Note: Descriptions are shown in the official language in which they were submitted.
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ANTI-GITR ANTIGEN-BINDING PROTEINS AND METHODS OF USE THEREOF
FIELD
[1] Provided herein are antigen-binding proteins (ABPs) with binding
specificity for
glucocorticoid-induced tumor necrosis factor receptor-related protein (GITR)
and
compositions comprising such ABPs, including pharmaceutical compositions,
diagnostic
compositions, and kits. Also provided are methods of making GITR ABPs, and
methods of
using GITR ABPs, for example, for therapeutic purposes, diagnostic purposes,
and
research purposes.
BACKGROUND
[2] GITR is a member of the tumor necrosis factor receptor (TNFR) superfamily.
GITR is
expressed in many cells of the innate and adaptive immune system, and membrane
surface
expression is increased in activated T cells. See Hanabuchib et al., Blood,
2006, 107:3617-
3623; and Nocentini et al., Eur. I Immunol., 2005, 35:1016-1022; each of which
is
incorporated by reference in its entirety. GITR is activated by GITR ligand
(GITRL).
[3] Agonism of GITR has a co-stimulatory effect on effector T cells. See
Schaer et al., Curr.
Op/n. Immunol., 2012, 24:217-224, incorporated by reference in its entirety.
GITR
agonists have been proposed as therapeutic agents for cancer therapy. See
Schaer et al.,
supra; Melero et al., Clin Cancer Res., 2013, 19:1044-1053; Cohen et al., I
Cl/n. Oncol.,
2007, 25:3058; Cohen et al., PLoS One, 2010, 5:e10436; Nocentini et al., Br.
Pharmacol., 2012, 165:2089-2099; and U.S. Pat. Pub. No. 2007/0098719; each of
which is
incorporated by reference in its entirety.
[4] Although antibody agonists of GITR have shown promise in mouse models, it
has been
difficult to obtain agonizing antibodies to human GITR. Nocentini et al. (Br.
Pharmacol., 2012, 165:2089-2099) have noted that lainti-GITR mAbs have much
weaker
triggering potential in humans than in mice." They speculate that this may be
a result of
the fact that human GITR must be multimerized into stable trimers or
superclusters (e.g.,
tetramers of trimers) in order to be robustly activated.
[5] Thus, there is a need for ABPs that can agonize human GITR more potently
than known
antibodies. Provided herein are ABPs that fulfill this need.
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SUMMARY
[6] Provided herein are ABPs that specifically bind GITR and methods of using
such ABPs.
[7] In one aspect is provided an isolated multivalent antigen binding protein
(ABP) that
specifically binds human GITR (hGITR; SEQ ID NO: 1), comprising the following
six
CDR sequences: (a) a CDR-H3 having the sequence XIX2X3X4X5RGYGDYGGHHAFDI,
wherein Xi is A or V, X2 is H, D, L, or R, X3 is E or D, X4 is R, N, S, or A,
and X5 is V, D
or G (SEQ ID NO:141); (b) a CDR-H2 having the sequence X1IX2X3SGX4TYYNPSLKS,
wherein Xi is G, L, or S, X2 is Y, A, or V, X3 is E, Y or H, and X4 is S or K
(SEQ ID
NO:142); (c) a CDR-H1 having the sequence XISISSX2X3X4X5WX6, wherein Xi is Y
or
G, X2 is G, S, or E, X3 is L, G, S, Y, or A, X4 is G, A, Y, M, or G, X5 is V,
A, or is absent,
and X6 is S or G (SEQ ID NO:143); (d) a CDR-L3 having the sequence
QQEYX1TPPX2,
wherein Xi is A or N and X2 is T or S (SEQ ID NO:144); (e) a CDR-L2 having the
sequence XIAX2SLX3X4, wherein Xi is A or S, X2 is D or S, X3 is Q, D, K, or E,
and X4 is
S or Y (SEQ ID NO:145); and (f) a CDR-L1 having the sequence XIAS X25I
X3X4YLN,
wherein Xi is G or R, X2 is Q or K, X3 is S, D, or N, and X4 is S or T (SEQ ID
NO:146).
[8] In one embodiment, the ABP of (a) comprises a VH sequence of SEQ ID NO:9
and a VL
sequence of SEQ ID NO:10; in another embodiment, the ABP of (b) comprises a
VII
sequence of SEQ ID NO:19 and a VL sequence of SEQ ID NO:20; in another
embodiment,
the ABP of (c) comprises a VII sequence of SEQ ID NO:26 and a VL sequence of
SEQ ID
NO:27; in another embodiment, the ABP of (d) comprises a VH sequence of SEQ ID
NO:26 or SEQ ID NO:34 and a VL sequence of SEQ ID NO:35; in another
embodiment,
the ABP of (e) comprises a VH sequence of SEQ ID NO:26 and a VL sequence of
SEQ ID
NO:40; in another embodiment, the ABP of (f) comprises a VH sequence of SEQ ID
NO:44
and a VL sequence of SEQ ID NO:45; in another embodiment, the ABP of (g)
comprises a
VH sequence of SEQ ID NO:44 and a VL sequence of SEQ ID NO:53; in another
embodiment, the ABP of (h) comprises a VII sequence of SEQ ID NO:58 and a VL
sequence of SEQ ID NO:10; in another embodiment, the ABP of (i) comprises a VH
sequence of SEQ ID NO:104 and a VL sequence of SEQ ID NO:10; and in another
embodiment, the ABP of (j) comprises a VII sequence of SEQ ID NO:105 and a VL
sequence of SEQ ID NO:10.
[9] In one embodiment, the ABP of (b) comprises a heavy chain of SEQ ID NO:7
and a light
chain of SEQ ID NO:8; In another embodiment, the ABP of (b) comprises a heavy
chain
of SEQ ID NO:17 and a light chain of SEQ ID NO:18. In another embodiment, the
ABP of
(c) comprises a heavy chain of SEQ ID NO:24 and a light chain of SEQ ID NO:25.
In
another embodiment, the ABP of (d) comprises (i) a heavy chain of SEQ ID NO:32
and a
light chain of SEQ ID NO:33, or (ii) a heavy chain of SEQ ID NO:37 and a light
chain of
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SEQ ID NO:33. In another embodiment, the ABP of (e) comprises (i) a heavy
chain of
SEQ ID NO:38 and a light chain of SEQ ID NO:39. In another embodiment, the ABP
of
(f) comprises a heavy chain of SEQ ID NO:42 and a light chain of SEQ ID NO:43;
in
another embodiment, the ABP of (g) comprises (i) a heavy chain of SEQ ID NO:51
and a
light chain of SEQ ID NO:52; In another embodiment, the ABP of (h) comprises
(i) a
heavy chain of SEQ ID NO:57 and a light chain of SEQ ID NO:8; In another
embodiment,
the ABP of (i) comprises (i) a heavy chain of SEQ ID NO:114 and a light chain
of SEQ ID
NO:8, or (ii) a heavy chain of SEQ ID NO:120 and a light chain of SEQ ID NO:8;
or (iii) a
heavy chain of SEQ ID NO:122 and a light chain of SEQ ID NO:8; or in another
embodiment, the ABP of (j) comprises (i) a heavy chain of SEQ ID NO:115 and a
light
chain of SEQ ID NO:8, or (ii) a heavy chain of SEQ ID NO:121 and a light chain
of SEQ
ID NO:8; or (iii) a heavy chain of SEQ ID NO:123 and a light chain of SEQ ID
NO:8.
[10] In another embodiment, the ABP comprises a heavy chain of SEQ ID NO:7
and a
light chain of SEQ ID NO:8; or a heavy chain of SEQ ID NO:17 and a light chain
of SEQ
ID NO:18; or a heavy chain of SEQ ID NO:24 and a light chain of SEQ ID NO:25;
a
heavy chain of SEQ ID NO:32 and a light chain of SEQ ID NO:33, or (ii) a heavy
chain of
SEQ ID NO:37 and a light chain of SEQ ID NO:33; a heavy chain of SEQ ID NO:38
and a
light chain of SEQ ID NO:39; a heavy chain of SEQ ID NO:42 and a light chain
of SEQ
ID NO:43; a heavy chain of SEQ ID NO:51 and a light chain of SEQ ID NO:52; a
heavy
chain of SEQ ID NO:57 and a light chain of SEQ ID NO:8; a heavy chain of SEQ
ID
NO:114 and a light chain of SEQ ID NO:8, or (ii) a heavy chain of SEQ ID
NO:120 and a
light chain of SEQ ID NO:8; or (iii) a heavy chain of SEQ ID NO:122 and a
light chain of
SEQ ID NO:8; or a heavy chain of SEQ ID NO:115 and a light chain of SEQ ID
NO:8, or
(ii) a heavy chain of SEQ ID NO:121 and a light chain of SEQ ID NO:8; or (iii)
a heavy
chain of SEQ ID NO:123 and a light chain of SEQ ID NO:8.
[11] In another aspect is provided an isolated multivalent antigen binding
protein (ABP)
that specifically binds human GITR (hGITR; SEQ ID NO: 1), comprising the
following
six CDR sequences: (a) a CDR-H3 having the sequence set forth in SEQ ID NO:66;
(b) a
CDR-H2 having the sequence set forth in SEQ ID NO:65; (c) a CDR-H1 having the
sequence set forth in SEQ ID NO:64; (d) a CDR-L3 having the sequence set forth
in SEQ
ID NO:69; (e) a CDR-L2 having the sequence set forth in SEQ ID NO:68; and (f)
a CDR-
Li having the sequence set forth in SEQ ID NO:67.
[12] In one embodiment, the ABP comprises a VII sequence of SEQ ID NO:62
and a VL
sequence of SEQ ID NO:63; a VH sequence of SEQ ID NO:70 and a VL sequence of
SEQ
ID NO: 63; or a VH sequence of SEQ ID NO: 97 and a VL sequence of SEQ ID NO
:63.
[13] In another embodiment, the ABP comprises (i) a heavy chain of SEQ ID
NO:171 and
a light chain of SEQ ID NO:172; a heavy chain of SEQ ID NO:173 and a light
chain of
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SEQ ID NO:174; a heavy chain sequence of SEQ ID NO:106 and a light chain
sequence of
SEQ ID NO:107; or (ii) a heavy chain sequence of SEQ ID NO:116 and a light
chain
sequence of SEQ ID NO:107.
[14] In another aspect is provided an isolated multivalent antigen binding
protein (ABP)
that specifically binds human GITR (hGITR; SEQ ID NO: 1), comprising the
following
six CDR sequences: (a) a CDR-H3 having the sequence set forth in SEQ ID NO:75;
(b) a
CDR-H2 having the sequence set forth in SEQ ID NO:74; (c) a CDR-H1 having the
sequence set forth in SEQ ID NO:73; (d) a CDR-L3 having the sequence set forth
in SEQ
ID NO:78; (e) a CDR-L2 having the sequence set forth in SEQ ID NO:77; and (f)
a CDR-
Li having the sequence set forth in SEQ ID NO:75.
[15] In one embodiment, the ABP comprises a VII sequence of SEQ ID NO:71
and a VL
sequence of SEQ ID NO:72; or a VH sequence of SEQ ID NO:98 and a VL sequence
of
SEQ ID NO:72.
[16] In another embodiment, the ABP comprises a heavy chain of SEQ ID
NO:173 and a
light chain of SEQ ID NO:109; or the ABP comprises (i) a heavy chain of SEQ ID
NO:108
and a light chain of SEQ ID NO:109; or (ii) a heavy chain of SEQ ID NO:117 and
a light
chain of SEQ ID NO:109.
[17] In another aspect is provided an isolated multivalent antigen binding
protein (ABP)
that specifically binds human GITR (hGITR; SEQ ID NO: 1), comprising the
following
six CDR sequences: (a) a CDR-H3 having the sequence set forth in SEQ ID NO:83;
(b) a
CDR-H2 having the sequence set forth in (i) SEQ ID NO:82, or (ii) SEQ ID
NO:100; (c) a
CDR-H1 having the sequence set forth in SEQ ID NO:81; (d) a CDR-L3 having the
sequence set forth in SEQ ID NO:86; (e) a CDR-L2 having the sequence set forth
in SEQ
ID NO:85; and (f) a CDR-L1 having the sequence set forth in SEQ ID NO:84.
[18] In one embodiment, the ABP comprises the CDR-H2 sequence of (b)(i) of
the above
paragraph and a VH sequence of SEQ ID NO:79 and a VL sequence of SEQ ID NO:80.
In
another embodiment, the ABP comprises the CDR-H2 sequence of (b)(i) of the
above
paragraph and a VH sequence of SEQ ID NO:87 and a VL sequence of SEQ ID NO:80.
In
another embodiment, the ABP comprises the CDR-H2 sequence of (b)(i) of the
above
paragraph and a VH sequence of SEQ ID NO:88 and a VL sequence of SEQ ID NO:80.
In
another embodiment, the ABP comprises the CDR-H2 sequence of (b)(ii) of the
above
paragraph and a VH sequence of SEQ ID NO:99 and a VL sequence of SEQ ID NO:80.
[19] In one embodiment, the ABP comprises a heavy chain of SEQ ID NO:174
and alight
chain of SEQ ID NO:111; In another embodiment, the ABP comprises a heavy chain
of
SEQ ID NO:175 and a light chain of SEQ ID NO: iii; in another embodiment, the
ABP
comprises a heavy chain of SEQ ID NO:176 and a light chain of SEQ ID NO: iii;
In
another embodiment, the ABP comprises a heavy chain of SEQ ID NO:110 and a
light
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chain of SEQ ID NO:111; or (ii) a heavy chain of SEQ ID NO:118 and a light
chain of
SEQ ID NO:111.
[20] In another aspect is provided an isolated multivalent antigen binding
protein (ABP)
that specifically binds human GITR (hGITR; SEQ ID NO:1), comprising the
following six
CDR sequences: (a) a CDR-H3 having the sequence set forth in SEQ ID NO:93; (b)
a
CDR-H2 having the sequence GIIPIFGEAQYAQX1FX2G, wherein Xi is K or R, and X2
is
Q or R (SEQ ID NO:215); (c) a CDR-H1 having the sequence set forth in SEQ ID
NO:91;
(d) a CDR-L3 having the sequence set forth in SEQ ID NO:94; (d) a CDR-L2
having the
sequence set forth in SEQ ID NO:85; and (e) a CDR-L1 having the sequence set
forth in
SEQ ID NO:84.
[21] In one embodiment, the ABP comprises: a CDR-H2 of SEQ ID NO:92; a CDR-
H2 of
SEQ ID NO:96; or a CDR-H2 of SEQ ID NO:102.
[22] In another embodiment, the ABP comprises a VII sequence of SEQ ID
NO:89 and a
VL sequence of SEQ ID NO:90. In another embodiment, the ABP comprises a VII
sequence of SEQ ID NO:95 and a VL sequence of SEQ ID NO:90. In another
embodiment,
the ABP comprises a VII sequence of SEQ ID NO:101 and a VL sequence of SEQ ID
NO:90.
[23] In another embodiment, the ABP comprises a heavy chain of SEQ ID
NO:177 and a
light chain of SEQ ID NO:113. In another embodiment, the ABP comprises a heavy
chain
of SEQ ID NO:178 and a light chain of SEQ ID NO:113. In another embodiment,
the ABP
comprises a heavy chain of SEQ ID NO:112 and a light chain of SEQ ID NO:113;
or (ii) a
heavy chain of SEQ ID NO:119 and a light chain of SEQ ID NO:113.
[24] In another aspect is provided an isolated multivalent antigen binding
protein (ABP)
that specifically binds human GITR (hGITR; SEQ ID NO: 1), comprising the
following
six CDR sequences: (a) a CDR-H3 having the sequence set forth in SEQ ID
NO:134; (b) a
CDR-H2 having the sequence set forth in SEQ ID NO:133;a CDR-H1 having the
sequence
set forth in SEQ ID NO:132; (c) a CDR-L3 having the sequence set forth in SEQ
ID
NO:135; (d) a CDR-L2 having the sequence set forth in SEQ ID NO:68; and (e) a
CDR-L1
having the sequence set forth in SEQ ID NO:67.
[25] In one embodiment, the ABP comprises (i) a VH sequence of SEQ ID
NO:126 and a
VL sequence of SEQ ID NO:128; or (ii) a VH sequence of SEQ ID NO:127 and a VL
sequence of SEQ ID NO:128.
[26] In one embodiment, the ABP comprises (i) a heavy chain of SEQ ID
NO:124 and a
light chain of SEQ ID NO:125; or (ii) a heavy chain of SEQ ID NO:136 and a
light chain
of SEQ ID NO:125.
[27] In another aspect is provided an isolated multivalent antigen binding
protein (ABP)
that specifically binds human GITR (hGITR; SEQ ID NO: 1), comprising: (a) a
CDR-H3
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having at least about 60%, at least about 70%, at least about 80%, at least
about 90%, or at
least about 95% identity to a CDR-H3 of a VH region selected from SEQ ID NOs:
9, 19,
26, 34, 44, 58, 62, 70, 71, 79, 87, 88, 89, 95, 97, 98, 99, 101, 104, 105,
126, and 127; (b) a
CDR-H2 having at least about 60%, at least about 70%, at least about 80%, at
least about
90%, or at least about 95% identity to a CDR-H2 of a VH region selected from
SEQ ID
NOs: 9, 19, 26, 34, 44, 58, 62, 70, 71, 79, 87, 88, 89, 95, 97, 98, 99, 101,
104, 105, 126,
and 127; (c) a CDR-H1 having at least about 60%, at least about 70%, at least
about 80%,
at least about 90%, or at least about 95% identity to a CDR-H1 of a VH region
selected
from SEQ ID NOs: 9, 19, 26, 34, 44, 58, 62, 70, 71, 79, 87, 88, 89, 95, 97,
98, 99, 101,
104, 105, 126, and 127; (d) a CDR-L3 having at least about 60%, at least about
70%, at
least about 80%, at least about 90%, or at least about 95% identity to a CDR-
L3 of a VL
region selected from SEQ ID NOs: 10, 20, 27, 35, 40, 45, 53, 63, 72, 80, 90,
and 128; (e) a
CDR-L2 having at least about 60%, at least about 70%, at least about 80%, at
least about
90%, or at least about 95% identity to a CDR-L2 of a VL region selected from
SEQ ID
NOs: 10, 20, 27, 35, 40, 45, 53, 63, 72, 80, 90, and 128; and (f) a CDR-L1
having at least
about 60%, at least about 70%, at least about 80%, at least about 90%, or at
least about
95% identity to a CDR-L1 of a VL region selected from SEQ ID NOs: 10, 20, 27,
35, 40,
45, 53, 63, 72, 80, 90, and 128.
[28] In one embodiment, the CDR-H3, CDR-H2, CDR-H1, CDR-L3, CDR-L2, and CDR-
Li are each identified according to a numbering scheme selected from the Kabat
numbering scheme, the Chothia numbering scheme, or the IMGT numbering scheme.
In
another embodiment, the CDR-H1 is identified as defined by both the Chothia
and Kabat
numbering schemes, inclusive of the boundaries of both numbering schemes.
[29] In one embodiment, the CDR-H3 comprises a CDR-H3 selected from SEQ ID
NOs:
13, 23, 30, 48, 61, 66, 75, 83, 93, 103, 131, or a variant thereof having 1,
2, or 3 amino
acid substitutions. In another embodiment, the CDR-H2 comprises a CDR-H3
selected
from SEQ ID NOs: 12, 22, 29, 47, 60, 65, 74, 82, 92, 96, 100, 102, 130, and
133, or a
variant thereof having 1, 2, or 3 amino acid substitutions. In another
embodiment, the
CDR-H1 comprises a CDR-H1 selected from SEQ ID NOs: 11, 21, 28, 46, 59, 64,
73, 81,
91, 129, 132, or a variant thereof having 1 or 2 amino acid substitutions. In
another
embodiment, the CDR-L3 comprises a CDR-L3 selected from SEQ ID NOs: 16, 17,
56,
69, 78, 86, 94, 135, or a variant thereof having 1 or 2 amino acid
substitutions. In another
embodiment, the CDR-L2 comprises a CDR-L2 selected from SEQ ID NOs: 15, 31,
41,
50, 55, 68, 77, and 85, or a variant thereof having 1 amino acid substitution.
In another
embodiment, the CDR-L1 comprises a CDR-L1 selected from SEQ ID NOs: 14, 36,
49,
54, 67, 76, and 84, or a variant thereof having 1 or 2 amino acid
substitutions. In one
embodiment, the amino acid substitutions are conservative amino acid
substitutions.
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[30] In an embodiment of any of the above aspects, the ABP: (a) competes
for binding to
GITR with an antibody selected from ABP1, ABP2, ABP3, ABP4, ABP5, ABP6, ABP7,
ABP8, ABP9, ABP10, ABP11, ABP12, ABP13, ABP14, ABP15, ABP16, ABP17,
ABP18, ABP19, ABP20, ABP21, ABP22, ABP23, ABP24, ABP25, ABP26, ABP27,
ABP28, ABP29, ABP30, ABP31, ABP32, ABP33, and ABP34, each as provided in
Appendix A of this disclosure; or (b) has at least three antigen-binding
domains that
specifically bind an epitope on GITR; or (c) has at least three antigen-
binding domains that
specifically bind a single epitope on GITR; or (d) has at least four antigen-
binding
domains that specifically bind an epitope on GITR; or (e) has at least four
antigen-binding
domains that specifically bind a single epitope on GITR; or (f) agonizes GITR
expressed
on the surface of a target cell; or (g) enhances the binding of GITRL to GITR;
or (h) co-
stimulates an effector T cell in combination with antigen presentation from an
antigen-
presenting cell; or (i) inhibits the suppression of an effector T cell by a
regulatory T cell; or
(j) reduces the number of regulatory T cells in a tissue or in systemic
circulation; (k) is
capable of binding to one or more of GITR (SEQ ID NO:1) residues from the
group
consisting of R56, C58, R59, D60, Y61, P62, E64, E65, C66, and C67; or (1) is
capable of
any combination of (a) - (k).
[31] In an embodiment of any of the above aspects, the GITR is selected
from hGITR
(SEQ ID NO: 1), hGITR-T43R (SEQ ID NO: 2), cGITR (SEQ ID NO: 3), mGITR (SEQ
ID NO: 4), and combinations thereof In another embodiment, the ABP (a)
specifically
binds cynomolgus monkey GITR (cGITR; SEQ ID NO: 3); (b) binds murine GITR
(mGITR; SEQ ID NO: 4) with an affinity lower (as indicated by higher KD) than
the
affinity of the ABP for hGITR, or does not bind mGITR; or (c) is capable of
any
combination of (a) - (b). In another embodiment, the ABP: (a) specifically
binds cGITR
(SEQ ID NO: 3); (b) binds mGITR (SEQ ID NO: 4) with an affinity lower (as
indicated by
higher KD) than the affinity of the ABP for hGITR and cGITR; and (c) enhances
binding
of GITRL to GITR.
[32] In another aspect is provided an ABP that competes for binding to GITR
with an ABP
of any one of claims 1-26, wherein the ABP: (a) specifically binds cGITR (SEQ
ID NO:
3); (b) binds mGITR (SEQ ID NO: 4) with an affinity lower (as indicated by
higher KD)
than the affinity of the ABP for hGITR and cGITR; and (c) enhances binding of
GITRL to
GITR. In one embodiment, the ABP comprises an antibody. In another embodiment,
the
antibody is a monoclonal antibody. In another embodiment, the antibody is
selected from a
human antibody, a humanized antibody or a chimeric antibody. In another
embodiment,
the ABP is multivalent. In another embodiment, the ABP comprises an antibody
fragment.
[33] In another embodiment, the ABP comprises an alternative scaffold. In
another
embodiment, the ABP comprises an immunoglobulin constant region. In another
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embodiment, the ABP comprises heavy chain constant region of a class selected
from IgA,
IgD, IgE, IgG, or IgM. In another embodiment, the ABP comprises a heavy chain
constant
region of the class IgG and a subclass selected from IgG4, IgGl, IgG2, or
IgG3.
[34] In an embodiment of any of the above aspects, at least one Fab is
fused to a C-
terminus of an Fc domain of an IgG. In another embodiment, the ABP further
comprises at
least one linker. In another embodiment, the IgG is an IgG4. In another
embodiment, the
IgG is an IgGl. In another embodiment, at least one Fab is fused to an N-
terminus of an Fc
domain of an IgG. In one embodiment, the at least one Fab is at least two
Fabs. In another
embodiment, the at least one Fab is at least three Fabs. In another
embodiment, the at least
one Fab is at least four Fabs. In another embodiment, two Fabs are
independently fused to
an N-terminus of the IgG. In another embodiment, two Fabs are independently
fused to a
C-terminus of the IgG. In another embodiment, a Fab is attached to each N-
terminus of the
IgG, a linker is attached to each said Fab, and a Fab is attached to each
linker. In another
embodiment, a Fab is attached to each C-terminus of the IgG, a linker is
attached to each
said Fab, and a Fab is attached to each linker. In another embodiment, wherein
each linker
comprises SEQ ID NO:5. In another embodiment, each linker comprises SEQ ID
NO:6.
[35] In an embodiment of any of the above aspects, the ABP comprises a
common light
chain antibody, an antibody with a knobs-into-holes modification, an scFy
attached to an
IgG, a Fab attached to an IgG, a diabody, a tetravalent bispecific antibody, a
DVD-Iirm, a
DART, a DuoBody0, a CovX-Body, an Fcab antibody, a TandAbO, a tandem Fab, a
ZybodyTm, or combinations thereof In one embodiment, the ABP binds more than
one
GITR molecule. In another embodiment, the ABP is independent of GITRL binding.
In
another embodiment, the ABP enhances binding of GITRL to GITR by at least
about 10%,
at least about 20%, at least about 30%, at least about 40%, at least about
50%, at least
about 60%, at least about 70%, at least about 80%, or at least about 90%. In
another
embodiment, the ABP enhances binding of GITRL to GITR by at least about 50%.
In
another embodiment, the target cell is selected from an effector T cell, a
regulatory T cell,
a natural killer (NK) cell, a natural killer T (NKT) cell, a dendritic cell,
and a B cell. In
another embodiment, the target cell is an effector T cell selected from a
helper (CD4+) T
cell, a cytotoxic (CD8+) T cell, and combinations thereof In another
embodiment, the
target cell is a regulatory T cell selected from a CD4+CD25+Foxp3+ regulatory
T cell, a
CD8+CD25+ regulatory T cell, and combinations thereof. In another embodiment,
the
tissue is a tumor.
[36] In an embodiment of any of the above aspects, the KD of the first
antigen-binding
domain for hGITR (SEQ ID NO: 1) or hGITR-T43R (SEQ ID NO: 2) is less than
about 20
nM. In one embodiment, the KD of the first antigen-binding domain for cGITR
(SEQ ID
NO: 3) is less than about 200 nM. In another embodiment, the KD of the second
antigen-
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binding domain for hGITR (SEQ ID NO: 1) or hGITR-T43R (SEQ ID NO: 2) is less
than
about 100 nM. In another embodiment, the KD of the second antigen-binding
domain for
cGITR (SEQ ID NO: 3) is less than about 1 [IM. In another embodiment, the ABP
comprises an Fc domain with reduced effector function when compared to an IgG1
Fc
domain. In another embodiment, the ABP comprises an aglycosylated Fc domain.
In
another embodiment, the ABP comprises an IgG1 Fc domain with an alanine at one
or
more of positions 234, 235, 265, and 297.
[37] In an embodiment of any of the above aspects, the GITR is expressed on
the surface
of a target cell. In one embodiment, the ABP multimerizes GITR expressed on
the surface
of a target cell. In one embodiment, the ABP multimerizes 2, 3, 4, 5, 6, 7, 8,
9, 10, 11, or
12 GITR molecules.
[38] In an embodiment of any of the above aspects, the ABP specifically
binds an epitope
of human GITR (hGITR; SEQ ID NO:1) and is capable of binding one or more
residues
from the group consisting of R56, C58, R59, D60, Y61, P62, E64, E65, C66, and
C67.
[39] In an embodiment of any of the above aspects, the ABP comprises an
immunoglobulin comprising at least two different (i.e., have different
sequences and/or
bind to different residues) heavy chain variable regions each paired with a
common light
chain variable region. In another embodiment, the common light chain variable
region
forms a distinct antigen-binding domain with each of the two different heavy
chain
variable regions. In another embodiment, the ABP comprises a first VH variable
domain
having SEQ ID NO:189, a second VH variable domain having SEQ ID NO:215, and a
common variable light chain having SEQ ID NO:190. In another embodiment, the
ABP
comprises a first VH variable domain having SEQ ID NO:199, a second VH
variable
domain having SEQ ID NO:216, and a common variable light chain having SEQ ID
NO :200.
[40] Provided in another aspect is a kit comprising an ABP of any of the
above aspects or
set forth in Appendix A, and instructions for use of the ABP. In one
embodiment, the ABP
is lyophilized. In another embodiment, the kit further comprises a fluid for
reconstitution
of the lyophilized ABP.
[41] It is known that when an antibody is expressed in cells, the antibody
is modified after
translation. Examples of the posttranslational modification include cleavage
of lysine at
the C terminal of the heavy chain by a carboxypeptidase; modification of
glutamine or
glutamic acid at the N terminal of the heavy chain and the light chain to
pyroglutamic acid
by pyroglutamylation; glycosylation; oxidation; deamidation; and glycation,
and it is
known that such posttranslational modifications occur in various antibodies
(See journal of
Pharmaceutical Sciences, 2008, Vol. 97, p. 2426-2447, incorporated by
reference in its
entirety). In some embodiments, an ABP of the invention is an antibody or
antigen-binding
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fragment thereof which has undergone posttranslational modification. Examples
of an
antibody or antigen binding fragment thereof which have undergone
posttranslational
modification include an antibody or antigen-binding fragments thereof which
have
undergone pyroglutamylation at the N terminal of the heavy chain variable
region and/or
deletion of lysine at the C terminal of the heavy chain. It is known in the
art that such
posttranslational modification due to pyroglutamylation at the terminal and
deletion of
lysine at the C terminal does not have any influence on the activity of the
antibody or
fragment thereof (Analytical Biochemistry, 2006, Vol. 348, p. 24-39,
incorporated by
reference in its entirety).
[42] In an embodiment of any of the above aspects, the ABP comprises a
polypeptide
sequence having a pyroglutamate (pE) residue at its N-terminus. In one
embodiment, the
ABP comprises a VH sequence in which an N-terminal Q is substituted with pE.
In
another embodiment, the ABP comprises a VL sequence in which an N-terminal E
is
substituted with pE. In another embodiment, the ABP comprises a heavy chain
sequence
in which an N-terminal Q is substituted with pE.
[43] In an embodiment of any of the above aspects, the ABP comprises a
light chain
sequence in which an N-terminal E is substituted with pE. In another
embodiment, the
ABP is for use as a medicament. In another embodiment, the ABP is for use in
the
treatment of a cancer or viral infection.
[44] In one embodiment, the ABP is for use in the treatment of a cancer,
wherein the
cancer is selected from a solid tumor and a hematological tumor.
[45] In another aspect is provided an isolated polynucleotide encoding an
ABP of any of
the above aspects or set forth in Appendix A, a VH thereof, a VL thereof, a
light chain
thereof, a heavy chain thereof or an antigen-binding portion thereof
[46] In another aspect is provided a vector comprising the polynucleotide
of the above
aspect. In another aspect is provided a host cell comprising the
polynucleotide or the
vector of any of the above aspects. In one embodiment, the host cell is
selected from a
bacterial cell, a fungal cell, and a mammalian cell. In another embodiment,
the host cell is
selected from an E. coli cell, a Saccharomyces cerevisiae cell, and a CHO
cell.
[47] In another aspect is provided a cell-free expression reaction
comprising the
polynucleotide or vector of the above aspects.
[48] In another aspect is provided a method of producing an ABP of any of
the above
aspects or set forth in Appendix A, comprising expressing the ABP in the host
cell and
isolating the expressed ABP.
[49] In another aspect is provided a pharmaceutical composition comprising
an ABP of
any of the above aspects or set forth in Appendix A, and a pharmaceutically
acceptable
excipient. In one embodiment, the amount of the ABP in the pharmaceutical
composition
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is sufficient to (a) reduce the suppression of effector T cells by regulatory
T cells; (b)
activate effector T cells; (c) reduce the number of regulatory T cells in a
tissue or
systemically; (d) induce or enhance proliferation of effector T cells; (e)
inhibit the rate of
tumor growth; (0 induce tumor regression; or (g) combinations thereof, in a
subject. In one
embodiment, the pharmaceutical composition is for use as a medicament, e.g.,
for use in
the treatment of a cancer or a viral infection. In one embodiment, the
pharmaceutical
composition is for use in the treatment of a cancer, wherein the cancer is
selected from a
solid tumor and a hematological tumor.
[50] In another embodiment, the amount of the ABP in the pharmaceutical
composition is
sufficient to (a) reduce the suppression of effector T cells by regulatory T
cells; (b) activate
effector T cells; (c) reduce the number of regulatory T cells in a tissue or
systemically; (d)
induce or enhance proliferation of effector T cells; (e) inhibit the rate of
tumor growth; (f)
induce tumor regression; or (g) combinations thereof, in a subject.
[51] In another aspect is provided a method of treating or preventing a
disease or condition
in a subject in need thereof, comprising administering to the subject an
effective amount of
an ABP of any of the above aspects or set forth in Appendix A, or a
pharmaceutical
composition thereof
[52] In another aspect is provided a method of method of increasing
activation of immune
cells in a subject, comprising administering to the subject an effective
amount of an ABP
of any of the above aspects or set forth in Appendix A, or a pharmaceutical
composition
thereof In one embodiment, disease or condition is a cancer.
[53] In another embodiment, the method induces or enhances an immune
response to a
cancer-associated antigen. In another embodiment, the ABP is administered in
an amount
sufficient to (a) reduce the suppression of effector T cells by regulatory T
cells; (b) activate
effector T cells; (c) reduce the number of regulatory T cells in a tissue or
systemically; (d)
induce or enhance proliferation of effector T cells; (e) inhibit the rate of
tumor growth; (f)
induce tumor regression; or (g) combinations thereof In another embodiment,
the cancer is
a solid cancer. In another embodiment, the cancer is a hematological cancer.
In another
embodiment, the method further comprises administering one or more additional
therapeutic agents. In one embodiment, the additional therapeutic agent is
selected from
radiation, a cytotoxic agent, a chemotherapeutic agent, a cytostatic agent, an
anti-hormonal
agent, an EGFR inhibitor, an immunostimulatory agent, an anti-angiogenic
agent, and
combinations thereof In one embodiment, the additional therapeutic agent is an
immunostimulatory agent. In one embodiment, the immunostimulatory agent
comprises
an agent that blocks signaling of an inhibitory receptor expressed by an
immune cell or a
ligand thereof In one embodiment, the inhibitory receptor expressed by an
immune cell or
ligand thereof is selected from CTLA-4, PD-1, PD-L1, NRP1, LAG-3, Tim3, TIGIT,
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neuritin, BTLA, KIR, and combinations thereof In one embodiment, the
immunostimulatory agent comprises an agonist to a stimulatory receptor
expressed by an
immune cell. In another embodiment, the stimulatory receptor expressed by an
immune
cell is selected from 0X40, ICOS, CD27, CD28, 4-1BB, CD40, and combinations
thereof
In another embodiment, the immunostimulatory agent comprises a cytokine. In
another
embodiment, the cytokine is selected from IL-2, IL-5, IL-7, IL-12, IL-15, IL-
21, and
combinations thereof In another embodiment, the immunostimulatory agent
comprises an
oncolytic virus. In another embodiment, the oncolytic virus is selected from
herpes
simplex virus, vesicular stomatitis virus, adenovirus, Newcastle disease
virus, vaccinia
virus, a maraba virus, and combinations thereof. In another embodiment, the
immunostimulatory agent comprises a T cell expressing a chimeric antigen
receptor. In
another embodiment, the immunostimulatory agent comprises a bi- or multi-
specific T cell
directed antibody. In another embodiment, the immunostimulatory agent
comprises an
anti-TGF-B antibody, a TGF-B trap, or a combination thereof In another
embodiment, the
immunostimulatory agent comprises a vaccine to a cancer-associated antigen.
[54] In another aspect is provided a method of modulating an immune
response in a
subject in need thereof, comprising administering to the subject an effective
amount of an
ABP of any of the above aspects or set forth in Appendix A, or a
pharmaceutical
composition thereof In one embodiment, the method further comprises
administering one
or more additional therapeutic agents to the subject. In one embodiment, the
additional
therapeutic agent is an agonist to a stimulatory receptor of an immune cell,
and the
stimulatory receptor of an immune cell is selected from 0X40, CD2, CD27, CDS,
ICAM-
1, LFA-1 (CD11a/CD18), ICOS (CD278), 4-1BB (CD137), CD28, CD30, CD40, BAFFR,
HVEM, CD7, LIGHT, NKG2C, SLAMF7, NKp80, CD160, B7-H3, CD83 ligand, and
combinations thereof. In one embodiment, the additional therapeutic agent is a
cytokine
selected from IL-2, IL-5, IL-7, IL-12, IL-15, IL-21, and combinations thereof
In one
embodiment, the additional therapeutic agent is an oncolytic virus selected
from herpes
simplex virus, vesicular stomatitis virus, adenovirus, Newcastle disease
virus, vaccinia
virus, a maraba virus, and combinations thereof. In one embodiment, the
additional
therapeutic agent is formulated in the same pharmaceutical composition as the
ABP. In
one embodiment, the additional therapeutic agent is formulated in a different
pharmaceutical composition from the ABP. In one embodiment, the additional
therapeutic
agent is administered prior to administering the ABP. In one embodiment, the
additional
therapeutic agent is administered after administering the ABP. In one
embodiment, the
additional therapeutic agent is administered contemporaneously with the ABP.
[55] In another aspect is provided an isolated multivalent antigen binding
protein (ABP)
that specifically binds human GITR (hGITR; SEQ ID NO: 1), wherein the ABP
competes
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for binding with one or more of ABP1, ABP2, ABP3, ABP4, ABP5, ABP6, ABP7,
ABP8,
ABP9, ABP10, ABP11, ABP12, ABP13, ABP14, ABP15, ABP16, ABP17, ABP18,
ABP19, ABP20, ABP21, ABP22, ABP23, ABP24, ABP25, ABP26, ABP27, ABP28,
ABP29, ABP30, ABP31, ABP32, ABP33, and ABP34, ABP50, ABP51, ABP52, ABP53,
ABP54, ABP55, ABP56, ABP57, ABP62, ABP63, ABP64, ABP65, ABP66, ABP67,
ABP68, ABP67, ABP68, ABP69, ABP70, ABP71, ABP72, ABP73, ABP74, ABP75,
ABP76, ABP77, ABP78, ABP79, ABP80, ABP812, ABP82, ABP83, ABP84, ABP85,
ABP86, ABP87, ABP88, ABP89, ABP90, ABP91, ABP92, ABP93, ABP94, ABP95,
ABP96, ABP97, ABP98, ABP99, ABP100, ABP102, ABP103, ABP104, ABP105,
ABP106, ABP107, ABP108, and ABP109, each as provided in Appendix A of this
disclosure.
1561 In another aspect is provided an anti-human GITR antibody or an
antigen-binding
fragment thereof comprising four heavy chain variable regions and four light
chain
variable regions, wherein the heavy chain variable region comprises a CDR-H3
consisting
of SEQ ID NO:13, a CDR-H2 consisting of SEQ ID NO:12 and a CDR-H1 consisting
of
SEQ ID NO:11; and the light chain variable region comprises a CDR-L3
consisting of
SEQ ID NO:16, a CDR-L2 consisting of SEQ ID NO:15, and a CDR-L1 consisting of
SEQ ID NO:14; and the one heavy chain variable region and the one light chain
variable
region constitute one antigen-binding site, and the antibody or the antigen-
binding
fragment comprises four antigen-binding sites.
[57] In one embodiment, the anti-human GITR antibody or the antigen-binding
fragment
thereof comprises four heavy chain variable regions and four light chain
variable regions,
in which the heavy chain variable region consists of SEQ ID NO: 9, the light
chain
variable region consists of SEQ ID NO: 10, and the one heavy chain variable
region and
the one light chain variable region constitute one antigen-binding site, and
the antibody or
the antigen-binding fragment comprises four antigen-binding sites.
[58] In one embodiment, the anti-human GITR antibody or the antigen-binding
fragment
thereof comprises four heavy chain variable regions and four light chain
variable regions,
in which the heavy chain variable region consists of SEQ ID NO: 9, wherein Q
at position
1 of the sequence is modified to pyroglutamate, the light chain variable
region consists of
SEQ ID NO: 10, and the one heavy chain variable region and the one light chain
variable
region constitute one antigen-binding site, and the antibody or the antigen-
binding
fragment comprises four antigen-binding sites.
[59] In one embodiment, the anti-human GITR antibody comprises two heavy
chains and
four light chains, each heavy chain comprises two structures consisting of a
heavy chain
variable region and a CH1 region, a CH2 region, and a CH3 region, and the C
terminus of
one of the structures is linked to the N terminus of the other structure
through a linker, and
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each light chain comprises a light chain variable region and a light chain
constant region
(left panel in FIG. 1B).
[60] In one embodiment, the anti-human GITR antibody comprises two heavy
chains and
four light chains, the heavy chains each comprising a first heavy chain
variable region and
a first CH1 region, a linker, a second heavy chain variable region, a second
CH1 region, a
CH2 region, and a CH3 region, and each light chain comprises a light chain
variable
region and a light chain constant region (left panel in FIG. 1B).
[61] In one embodiment, the anti-human GITR antibody comprises two heavy
chains and
four light chains; each heavy chain comprises two structures consisting of a
heavy chain
variable region comprising a CDR-H3 consisting of SEQ ID NO:13, a CDR-H2
consisting
of SEQ ID NO:12 and a CDR-H1 consisting of SEQ ID NO:11 and a CH1 region, a
CH2
region, and a CH3 region, and the carboxy terminus (C terminus) of one of the
structures is
linked to the amino terminus (N terminus) of the other structure through a
linker; and each
light chain comprises a light chain variable region comprising a CDR-L3
consisting of
SEQ ID NO:16, a CDR-L2 consisting of SEQ ID NO:15, and a CDR-L1 consisting of
SEQ ID NO:14.
[62] In one embodiment, the anti-human GITR antibody comprises two heavy
chains and
four light chains; each heavy chain comprising a first heavy chain variable
region and a
second heavy chain variable region, each comprising a CDR-H3 consisting of SEQ
ID
NO:13, a CDR-H2 consisting of SEQ ID NO:12 and a CDR-H1 consisting of SEQ ID
NO:11; a first CH1 region, a linker, a second CH1 region, a CH2 region, and a
CH3
region; and each light chain comprises a light chain variable region
comprising a CDR-L3
consisting of SEQ ID NO:16, a CDR-L2 consisting of SEQ ID NO:15, and a CDR-L1
consisting of SEQ ID NO:14.
[63] In one embodiment, the anti-human GITR antibody comprising two heavy
chains and
four light chains, in which each heavy chain comprises two structures
consisting of a
heavy chain variable region of SEQ ID NO: 9 and a CH1 region, a CH2 region,
and a CH3
region, and the C terminus of one of the structures is linked to the N
terminus of the other
structure through a linker, and each light chain comprises a light chain
variable region of
SEQ ID NO: 10, and a light chain constant region.
[64] In one embodiment, the anti-human GITR antibody comprises two heavy
chains and
four light chains, each heavy chain comprising a first heavy chain variable
region and a
second heavy chain variable region, each as set forth as SEQ ID NO: 9; a first
CH1 region,
a linker, a second CH1 region, a CH2 region, and a CH3 region; and wherein
each light
chain comprises a light chain variable region of SEQ ID NO: 10, and a light
chain constant
region.
[65] In one embodiment, the anti-human GITR antibody comprising two heavy
chains and
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four light chains, in which each heavy chain comprises two structures
consisting of a
heavy chain variable region of SEQ ID NO: 9 and a CH1 region, a CH2 region,
and a CH3
region, and the C terminus of one of the structures is linked to the N
terminus of the other
structure through a linker, wherein Q at position 1 of the sequence is
modified to
pyroglutamate, and each light chain comprises a light chain variable region of
SEQ ID
NO: 10, and a light chain constant region.
[66] In one embodiment, the anti-human GITR antibody comprises two heavy
chains and
four light chains, each heavy chain comprising a first heavy chain variable
region and a
second heavy chain variable region, each as set forth as SEQ ID NO: 9; a first
CH1 region,
a linker, a second CH1 region, a CH2 region, and a CH3 region; wherein Q at
position 1 of
the sequence is modified to pyroglutamate, and each light chain comprises a
light chain
variable region of SEQ ID NO: 10, and a light chain constant region.
[67] In one embodiment, the anti-human GITR antibody comprises two heavy
chains, each
having two variable regions, of SEQ ID NO: 7 and four light chains of SEQ ID
NO: 8.
[68] In one embodiment, the anti-human GITR antibody comprises two heavy
chains, each
having two variable regions, of SEQ ID NO: 7, wherein the Q at position 1 of
the sequence
is modified to pyroglutamate and four light chains of SEQ ID NO: 8.
[69] In one embodiment, the anti-human GITR antibody comprises two heavy
chains, each
having two variable regions, consisting of the amino acid sequence ranging
from Q at
position 1 to G at position 686 of SEQ ID NO: 7, and four light chains of SEQ
ID NO: 8.
[70] In one embodiment, the anti-human GITR antibody comprises two heavy
chains
consisting of the amino acid sequence ranging from Q at position 1 to G at
position 686 of
SEQ ID NO: 7, wherein the Q is modified to pyroglutamate, and four light
chains of SEQ
ID NO: 8.
BRIEF DESCRIPTION OF THE DRAWINGS
[71] FIG. 1 provides a schematic illustration of a mechanism of action of
certain
illustrative GITR ABPs provided herein. FIG. lA shows three GITR molecules
(one
labeled 101) embedded in a cell membrane (102). A cartoon of a traditional
format
antibody is shown binding just two GITR molecules (103). FIG. 1B shows a
cartoon of
the tetravalent monospecific (TM) format antibodies disclosed herein: the left
panel (105)
shows a TM comprising two N-terminal IgG1 Fabs, with a C-terminal IgG4 5228P
antibody; the right panel (106) shows a TM comprising two C-terminal IgG1 Fabs
and an
N-terminal IgG4 5228P antibody. The antigen binding domains are illustrated by
open
circles (107). FIG. 1C shows a cartoon of the tetravalent bispecific format
antibodies
disclosed herein: the left panel (105) shows a bispecific antibody comprising
two N-
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terminal IgG1 Fabs, with a C-terminal IgG4 S228P antibody; the right panel
(106) shows a
bispecific antibody comprising two C-terminal IgG1 Fabs and an N-terminal IgG4
S228P
antibody. The antigen binding domains having specificity for two non-
overlapping
epitope specificities are illustrated by open circles (107 and 108). FIG. 1D
shows
multimerization of GITR following binding of three illustrative tetravalent
monospecific
(TM) format ABPs. Such multimerization is expected to agonize GITR signaling,
as
described elsewhere in this disclosure.
[72] FIG. 2 is a graph showing exemplary results of determination of KD by
OCTET for
N-terminal Fab format ABPs 1-8.
[73] FIG. 3 is a series of graphs showing the results of FACS analysis
demonstrating
binding of an exemplary panel of N-Terminal Fab TM format ABPs to CD4+ (Figure
3A)
and CD8+ (Figure 3B) T cells. The distribution of CD4+ and CD8+ is shown in
the top
left panel of each Figure. In the top row, from left to right, is shown an
anti-GITR positive
control, and anti-human IgG4 isotype control, ABP9, ABP2, ABP1, and ABP7. On
the
bottom row is shown ABP8, ABP3, ABP4, ABP5, ABP6, ABP23, and ABP24. The
percentage of positively stained IgG4+ CD4/8+ cells is indicated; MRI is
indicated in
brackets..
[74] FIG. 4 is a series of graphs showing the activity of eight optimized
agonist antibodies
in the N-terminal Fab TM format. HT1080 cells that stably express human (left
panels) or
cynomolgus monkey (right panels) GITR were then incubated with ABP1 (FIG. 4A),
ABP2 (FIG. 4B), ABP3 (FIG. 4C), ABP4 (FIG. 4D), ABP5 (FIG. 4E), ABP6 (FIG.
4F),
ABP7 (FIG. 4G), and ABP8 (FIG 4H) (shown as circles in the FIGs), and IL-8
induction
was measured. GITRL was used as a control (squares). A table of EC50 values is
shown
on the bottom of each panel of each FIG.
[75] FIG. 5 is a series of graphs comparing the agonist activity in GITR-
expressing
HT1080 cells of a parental N-terminal Fab TM format antibody, ABP43, to a
number of
further optimized antibodies having either N-terminal or C-terminal format.
FIG. 5A
shows ABP43 (squares), ABP23 (circles), ABP24 (triangles), and ABP29 (open
circles),
ABP30 (open triangles), ABP31 (open circles), and ABP32 (open triangles), all
in
comparison to GITRL (+ sign). FIG. 5B shows ABP19 (N-terminal Fab, triangles)
and
ABP25 (C terminal Fab, upside down triangles). GITRL is shown as plus signs.
FIG. 5C
shows ABP21 (N-terminal Fab, triangles) and ABP27 (C terminal Fab, upside down
triangles). GITRL is shown as plus signs. FIG. 5D shows ABP20 (N-terminal Fab,
triangles) and ABP26 (C terminal Fab, upside down triangles). GITRL is shown
as plus
signs. FIG. 5E shows ABP22 (N-terminal Fab, triangles) and ABP28 (C terminal
Fab,
upside down triangles). GITRL is shown as plus signs. IgG4 control is shown as
X sign
in each figure.
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[76] FIG. 6 shows the results of EC50 determination for ABP33 and ABP34 in
the
HT1080 assay as described in the Examples. ABP33 (tetravalent version
combining the
IgG4 ABP61 with the IgG1 Fab of ABP58 on the N-terminus) and ABP34
(tetravalent
version combining the IgG4 of ABP61 with the IgG1 Fab of ABP58 on the C-
terminus)
were compared to the bivalent ABPs 59 and 61 (IgG4 S228P), which were the
basis for
ABP33 and ABP34. As shown in the FIG., the bispecific tetravalent antibodies
both had
superior EC50, as measured by IL-8 induction, when compared to the individual
bivalent
antibodies used to construct the bispecific tetravalent antibodies.
[77] FIG. 7 shows the results of EC50 determination in a Jurkat T cell
assay as described
for the HT1080 cells above. Optimized N terminal Fab TM format ABPs were
compared
to GITRL for their ability to agonize GITR, as measured by IL-8 production.
FIG. 7A-7H
show ABPs 1-8, respectively.
[78] FIG. 8A shows FACS analysis from triplicate quantifications of T cells
isolated from
two human donors, and shows the percentage of GITR+ CD4+ cells (left) and CD8+
cells
(right) at various time points, +/- stimulation with PHA.
[79] FIGs 8B- 8M show results of treatment of T-blasts from 4 different
donors treated
with controls or ABPs and the resultant IL-2 production (data normalized to
levels of IL-2
production achieved in control media). In each FIG. the top row, from left to
right is FACS
measurement of the ABP binding to CD4+ cells, IL-2 production in cells from
Donor 1,
IL-2 production in cells from Donor 2. In the bottom row, from left to right,
is FACS
measurement of the ABP binding to CD8+ cells, IL-2 production in cells from
Donor 3,
IL-2 production in cells from Donor 4. Shown are data as follows: 8B: IgG4
isotype
control; 8C: SEC4 antibody; 8D: IgG4 TM format negative control; 8E: ABP9 (TM
Format IgG4 non-optimized parent of ABPs 1-8). 8F: ABP1; 8G: ABP2; 8H: ABP3;
81:
ABP4; 8J: ABP5; 8K:ABP6; 8L: ABP7; 8M: ABP8.
[80] FIG. 9A-9H is a series of graphs showing a comparison of the activity
of the
optimized N-terminal Fab TM ABPs ("IgG4 TM") against the corresponding
optimized
non-TM IgG1 N297A ABPs ("IgGl"). HT1080 cells that stably express human (left
panel)
or cynomolgus (right panel) were then treated with each N-terminal Fab TM ABP
and
corresponding IgG1 ABP as follows: ABP1 IgG4 TM /ABP35 IgG1 (FIG. 9A), ABP2
IgG4 TM /ABP36 IgG1 (FIG. 9B), ABP3 IgG4 TM /ABP37 IgG1 (FIG. 9C), ABP4 IgG4
TM /ABP38 IgG1 (FIG. 9D), ABP5 IgG4 TM /ABP38 P45L IgG1 (FIG. 9E), ABP6 IgG4
TM /ABP39 IgG1 (FIG. 9F), ABP7 IgG4 TM /ABP40 IgG1 (FIG. 9G) and ABP8 IgG4
TM /ABP41 IgG1 (FIG. 9H) and IgG4 as a positive control. Induction of IL-8 by
GITRL
is shown as circles, by IgG4 TM format ABPs as triangles, by the corresponding
IgG1
ABPs as diamonds, and by IgG4 control antibodies as squares. A table of EC50
values is
shown on the bottom of each panel of each FIG.
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[81] FIG. 10 is a graph showing EC50data in the HT1080 assay as described
comparing a
non-TM parental antibody with corresponding TM format versions. ABP43
(diamonds) is
the non-TM IgG4 S228P parent of ABP9 (IgG4 N-terminal Fab, squares) and ABP10
(IgG4 C-terminal Fab, circles). IL-8 induction by GITRL (positive control) is
shown as a
single data point (star). IL-8 production is shown in pg/mL.
[82] FIG. 11A is a graph showing induction of IL-8 by representative
benchmark
antibodies SEC4 and SEC9 in HT1080 cells that were engineered to stably
express human
GITR. Cultured cells were treated for six hours with a range of concentrations
of two
benchmark agonist antibodies SEC4 (35E6 formatted to have mouse variable
regions with
human IgG4 S228P/kappa regions, diamonds) and SEC9 (humanized 6C8 N62Q IgG1
N297A, circles), as well as an IgG4 negative control (closed triangles), an
IgG1 negative
control (open triangles), and trimeric human GITR ligand ("hGITRL", squares)
as a
positive control. Induction of IL-8 was measured by ELISA. As shown in the
Figure,
GITRL had a better EC50 (inset) and max induction compared to both SEC4 and
SEC9.
FIGs 11B-11I show comparison of ABPs 1-8, respectively, with SEC4 and SEC9. TM
ABPs are indicated with circles, SEC4 with squares and SEC9 with triangles. IL-
8
production is shown in pg/mL.
[83] FIG. 12 is two graphs showing cytokine production in stimulated
isolated human
NSCLC adenocarcinoma cells after treatment with TM-format ABP1 alone or in
combination with pembrolizumab. Cells were either unstimulated controls, or
were
stimulated with lug/mL aCD3 (Soluble) +2 ug/mL aCD28 (Soluble) + IL-2
(50ng/mL);
cells either received no immunotherapy treatment (for assessment of checkpoint
protein
levels), pembrolizumab (10 g/mL), TM format ABP control (2 g/mL), ABP1 (2
g/mL),
or ABP1 + pembrolizumab. Cells were incubated for 48 hours before supernatants
were
collected and cells were stained for checkpoint expression. In some samples,
brefeldin A
(an inhibitor of cytokine secretion) was added to the last five hours of
stimulation for
detection of cytokines by intracellular cytokine staining. Shown are TNF
production (FIG.
12A) and IFNy production (FIG. 12B).
[84] FIG. 13 is a series of graphs showing GITR clustering and
internalization after
antibody binding. GITR internalization was measured in CD4+ cells (FIGs 13A-E)
and
CD8+ cells (Figures 13F-J) from either Donor 1 (Figures 13A-B, 13F-G) or Donor
2
(Figures 13C-D, 13H-I). Cells were treated with either ABP1 TM format antibody
or
ABP35 standard bivalent antibody. As can be observed, incubation with either
ABP1 or
ABP35 inhibits the subsequent staining with ABP1Dylight650 (Figures 13A, 13C,
13F,
13H) but only incubation with ABP1 induces GITR internalization as measured by
staining with non-competitive clone 108-17 (Figures 13B, 13D, 13G, 131). A
determination of the EC50 of ABP1 on cells from both donors is shown in Figure
13E
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(CD4+ cells) and Figure 13J (CD8+ cells).
[85] FIG. 14 shows production of cytokine (IL-2) from activated T- blasts
from two
healthy human donors (Donor 1 and Donor 2, FIG. 14A and 14B, respectively)
after
treatment with anti-GITR IgG4 TM format antibody ABP1, its IgG1 format
counterpart
ABP35, and IgG4 TM format and IgG1 isotype controls. Activated CD4+/CD8+ T
cells
were treated with medium alone, recombinant GITR-Ligand, ABP1, hIgG4 TM Format
isotype control, ABP35, or hIgG1 standard format isotype control at nine doses
each: 10
ug/mL, 2 ug/mL, 0.4 ug/mL, 80 ng/mL, 16 ng/mL, 3.2 ng/mL, 0.64 ng/mL, 0.13
ng/mL,
and 0.026 ng/mL. Cells were stimulated by adding 1 ug/m1 anti-CD3 antibodies
and 2
ug/m1 anti-CD28 antibodies. FIG. 14C shows the same data for ABP1 as in FIG.
14A and
14B with a calculation of the EC50 determination in each donor.
[86] FIG. 15 shows production of cytokine (IL-2) from activated T blasts
from two
healthy human donors (Donor 1 and Donor 2, FIG. 15A and 15B, respectively)
after
treatment with anti-GITR IgG4 TM format antibody ABP1, IgG4 TM format control
("IsoTM"), SEC4, SEC9, recombinant hGITRL and IgG1 and IgG4 isotype controls.
Activated CD4+/CD8+ T cells were treated with medium alone, recombinant GITR-
Ligand, ABP1, hIgG4 TM Format isotype control, SEC4, SEC9, IgG1 standard
format
isotype control, or hIgG1 standard format isotype control at nine doses each:
10 ug/mL, 2
ug/mL, 0.4 ug/mL, 80 ng/mL, 16 ng/mL, 3.2 ng/mL, 0.64 ng/mL, 0.13 ng/mL, and
0.026
ng/mL. Cells were stimulated by adding 1 ug/m1 anti-CD3 antibodies and 2 ug/m1
anti-
CD28 antibodies. FIGS 15C (Donor 1) and 15D (Donor 2) show the same data for
ABP1
as in 15A and 15B with a calculation of the EC50 determination in each donor.
DETAILED DESCRIPTION
Definitions
[87] Unless otherwise defined, all terms of art, notations and other
scientific terminology
used herein are intended to have the meanings commonly understood by those of
skill in
the art to which this invention pertains. In some cases, terms with commonly
understood
meanings are defined herein for clarity and/or for ready reference, and the
inclusion of
such definitions herein should not necessarily be construed to represent a
difference over
what is generally understood in the art. The techniques and procedures
described or
referenced herein are generally well understood and commonly employed using
conventional methodologies by those skilled in the art, such as, for example,
the widely
utilized molecular cloning methodologies described in Sambrook et al.,
Molecular
Cloning: A Laboratory Manual 4th ed. (2012) Cold Spring Harbor Laboratory
Press, Cold
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Spring Harbor, NY. As appropriate, procedures involving the use of
commercially
available kits and reagents are generally carried out in accordance with
manufacturer-
defined protocols and conditions unless otherwise noted.
[88] As used herein, the singular forms "a," "an," and "the" include the
plural referents
unless the context clearly indicates otherwise. The terms "include," "such
as," and the like
are intended to convey inclusion without limitation, unless otherwise
specifically
indicated.
[89] As used herein, the term "comprising" also specifically includes
embodiments
µ`consisting of' and "consisting essentially of' the recited elements, unless
specifically
indicated otherwise. For example, a multispecific ABP "comprising a diabody"
includes a
multispecific ABP "consisting of a diabody" and a multispecific ABP
"consisting
essentially of a diabody."
[90] The term "about" indicates and encompasses an indicated value and a
range above
and below that value. In certain embodiments, the term "about" indicates the
designated
value 10%, 5%, or 1%. In certain embodiments, the term "about" indicates
the
designated value one standard deviation of that value.
[91] The terms "GITR," "GITR protein," and "GITR antigen" are used
interchangeably
herein to refer to human GITR, or any variants (e.g., splice variants and
allelic variants),
isoforms, and species homologs of human GITR that are naturally expressed by
cells, or
that are expressed by cells transfected with a gitr gene. In some aspects, the
GITR protein
is a GITR protein naturally expressed by a primate (e.g., a monkey or a
human), a rodent
(e.g., a mouse or a rat), a dog, a camel, a cat, a cow, a goat, a horse, or a
sheep. In some
aspects, the GITR protein is human GITR (hGITR; SEQ ID NO: 1). In some
aspects, the
GITR protein is a human GITR T43R variant (hGITR-T43R; SEQ ID NO: 2). In some
aspects, the GITR protein comprises the extracellular domain of hGITR, located
at
positions 26 - 162 of SEQ ID NOs: 1-2. In some aspects, the GITR protein is a
cynomolgus monkey GITR (cGITR; SEQ ID NO: 3). In some aspects, the GITR
protein
comprises the extracellular domain of cGITR, located at positions 20 - 156 of
SEQ ID
NOs: 3. In some aspects, the GITR protein is a murine GITR (mGITR; SEQ ID NO:
4). In
some aspects, the GITR protein comprises the extracellular domain of mGITR,
located at
positions 20 - 153 of SEQ ID NOs: 4. In some aspects, the GITR protein is a
full-length or
unprocessed GITR protein. In some aspects, the GITR protein is a truncated or
processed
GITR protein produced by post-translational modification. GITR is also known
by a
variety of synonyms, including tumor necrosis factor receptor superfamily,
member 18
(TNFRSF18); AITR, glucocorticoid-induced TNFR-related protein; activation-
inducible
TNFR family receptor; TNF receptor superfamily activation-inducible protein;
CD357;
and GITR-D.
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[92] The term "immunoglobulin" refers to a class of structurally related
proteins generally
comprising two pairs of polypeptide chains: one pair of light (L) chains and
one pair of
heavy (H) chains. In an "intact immunoglobulin," all four of these chains are
interconnected by disulfide bonds. The structure of immunoglobulins has been
well
characterized. See, e.g., Paul, Fundamental Immunology 7th ed., Ch. 5 (2013)
Lippincott
Williams & Wilkins, Philadelphia, PA. Briefly, each heavy chain typically
comprises a
heavy chain variable region (VH) and a heavy chain constant region (CH). The
heavy chain
constant region typically comprises three domains, abbreviated Cm, CH2, and
CH3. Each
light chain typically comprises a light chain variable region (VL) and a light
chain constant
region. The light chain constant region typically comprises one domain,
abbreviated CL.
[93] The term "antigen-binding protein" (ABP) refers to a protein
comprising one or more
antigen-binding domains that specifically bind to an antigen or epitope. In
some
embodiments, the antigen-binding domain binds the antigen or epitope with
specificity and
affinity similar to that of naturally occurring antibodies. In some
embodiments, the ABP
comprises, consists of, or consists essentially of an antibody. In some
embodiments, the
ABP comprises, consists of, or consists essentially of an antibody fragment.
In some
embodiments, the ABP comprises, consists of, or consists essentially of an
alternative
scaffold. A "GITR ABP," "anti-GITR ABP," or "GITR-specific ABP" is an ABP, as
provided herein, which specifically binds to the antigen GITR. In some
embodiments, the
ABP binds the extracellular domain of GITR. In certain embodiments, a GITR ABP
provided herein binds to an epitope of GITR that is conserved between or among
GITR
proteins from different species.
[94] The term "antibody" is used herein in its broadest sense and includes
certain types of
immunoglobulin molecules comprising one or more antigen-binding domains that
specifically bind to an antigen or epitope. An antibody specifically includes
intact
antibodies (e.g., intact immunoglobulins), antibody fragments, and multi-
specific
antibodies. One example of an antigen-binding domain is an antigen-binding
domain
formed by a VH -VL dimer. An antibody is one type of ABP.
[95] The term "alternative scaffold" refers to a molecule in which one or
more regions
may be diversified to produce one or more antigen-binding domains that
specifically bind
to an antigen or epitope. In some embodiments, the antigen-binding domain
binds the
antigen or epitope with specificity and affinity similar to that of naturally
occurring
antibodies. Exemplary alternative scaffolds include those derived from
fibronectin (e.g.,
AdnectinsTm), the I3-sandwich (e.g., iMab), lipocalin (e.g., Anticalins ),
EETI-II/AGRP,
BPTI/LACI-D1/ITI-D2 (e.g., Kunitz domains), thioredoxin peptide aptamers,
protein A
(e.g., Affibody ), ankyrin repeats (e.g., DARPins), gamma-B-
crystallin/ubiquitin (e.g.,
Affilins), CTLD3 (e.g., Tetranectins), Fynomers, and (LDLR-A module) (e.g.,
Avimers).
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Additional information on alternative scaffolds is provided in Binz et al.,
Nat. Biotechnol.,
2005 23:1257-1268; Skerra, Current Op/n. in Biotech., 2007 18:295-304; and
Silacci et al.,
I Biol. Chem., 2014, 289:14392-14398; each of which is incorporated by
reference in its
entirety. An alternative scaffold is one type of ABP.
[96] The term "antigen-binding domain" means the portion of an ABP that is
capable of
specifically binding to an antigen or epitope.
[97] The terms "full length antibody," "intact antibody," and "whole
antibody" are used
herein interchangeably to refer to an antibody having a structure
substantially similar to a
naturally occurring antibody structure and having heavy chains that comprise
an Fc region.
[98] The term "Fc region" means the C-terminal region of an immunoglobulin
heavy
chain that, in naturally occurring antibodies, interacts with Fc receptors and
certain
proteins of the complement system. The structures of the Fc regions of various
immunoglobulins, and the glycosylation sites contained therein, are known in
the art. See
Schroeder and Cavacini, I Allergy Cl/n. Immunol., 2010, 125:S41-52,
incorporated by
reference in its entirety. The Fc region may be a naturally occurring Fc
region, or an Fc
region modified as described elsewhere in this disclosure.
[99] The VH and VL regions may be further subdivided into regions of
hypervariability
("hypervariable regions (HVRs);" also called "complementarity determining
regions"
(CDRs)) interspersed with regions that are more conserved. The more conserved
regions
are called framework regions (FRs). Each VH and VL generally comprises three
CDRs and
four FRs, arranged in the following order (from N-terminus to C-terminus): FR1
- CDR1 -
FR2 - CDR2 - FR3 - CDR3 - FR4. The CDRs are involved in antigen binding, and
influence antigen specificity and binding affinity of the antibody. See Kabat
et al.,
Sequences of Proteins of Immunological Interest 5th ed. (1991) Public Health
Service,
National Institutes of Health, Bethesda, MD, incorporated by reference in its
entirety.
[100] The light chain from any vertebrate species can be assigned to one of
two types,
called kappa (K) and lambda (2), based on the sequence of its constant domain.
[101] The heavy chain from any vertebrate species can be assigned to one of
five different
classes (or isotypes): IgA, IgD, IgE, IgG, and IgM. These classes are also
designated a, 6,
e, y, and a, respectively. The IgG and IgA classes are further divided into
subclasses on the
basis of differences in sequence and function. Humans express the following
subclasses:
IgGl, IgG2, IgG3, IgG4, IgAl, and IgA2.
[102] The amino acid sequence boundaries of a CDR can be determined by one
of skill in
the art using any of a number of known numbering schemes, including those
described by
Kabat et al., supra ("Kabat" numbering scheme); Al-Lazikani et al., 1997,1
Mol. Biol.,
273:927-948 ("Chothia" numbering scheme); MacCallum et al., 1996,1 Mol. Biol.
262:732-745 ("Contact" numbering scheme); Lefranc et al., Dev. Comp. Immunol.,
2003,
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27:55-77 ("IMGT" numbering scheme); and Honegge and Pliickthun, I Mol. Biol.,
2001,
309:657-70 ("AHo" numbering scheme); each of which is incorporated by
reference in its
entirety.
[103] Table 1 provides the positions of CDR-L1, CDR-L2, CDR-L3, CDR-H1, CDR-
H2,
and CDR-H3 as identified by the Kabat and Chothia schemes. For CDR-H1, residue
numbering is provided using both the Kabat and Chothia numbering schemes.
[104] Unless otherwise specified, the numbering scheme used for
identification of a
particular CDR herein is the Kabat/Chothia numbering scheme. Where the
residues
encompassed by these two numbering schemes diverge (e.g., CDR-H1 and/or CDR-
H2),
the numbering scheme is specified as either Kabat or Chothia. For convenience,
CDR-H3
is sometimes referred to herein as either Kabat or Chothia. However, this is
not intended to
imply differences in sequence where they do not exist, and one of skill in the
art can
readily confirm whether the sequences are the same or different by examining
the
sequences.
[105] CDRs may be assigned, for example, using antibody numbering
software, such as
Abnum, available at www.bioinf.org.uk/abs/abnum/, and described in Abhinandan
and
Martin, Immunology, 2008, 45:3832-3839, incorporated by reference in its
entirety.
Table 1. Residues in CDRs according to Kabat and Chothia numbering schemes.
CDR Kabat Chothia
Li L24-L34 L24-L34
L2 L50-L56 L50-L56
L3 L89-L97 L89-L97
H31-H35B
H1 (Kabat Numbering) H26-H32 or H34*
H1 (Chothia Numbering) H31-H35 H26-H32
H2 H50-H65 H52-H56
H3 H95-H102 H95-H102
* The C-terminus of CDR-H1, when numbered using the Kabat numbering
convention, varies
between H32 and H34, depending on the length of the CDR.
[106] The "EU numbering scheme" is generally used when referring to a
residue in an
antibody heavy chain constant region (e.g., as reported in Kabat et al.,
supra). Unless
stated otherwise, the EU numbering scheme is used to refer to residues in
antibody heavy
chain constant regions described herein.
[107] An "antibody fragment" or "antigen-binding fragment" comprises a
portion of an
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intact antibody, such as the antigen-binding or variable region of an intact
antibody.
Antibody fragments include, for example, Fv fragments, Fab fragments, F(ab')2
fragments,
Fab' fragments, scFv (sFv) fragments, and scFv-Fc fragments.
[108] "Fv" fragments comprise a non-covalently-linked dimer of one heavy
chain variable
domain and one light chain variable domain.
[109] "Fab" fragments comprise, in addition to the heavy and light chain
variable domains,
the constant domain of the light chain and the first constant domain (Cm) of
the heavy
chain. Fab fragments may be generated, for example, by recombinant methods or
by
papain digestion of a full-length antibody.
[110] "F(ab')2" fragments contain two Fab' fragments joined, near the hinge
region, by
disulfide bonds. F(ab')2 fragments may be generated, for example, by
recombinant
methods or by pepsin digestion of an intact antibody. The F(ab') fragments can
be
dissociated, for example, by treatment with B-mercaptoethanol.
[111] "Single-chain Fv" or "sFv" or "scFv" antibody fragments comprise a VH
domain and
a VL domain in a single polypeptide chain. The VH and VL are generally linked
by a
peptide linker. See Phickthun A. (1994). In some embodiments, the linker is a
(GGGGS).
(SEQ ID NO: 5). In other embodiments, the linker is GGGGSGGGGSGGGGS (SEQ ID
NO:6). In some embodiments, n = 1, 2, 3, 4, 5, or 6. See Antibodies from
Escherichia coil.
In Rosenberg M. & Moore G.P. (Eds.), The Pharmacology ofMonoclonal Antibodies
vol.
113 (pp. 269-315). Springer-Verlag, New York, incorporated by reference in its
entirety.
[112] "scFv-Fc" fragments comprise an scFv attached to an Fc domain. For
example, an Fc
domain may be attached to the C-terminal of the scFv. The Fc domain may follow
the VH
or VL, depending on the orientation of the variable domains in the scFv (i.e.,
VH -VL or VL
-VH). Any suitable Fc domain known in the art or described herein may be used.
In some
cases, the Fc domain comprises an IgG4 Fc domain.
[113] The term "single domain antibody" refers to a molecule in which one
variable domain
of an antibody specifically binds to an antigen without the presence of the
other variable
domain. Single domain antibodies, and fragments thereof, are described in
Arabi
Ghahroudi et al., FEBS Letters, 1998, 414:521-526 and Muyldermans et al.,
Trends in
Biochem. Sc., 2001, 26:230-245, each of which is incorporated by reference in
its entirety.
[114] A "multispecific ABP" is an ABP that comprises two or more different
antigen-
binding domains that collectively specifically bind two or more different
epitopes. The two
or more different epitopes may be epitopes on the same antigen (e.g., a single
GITR
molecule expressed by a cell) or on different antigens (e.g., different GITR
molecules
expressed by the same cell). In some aspects, a multi-specific ABP binds two
different
epitopes (i.e., a "bispecific ABP"). In some aspects, a multi-specific ABP
binds three
different epitopes (i.e., a "trispecific ABP"). In some aspects, a multi-
specific ABP binds
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four different epitopes (i.e., a "quadspecific ABP"). In some aspects, a multi-
specific ABP
binds six different epitopes (i.e., a "quintspecific ABP"). In some aspects, a
multi-specific
ABP binds 6, 7, 8, or more different epitopes. Each binding specificity may be
present in
any suitable valency. Examples of multispecific ABPs are provided elsewhere in
this
disclosure.
[115] A "monospecific ABP" is an ABP that comprises a binding site that
specifically binds
to a single epitope. An example of a monospecific ABP is a naturally occurring
IgG
molecule which, while divalent, recognizes the same epitope at each antigen-
binding
domain. The binding specificity may be present in any suitable valency.
[116] The term "monoclonal antibody" refers to an antibody from a
population of
substantially homogeneous antibodies. A population of substantially
homogeneous
antibodies comprises antibodies that are substantially similar and that bind
the same
epitope(s), except for variants that may normally arise during production of
the
monoclonal antibody. Such variants are generally present in only minor
amounts. A
monoclonal antibody is typically obtained by a process that includes the
selection of a
single antibody from a plurality of antibodies. For example, the selection
process can be
the selection of a unique clone from a plurality of clones, such as a pool of
hybridoma
clones, phage clones, yeast clones, bacterial clones, or other recombinant DNA
clones. The
selected antibody can be further altered, for example, to improve affinity for
the target
("affinity maturation"), to humanize the antibody, to improve its production
in cell culture,
and/or to reduce its immunogenicity in a subject.
[117] The term "chimeric antibody" refers to an antibody in which a portion
of the heavy
and/or light chain is derived from a particular source or species, while the
remainder of the
heavy and/or light chain is derived from a different source or species.
[118] "Humanized" forms of non-human antibodies are chimeric antibodies
that contain
minimal sequence derived from the non-human antibody. A humanized antibody is
generally a human antibody (recipient antibody) in which residues from one or
more CDRs
are replaced by residues from one or more CDRs of a non-human antibody (donor
antibody). The donor antibody can be any suitable non-human antibody, such as
a mouse,
rat, rabbit, chicken, or non-human primate antibody having a desired
specificity, affinity,
or biological effect. In some instances, selected framework region residues of
the recipient
antibody are replaced by the corresponding framework region residues from the
donor
antibody. Humanized antibodies may also comprise residues that are not found
in either
the recipient antibody or the donor antibody. Such modifications may be made
to further
refine antibody function. For further details, see Jones et al., Nature, 1986,
321:522-525;
Riechmann et al., Nature, 1988, 332:323-329; and Presta, Curr. Op. Struct.
Biol., 1992,
2:593-596, each of which is incorporated by reference in its entirety.
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[119] A "human antibody" is one which possesses an amino acid sequence
corresponding
to that of an antibody produced by a human or a human cell, or derived from a
non-human
source that utilizes a human antibody repertoire or human antibody-encoding
sequences
(e.g., obtained from human sources or designed de novo). Human antibodies
specifically
exclude humanized antibodies.
[120] An "isolated ABP" or "isolated nucleic acid" is an ABP or nucleic
acid that has been
separated and/or recovered from a component of its natural environment.
Components of
the natural environment may include enzymes, hormones, and other proteinaceous
or
nonproteinaceous materials. In some embodiments, an isolated ABP is purified
to a degree
sufficient to obtain at least 15 residues of N-terminal or internal amino acid
sequence, for
example by use of a spinning cup sequenator. In some embodiments, an isolated
ABP is
purified to homogeneity by gel electrophoresis (e.g., SDS-PAGE) under reducing
or
nonreducing conditions, with detection by Coomassie0 blue or silver stain. An
isolated
ABP includes an ABP in situ within recombinant cells, since at least one
component of the
ABP's natural environment is not present. In some aspects, an isolated ABP or
isolated
nucleic acid is prepared by at least one purification step. In some
embodiments, an isolated
ABP or isolated nucleic acid is purified to at least 80%, 85%, 90%, 95%, or
99% by
weight. In some embodiments, an isolated ABP or isolated nucleic acid is
purified to at
least 80%, 85%, 90%, 95%, or 99% by volume. In some embodiments, an isolated
ABP or
isolated nucleic acid is provided as a solution comprising at least 85%, 90%,
95%, 98%,
99% to 100% ABP or nucleic acid by weight. In some embodiments, an isolated
ABP or
isolated nucleic acid is provided as a solution comprising at least 85%, 90%,
95%, 98%,
99% to 100% ABP or nucleic acid by volume.
[121] "Affinity" refers to the strength of the sum total of non-covalent
interactions between
a single binding site of a molecule (e.g., an ABP) and its binding partner
(e.g., an antigen
or epitope). Unless indicated otherwise, as used herein, "affinity" refers to
intrinsic binding
affinity, which reflects a 1:1 interaction between members of a binding pair
(e.g., ABP and
antigen or epitope). The affinity of a molecule X for its partner Y can be
represented by
the dissociation equilibrium constant (KD). The kinetic components that
contribute to the
dissociation equilibrium constant are described in more detail below. Affinity
can be
measured by common methods known in the art, including those described herein.
Affinity
can be determined, for example, using surface plasmon resonance (SPR)
technology (e.g.,
BIACORE ) or biolayer interferometry (e.g., FORTEBI0 ).
[122] With regard to the binding of an ABP to a target molecule, the terms
"bind," "specific
binding," "specifically binds to," "specific for," "selectively binds," and
"selective for" a
particular antigen (e.g., a polypeptide target) or an epitope on a particular
antigen mean
binding that is measurably different from a non-specific or non-selective
interaction (e.g.,
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with a non-target molecule). Specific binding can be measured, for example, by
measuring
binding to a target molecule and comparing it to binding to a non-target
molecule. Specific
binding can also be determined by competition with a control molecule that
mimics the
epitope recognized on the target molecule. In that case, specific binding is
indicated if the
binding of the ABP to the target molecule is competitively inhibited by the
control
molecule. In some aspects, the affinity of a GITR ABP for a non-target
molecule is less
than about 50% of the affinity for GITR. In some aspects, the affinity of a
GITR ABP for a
non-target molecule is less than about 40% of the affinity for GITR. In some
aspects, the
affinity of a GITR ABP for a non-target molecule is less than about 30% of the
affinity for
GITR. In some aspects, the affinity of a GITR ABP for a non-target molecule is
less than
about 20% of the affinity for GITR. In some aspects, the affinity of a GITR
ABP for a
non-target molecule is less than about 10% of the affinity for GITR. In some
aspects, the
affinity of a GITR ABP for a non-target molecule is less than about 1% of the
affinity for
GITR. In some aspects, the affinity of a GITR ABP for a non-target molecule is
less than
about 0.1% of the affinity for GITR.
[123] The term "kd" (sec-1), as used herein, refers to the dissociation
rate constant of a
particular ABP -antigen interaction. This value is also referred to as the
koff value.
[124] The term "ka" (M-lxsec-1), as used herein, refers to the association
rate constant of a
particular ABP -antigen interaction. This value is also referred to as the
k.11 value.
[125] The term "KD" (M), as used herein, refers to the dissociation
equilibrium constant of
a particular ABP -antigen interaction. KD = kdka.
[126] The term "KA" (M-1), as used herein, refers to the association
equilibrium constant of
a particular ABP -antigen interaction. KA = kaika.
[127] An "affinity matured" ABP is one with one or more alterations (e.g.,
in one or more
CDRs or FRs) that result in an improvement in the affinity of the ABP for its
antigen,
compared to a parent ABP which does not possess the alteration(s). In one
embodiment, an
affinity matured ABP has nanomolar or picomolar affinity for the target
antigen. Affinity
matured ABPs may be produced using a variety of methods known in the art. For
example,
Marks et al. (Bio/Technology, 1992, 10:779-783, incorporated by reference in
its entirety)
describes affinity maturation by VH and VL domain shuffling. Random
mutagenesis of
CDR and/or framework residues is described by, for example, Barbas et al.
(Proc. Nat.
Acad. Sci. USA., 1994, 91:3809-3813); Schier et al., Gene, 1995, 169:147-155;
Yelton et
al., 1 Immunol., 1995, 155:1994-2004; Jackson et al., 1 Immunol., 1995,
154:3310-33199;
and Hawkins et al, I Mol. Biol., 1992, 226:889-896; each of which is
incorporated by
reference in its entirety.
[128] An "immunoconjugate" is an ABP conjugated to one or more heterologous
molecule(s).
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[129] "Effector functions" refer to those biological activities mediated by
the Fc region of
an antibody, which activities may vary depending on the antibody isotype.
Examples of
antibody effector functions include Clq binding to activate complement
dependent
cytotoxicity (CDC), Fc receptor binding to activate antibody-dependent
cellular
cytotoxicity (ADCC), and antibody dependent cellular phagocytosis (ADCP).
[130] When used herein in the context of two or more ABPs, the term "competes
with" or
µ`cross-competes with" indicates that the two or more ABPs compete for binding
to an
antigen (e.g., GITR). In one exemplary assay, GITR is coated on a surface and
contacted
with a first GITR ABP, after which a second GITR ABP is added. In another
exemplary
assay, a first GITR ABP is coated on a surface and contacted with GITR, and
then a
second GITR ABP is added. If the presence of the first GITR ABP reduces
binding of the
second GITR ABP, in either assay, then the ABPs compete. The term "competes
with"
also includes combinations of ABPs where one ABP reduces binding of another
ABP, but
where no competition is observed when the ABPs are added in the reverse order.
However,
in some embodiments, the first and second ABPs inhibit binding of each other,
regardless
of the order in which they are added. In some embodiments, one ABP reduces
binding of
another ABP to its antigen by at least 50%, at least 60%, at least 70%, at
least 80%, or at
least 90%.
[131] The term "epitope" means a portion of an antigen the specifically
binds to an ABP.
Epitopes frequently consist of surface-accessible amino acid residues and/or
sugar side
chains and may have specific three dimensional structural characteristics, as
well as
specific charge characteristics. Conformational and non-conformational
epitopes are
distinguished in that the binding to the former but not the latter may be lost
in the presence
of denaturing solvents. An epitope may comprise amino acid residues that are
directly
involved in the binding, and other amino acid residues, which are not directly
involved in
the binding. The epitope to which an ABP binds can be determined using known
techniques for epitope determination such as, for example, testing for ABP
binding to
GITR variants with different point-mutations, or to chimeric GITR variants.
[132] Percent "identity" between a polypeptide sequence and a reference
sequence, is
defined as the percentage of amino acid residues in the polypeptide sequence
that are
identical to the amino acid residues in the reference sequence, after aligning
the sequences
and introducing gaps, if necessary, to achieve the maximum percent sequence
identity.
Alignment for purposes of determining percent amino acid sequence identity can
be
achieved in various ways that are within the skill in the art, for instance,
using publicly
available computer software such as BLAST, BLAST-2, ALIGN, MEGALIGN
(DNASTAR), CLUSTALW, CLUSTAL OMEGA, or MUSCLE software. Those skilled in
the art can determine appropriate parameters for aligning sequences, including
any
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algorithms needed to achieve maximal alignment over the full length of the
sequences
being compared.
[133] A
"conservative substitution" or a "conservative amino acid substitution,"
refers to
the substitution an amino acid with a chemically or functionally similar amino
acid.
Conservative substitution tables providing similar amino acids are well known
in the art.
By way of example, the groups of amino acids provided in Tables 2-4 are, in
some
embodiments, considered conservative substitutions for one another.
Table 2. Selected groups of amino acids that are considered conservative
substitutions for one
another, in certain embodiments.
Acidic Residues '13, and E
Basic Residues 1K, R, and H
Hydrophilic Uncharged Residues S, T, N, and Q
Aliphatic Uncharged Residues G, A, V, L, and I
Non-polar Uncharged Residues C, M, and P
Aromatic Residues Y, and W
Table 3. Additional selected groups of amino acids that are considered
conservative substitutions for
one another, in certain embodiments.
Group 1 A, S, and T
Group 2 D and E
Group 3 N and Q
Group 4 R and K
Group 5 I, L, and M
Group 6 F, Y, and W
Table 4. Further selected groups of amino acids that are considered
conservative substitutions for one
another, in certain embodiments.
Group A A and G
Group B D and E
Group C N and Q
Group D R, K, and H
Group E I, L, M, V
Group F F, Y, and W
Group G S and T
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Group H C and M
[134] Additional conservative substitutions may be found, for example, in
Creighton,
Proteins: Structures and Molecular Properties 2nd ed. (1993) W. H. Freeman &
Co., New
York, NY. An ABP generated by making one or more conservative substitutions of
amino
acid residues in a parent ABP is referred to as a "conservatively modified
variant."
[135] The term "amino acid" refers to the twenty common naturally occurring
amino acids.
Naturally occurring amino acids include alanine (Ala; A), arginine (Arg; R),
asparagine
(Asn; N), aspartic acid (Asp; D), cysteine (Cys; C); glutamic acid (Glu; E),
glutamine
(Gln; Q), Glycine (Gly; G); histidine (His; H), isoleucine (Ile; I), leucine
(Leu; L), lysine
(Lys; K), methionine (Met; M), phenylalanine (Phe; F), proline (Pro; P),
serine (Ser; S),
threonine (Thr; T), tryptophan (Trp; W), tyrosine (Tyr; Y), and valine (Val;
V).
[136] The term "vector," as used herein, refers to a nucleic acid molecule
capable of
propagating another nucleic acid to which it is linked. The term includes the
vector as a
self-replicating nucleic acid structure as well as the vector incorporated
into the genome of
a host cell into which it has been introduced. Certain vectors are capable of
directing the
expression of nucleic acids to which they are operatively linked. Such vectors
are referred
to herein as "expression vectors."
[137] The terms "host cell," "host cell line," and "host cell culture" are
used
interchangeably and refer to cells into which exogenous nucleic acid has been
introduced,
including the progeny of such cells. Host cells include "transformants" (or
"transformed
cells") and "transfectants" (or "transfected cells"), which each include the
primary
transformed or transfected cell and progeny derived therefrom. Such progeny
may not be
completely identical in nucleic acid content to a parent cell, and may contain
mutations.
[138] The term "treating" (and variations thereof such as "treat" or
"treatment") refers to
clinical intervention in an attempt to alter the natural course of a disease
or condition in a
subject in need thereof. Treatment can be performed both for prophylaxis and
during the
course of clinical pathology. Desirable effects of treatment include
preventing occurrence
or recurrence of disease, alleviation of symptoms, diminishment of any direct
or indirect
pathological consequences of the disease, preventing metastasis, decreasing
the rate of
disease progression, amelioration or palliation of the disease state, and
remission or
improved prognosis.
[139] As used herein, the term "therapeutically effective amount" or
"effective amount"
refers to an amount of an ABP or pharmaceutical composition provided herein
that, when
administered to a subject, is effective to treat a disease or disorder.
[140] As used herein, the term "subject" means a mammalian subject.
Exemplary subjects
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include humans, monkeys, dogs, cats, mice, rats, cows, horses, camels, goats,
rabbits, and
sheep. In certain embodiments, the subject is a human. In some embodiments the
subject
has a disease or condition that can be treated with an ABP provided herein. In
some
aspects, the disease or condition is a cancer.
[141] The term "package insert" is used to refer to instructions
customarily included in
commercial packages of therapeutic or diagnostic products (e.g., kits) that
contain
information about the indications, usage, dosage, administration, combination
therapy,
contraindications and/or warnings concerning the use of such therapeutic or
diagnostic
products.
[142] The term "cytotoxic agent," as used herein, refers to a substance
that inhibits or
prevents a cellular function and/or causes cell death or destruction.
[143] A "chemotherapeutic agent" refers to a chemical compound useful in
the treatment of
cancer. Chemotherapeutic agents include "anti-hormonal agents" or "endocrine
therapeutics" which act to regulate, reduce, block, or inhibit the effects of
hormones that
can promote the growth of cancer.
[144] The term "cytostatic agent" refers to a compound or composition which
arrests
growth of a cell either in vitro or in vivo. In some embodiments, a cytostatic
agent is an
agent that reduces the percentage of cells in S phase. In some embodiments, a
cytostatic
agent reduces the percentage of cells in S phase by at least about 20%, at
least about 40%,
at least about 60%, or at least about 80%.
[145] The term "tumor" refers to all neoplastic cell growth and
proliferation, whether
malignant or benign, and all pre-cancerous and cancerous cells and tissues.
The terms
µ`cancer," "cancerous," "cell proliferative disorder," "proliferative
disorder" and "tumor"
are not mutually exclusive as referred to herein. The terms "cell
proliferative disorder" and
"proliferative disorder" refer to disorders that are associated with some
degree of abnormal
cell proliferation. In some embodiments, the cell proliferative disorder is a
cancer.
[146] The term "pharmaceutical composition" refers to a preparation which
is in such form
as to permit the biological activity of an active ingredient contained therein
to be effective
in treating a subject, and which contains no additional components which are
unacceptably
toxic to the subject.
[147] The terms "modulate" and "modulation" refer to reducing or inhibiting
or,
alternatively, activating or increasing, a recited variable.
[148] The terms "increase" and "activate" refer to an increase of 10%, 20%,
30%, 40%,
50%, 60%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 2-fold, 3-fold, 4-fold, 5-fold,
10-fold,
20-fold, 50-fold, 100-fold, or greater in a recited variable, such as GITR
signaling activity.
[149] The terms "reduce" and "inhibit" refer to a decrease of 10%, 20%,
30%, 40%, 50%,
60%, 70%, 75%, 80%, 85%, 90%, 95%, 2-fold, 3-fold, 4-fold, 5-fold, 10-fold, 20-
fold, 50-
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fold, 100-fold, or greater in a recited variable, such as (a) a number of
regulatory T cells
and/or (b) the symptoms of a disease or condition, such as the presence or
size of
metastases or the size of a primary tumor.
[150] The term "agonize" refers to the activation of receptor signaling to
induce a
biological response associated with activation of the receptor. An "agonist"
is an entity,
such as an ABP provided herein, that binds to and agonizes a receptor.
[151] The term "antagonize" refers to the inhibition of receptor signaling
to inhibit a
biological response associated with activation of the receptor. An
"antagonist" is an entity,
such as an ABP, that binds to and antagonizes a receptor.
[152] The term "multimerize" refers to the act of forming "multimers" of an
entity by
assembling the entity to form a supra-entity structure held together by non-
covalent or
covalent interactions. Multimers include "homo-multimers," which are
assemblies formed
from multiple units of the same entity, or "hetero-multimers," which are
assemblies
comprising at least one unit of a first entity and at least one unit of a
second entity. When
used herein to refer to GITR, the term multimerize refers to the assembly of
multiple GITR
molecules expressed on the surface of a cell that is induced, for example, by
binding of an
ABP provided herein or by GITRL. Such multimerization is associated with the
activation
of GITR signaling. See Nocentini et al., Br. I Pharmacol., 2012, 165:2089-
2099,
incorporated by reference in its entirety.
[153] The term "effector T cell" includes T helper (i.e., CD4+) cells and
cytotoxic (i.e.,
CD8+) T cells. CD4+ effector T cells contribute to the development of several
immunologic processes, including maturation of B cells into plasma cells and
memory B
cells, and activation of cytotoxic T cells and macrophages. CD8+ effector T
cells destroy
virus-infected cells and tumor cells. See Seder and Ahmed, Nature Immunol.,
2003, 4:835-
842, incorporated by reference in its entirety, for additional information on
effector T cells.
[154] The term "regulatory T cell" includes cells that regulate
immunological tolerance, for
example, by suppressing effector T cells. In some aspects, the regulatory T
cell has a
CD4+CD25+Foxp3+ phenotype. In some aspects, the regulatory T cell has a
CD8+CD25+
phenotype. See Nocentini et al., Br. I Pharmacol., 2012, 165:2089-2099,
incorporated by
reference in its entirety, for additional information on regulatory T cells
expressing GITR.
[155] The term "dendritic cell" refers to a professional antigen-presenting
cell capable of
activating a naive T cell and stimulating growth and differentiation of a B
cell.
GITR Antigen-Binding Proteins
1.1. GITR Binding and Target Cells
[156] Provided herein are ABPs that specifically bind to GITR. In some
aspects, the GITR
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is hGITR (SEQ ID NO: 1). In some aspects, the GITR is hGITR-T43R (SEQ ID NO:
2). In
some aspects, the GITR is cGITR (SEQ ID NO: 3). In some embodiments, the GITR
is
mGITR (SEQ ID NO: 4).
[157] In some embodiments, the ABPs provided herein specifically bind to
hGITR (SEQ ID
NO: 1), hGITR-T43R (SEQ ID NO: 2), cGITR (SEQ ID NO: 3), and mGITR (SEQ ID
NO: 4). In some embodiments, the ABPs provided herein specifically bind to
hGITR (SEQ
ID NO: 1), hGITR-T43R (SEQ ID NO: 2), and cGITR (SEQ ID NO: 3). In some
embodiments, the ABPs provided herein specifically bind to hGITR (SEQ ID NO:
1) and
hGITR-T43R (SEQ ID NO: 2). In some embodiments, the ABPs provided herein do
not
bind mGITR (SEQ ID NO: 4).
[158] In some embodiments, the ABPs provided herein specifically bind to
the extracellular
domain of GITR.
[159] The GITR may be expressed on the surface of any suitable target cell.
In some
embodiments, the target cell is an effector T cell. In some embodiments, the
target cell is a
regulatory T cell. In some embodiments, the target cell is a natural killer
(NK) cell. In
some embodiments, the target cell is a natural killer T (NKT) cell. In some
embodiments,
the target cell is a dendritic cell. In some aspects, the dendritic cell is a
plasmacytoid
dendritic cell. In some embodiments, the target cell is a B cell. In some
aspects, the B cell
is a plasma cell. See Nocentini et al., Br. I Pharmacol., 2012, 165:2089-2099,
incorporated by reference in its entirety.
[160] In some embodiments, the ABPs provided herein specifically bind to a
GITR
monomer.
[161] In some embodiments, the ABPs provided herein specifically bind to a
GITR
multimer. In some aspects, the multimer comprises two GITR molecules. In some
aspects,
the multimer comprises three GITR molecules. In some aspects, the multimer
comprises
four GITR molecules. In some aspects, the multimer comprises five GITR
molecules. In
some aspects, the multimer comprises six GITR molecules. In some aspects, the
multimer
comprises more than six GITR molecules.
[162] In some embodiments, the ABPs provided herein comprise, consist of,
or consist
essentially of an immunoglobulin molecule. In some aspects, the immunoglobulin
molecule comprises, consists of, or consists essentially of an antibody.
[163] In some embodiments, the ABPs provided herein comprise a light chain.
In some
aspects, the light chain is a kappa light chain. In some aspects, the light
chain is a lambda
light chain.
[164] In some embodiments, the ABPs provided herein comprise a heavy chain.
In some
aspects, the heavy chain is an IgA. In some aspects, the heavy chain is an
IgD. In some
aspects, the heavy chain is an IgE. In some aspects, the heavy chain is an
IgG. In some
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aspects, the heavy chain is an IgM. In some aspects, the heavy chain is an
IgGl. In some
aspects, the heavy chain is an IgG2. In some aspects, the heavy chain is an
IgG3. In some
aspects, the heavy chain is an IgG4. In some aspects, the heavy chain is an
IgAl. In some
aspects, the heavy chain is an IgA2.
[165] In some embodiments, the ABPs provided herein comprise, consist of,
or consist
essentially of an antibody fragment. In some aspects, the antibody fragment is
an Fv
fragment. In some aspects, the antibody fragment is a Fab fragment. In some
aspects, the
antibody fragment is a F(ab')2fragment. In some aspects, the antibody fragment
is a Fab'
fragment. In some aspects, the antibody fragment is an scFv (sFv) fragment. In
some
aspects, the antibody fragment is an scFv-Fc fragment. In some aspects, the
antibody
fragment is a fragment of a single domain antibody.
[166] In some embodiments, the ABPs provided herein are monoclonal
antibodies. In some
embodiments, the ABPs provided herein are polyclonal antibodies.
[167] In some embodiments, the ABPs provided herein comprise, consist of,
or consist
essentially of chimeric antibodies. In some embodiments, the ABPs provided
herein
comprise, consist of, or consist essentially of humanized antibodies. In some
embodiments,
the ABPs provided herein comprise, consist of, or consist essentially of human
antibodies.
[168] In some embodiments, the ABPs provided herein comprise, consist of,
or consist
essentially of affinity matured ABPs. In some aspects, the ABPs are affinity
matured ABPs
derived from an ABP provided herein.
[169] In some embodiments, the ABPs provided herein comprise, consist of,
or consist
essentially of an alternative scaffold. Any suitable alternative scaffold may
be used. In
some aspects, the ABPs provided herein comprise, consist of, or consist
essentially of an
alternative scaffold selected from an AdnectinTm, an iMab, an Anticalin , an
EETI-
II/AGRP, a Kunitz domain, a thioredoxin peptide aptamer, an Affibody , a
DARPin, an
Affilin, a Tetranectin, a Fynomer, and an Avimer.
[170] In some embodiments, an ABP provided herein inhibits binding of GITR
to GITRL.
In some aspects, the ABP inhibits binding of GITR to GITRL by at least about
50%. In
some aspects, the ABP inhibits binding of GITR to GITRL by at least about 75%.
In some
aspects, the ABP inhibits binding of GITR to GITRL by at least about 90%. In
some
aspects, the ABP inhibits binding of GITR to GITRL by at least about 95%.
1.2. Monospecific and Multispecific GITR Antigen-Binding Proteins
[171] In some embodiments, the ABPs provided herein are monospecific ABPs.
In some
aspects, the monospecific ABPs bind to the same epitope on two or more
different GITR
molecules. In some aspects, the monospecific ABPs bind to the same epitope on
two GITR
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molecules. In some aspects, the monospecific ABPs bind to the same epitope on
three
GITR molecules. In some aspects, the monospecific ABPs bind to the same
epitope on
four GITR molecules. In some aspects, the monospecific ABPs bind to the same
epitope
on five GITR molecules. In some aspects, the monospecific ABPs bind to the
same epitope
on six GITR molecules. In some aspects, the monospecific ABPs bind to the same
epitope
on more than six GITR molecules.
[172] In some embodiments, the monospecific ABPs provided herein are
multivalent. As
used herein, the term "multivalent" refers to an antibody with, e.g., more
than two binding
regions (i.e., that comprise VH and VL regions). In some aspects, the
monospecific ABPs
are bivalent. In some aspects, the monospecific ABPs are trivalent. In some
aspects, the
monospecific ABPs are tetravalent. In some aspects, the monospecific ABPs are
pentavalent. In some aspects, the monospecific ABPs are hexavalent. In some
aspects, the
monospecific ABPs are septivalent. In some aspects, the monospecific ABPs are
octavalent.
[173] In some embodiments, the monospecific multivalent ABPs disclosed
herein are
tetravalent.
[174] In some embodiments, the ABPs provided herein are multispecific ABPs.
In some
aspects, the multispecific ABPs bind to two or more epitopes on two or more
different
GITR molecules. In some aspects, the multispecific ABPs bind to two or more
epitopes on
two GITR molecules. In some aspects, the multispecific ABPs bind to two or
more
epitopes on three GITR molecules. In some aspects, the multispecific ABPs bind
to two or
more epitopes on four GITR molecules. In some aspects, the multispecific ABPs
bind to
two or more epitopes on five GITR molecules. In some aspects, the
multispecific ABPs
bind to two or more epitopes on six GITR molecules. In some aspects, the
multispecific
ABPs bind to two or more epitopes on seven GITR molecules. In some aspects,
the
multispecific ABPs bind to two or more epitopes on eight GITR molecules. In
some
aspects, the multispecific ABPs bind to two or more epitopes on nine GITR
molecules. In
some aspects, the multispecific ABPs bind to two or more epitopes on ten GITR
molecules. In some aspects, the multispecific ABPs bind to two or more
epitopes on
eleven GITR molecules. In some aspects, the multispecific ABPs bind to two or
more
epitopes on twelve GITR molecules. In some aspects, the multispecific ABPs
bind to two
or more epitopes on more than twelve GITR molecules.
[175] The multispecific ABPs provided herein may bind any suitable number
of epitopes on
GITR. In some aspects, the multispecific ABPs bind two epitopes on GITR. In
some
aspects, the multispecific ABPs bind three epitopes on GITR. In some aspects,
the
multispecific ABPs bind four epitopes on GITR. In some aspects, the
multispecific ABPs
bind five epitopes on GITR. In some aspects, the multispecific ABPs bind six
epitopes on
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GITR. In some aspects, the multispecific ABPs bind seven epitopes on GITR. In
some
aspects, the multispecific ABPs bind eight epitopes on GITR. In some aspects,
the
multispecific ABPs bind nine epitopes on GITR. In some aspects, the
multispecific ABPs
bind ten epitopes on GITR. In some aspects, the multispecific ABPs bind eleven
epitopes
on GITR. In some aspects, the multispecific ABPs bind twelve epitopes on GITR.
In some
aspects, the multispecific ABPs bind more than twelve epitopes on GITR.
[176] In some aspects, a multispecific ABP provided herein binds at least
two different
epitopes on at least two different GITR molecules. In some aspects, a
multispecific ABP
provided herein binds at least three different epitopes on at least three
different GITR
molecules. In some aspects, a multispecific ABP provided herein binds at least
four
different epitopes on at least four different GITR molecules. In some aspects,
a
multispecific ABP provided herein binds at least five different epitopes on at
least five
different GITR molecules.
[177] In some embodiments, a multispecific ABP provided herein comprises a
first antigen-
binding domain that specifically binds a first epitope on GITR and a second
antigen-
binding domain that specifically binds a second epitope on GITR, wherein the
first epitope
and the second epitope are different. In some aspects, the multispecific ABP
further
comprises one or more additional antigen-binding domains that specifically
bind to one or
more additional epitopes on GITR, wherein each of the additional epitopes are
different
from the epitopes bound by the first antigen-binding domain, the second
antigen-binding
domain, or any other further antigen-binding domains of the ABP.
[178] In some embodiments, a multispecific ABP provided herein binds to an
epitope on a
GITR molecule and an epitope on another molecule that is not GITR. Any
suitable non-
GITR molecule may be bound by an ABP provided herein. In some aspects, the non-
GITR
molecule is another member of the tumor necrosis factor receptor superfamily
(TNFRSF).
In some aspects, the other member of the TNFRSF is selected from CD27, CD40,
EDA2R,
EDAR, FAS, LTBR, NGFR, RELT, TNFRSF1A, TNFRSF1B, TNFRSF4, TNFRSF6B,
TNFRSF8, TNFRSF9, TNFRSF10A, TNFRSF10B, TNFRSF10C, TNFRSF10D,
TFNRSF11A, TNFRSF11B, TNFRSF12A, TNFRSF13B, TNFRSF13C, TNFRSF14,
TNFRSF17, TNFRSF18, TNFRSF19, TNFRSF21, and TNFRSF25.
[179] Many multispecific ABP constructs are known in the art, and the ABPs
provided
herein may be provided in the form of any suitable multispecific suitable
construct.
[180] In some embodiments, the multispecific ABP comprises an
immunoglobulin
comprising at least two different heavy chain variable regions each paired
with a common
light chain variable region (i.e., a "common light chain antibody"). The
common light
chain variable region forms a distinct antigen-binding domain with each of the
two
different heavy chain variable regions. See Merchant et al., Nature
Biotechnol., 1998,
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16:677-681, incorporated by reference in its entirety.
[181] In some embodiments, the multivalent ABPs disclosed herein comprise
an
immunoglobulin comprising an antibody or fragment thereof attached to one or
more of
the N- or C-termini of the heavy or light chains of such immunoglobulin. See,
e.g.,U U.S.
Patent No. 8,722,859 and Coloma and Morrison, Nature Biotechnol., 1997, 15:159-
163,
each incorporated by reference in its entirety. In some aspects, such ABP
comprises a
tetravalent bispecific antibody. In some aspects, such ABP comprises a
tetravalent
monospecific (TM) antibody.
[182] In some embodiments, the multivalent ABP comprises a hybrid
immunoglobulin
comprising at least two different heavy chain variable regions and at least
two different
light chain variable regions. See Milstein and Cuello, Nature, 1983, 305:537-
540; and
Staerz and Bevan, Proc. Natl. Acad. Sci. USA, 1986, 83:1453-1457; each of
which is
incorporated by reference in its entirety.
[183] In some embodiments, the multivalent ABP comprises immunoglobulin chains
with
alterations to reduce the formation of side products that do not have
multispecificity. In
some aspects, the ABPs comprise one or more "knobs-into-holes" modifications
as
described in U.S. Pat. No. 5,731,168, incorporated by reference in its
entirety.
[184] In some embodiments, the multivalent ABP comprises immunoglobulin
chains with
one or more electrostatic modifications to promote the assembly of Fc hetero-
multimers.
See WO 2009/089004, incorporated by reference in its entirety.
[185] In some embodiments, the multivalent ABP comprises a bispecific
single chain
molecule. See Traunecker et al., EiVIBO 1, 1991, 10:3655-3659; and Gruber et
al., I
Immunol., 1994, 152:5368-5374; each of which is incorporated by reference in
its entirety.
[186] In some embodiments, the multivalent ABP comprises a heavy chain
variable domain
and a light chain variable domain, or a single domain antibody VHH domain,
connected by
a polypeptide linker, where the length of the linker is selected to promote
assembly of
multivalent ABPs with the desired multispecificity. For example, monospecific
scFvs
generally form when a heavy chain V residue prevents pairing of heavy and
light chain
variable domains on the same polypeptide chain, thereby allowing pairing of
heavy and
light chain variable domains from one chain with the complementary domains on
another
chain. The resulting ABPs therefore have multispecificity, with the
specificity of each
binding site contributed by more than one polypeptide chain. Polypeptide
chains
comprising heavy and light chain variable domains that are joined by linkers
between 3
and 12 amino acid residues form predominantly dimers (termed diabodies). With
linkers
between 0 and 2 amino acid residues, trimers (termed triabodies) and tetramers
(termed
tetrabodies) are favored. However, the exact type of oligomerization appears
to depend on
the amino acid residue composition and the order of the variable domain in
each
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polypeptide chain (e.g., VH-linker-VL vs. VL-linker-VH), in addition to the
linker length. A
skilled person can select the appropriate linker length based on the desired
multispecificity.
[187] In some embodiments, the monospecific or multispecific ABP comprises
a diabody.
See Hollinger et al., Proc. Natl. Acad. Sci. USA, 1993, 90:6444-6448,
incorporated by
reference in its entirety. In some embodiments, the monospecific or
multispecific ABP
comprises a triabody. See Todorovska et al., I Immunol. Methods, 2001, 248:47-
66,
incorporated by reference in its entirety. In some embodiments, the
monospecific or
multispecific ABP comprises a tetrabody. See id, incorporated by reference in
its entirety.
[188] In some embodiments, the multispecific ABP comprises a trispecific
F(ab')3
derivative. See Tuft et al. I Immunol., 1991, 147:60-69, incorporated by
reference in its
entirety.
[189] In some embodiments, the monospecific or multispecific ABP comprises
a cross-
linked antibody. See U.S. Patent No. 4,676,980; Brennan et al., Science, 1985,
229:81-83;
Staerz, et al. Nature, 1985, 314:628-631; and EP 0453082; each of which is
incorporated
by reference in its entirety.
[190] In some embodiments, the monospecific or multispecific ABP comprises
antigen-
binding domains assembled by leucine zippers. See Kostelny et al., I Immunol.,
1992,
148:1547-1553, incorporated by reference in its entirety.
[191] In some embodiments, the monospecific or multispecific ABP comprises
complementary protein domains. In some aspects, the complementary protein
domains
comprise an anchoring domain (AD) and a dimerization and docking domain (DDD).
In
some embodiments, the AD and DDD bind to each other and thereby enable
assembly of
multispecific ABP structures via the "dock and lock" (DNL) approach. ABPs of
many
specificities may be assembled, including bispecific ABPs, trispecific ABPs,
tetraspecific
ABPs, pentaspecific ABPs, and hexaspecific ABPs. Multispecific ABPs comprising
complementary protein domains are described, for example, in U.S. Pat. Nos.
7,521,056;
7,550,143; 7,534,866; and 7,527,787; each of which is incorporated by
reference in its
entirety.
[192] In some embodiments, the monospecific or multispecific ABP comprises
a hybrid of
an antibody molecule and a non-antibody molecule with specificity for GITR or
another
target. See WO 93/08829, incorporated by reference in its entirety, for
examples of such
ABPs. In some aspects the non-antibody molecule is GITRL.
[193] In some embodiments, the monospecific or multispecific ABP comprises
a dual
action Fab (DAF) antibody as described in U.S. Pat. Pub. No. 2008/0069820,
incorporated
by reference in its entirety.
[194] In some embodiments, the multispecific ABP comprises an antibody
formed by
reduction of two parental molecules followed by mixing of the two parental
molecules and
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reoxidation to assembly a hybrid structure. See Carlring et al., PLoS One,
2011, 6:e22533,
incorporated by reference in its entirety.
[195] In some embodiments, the monospecific or multispecific ABP comprises
a DVD-
Tem. A DVD-Iirm is a dual variable domain immunoglobulin that can bind to two
or more
antigens. DVD-IgsTm are described in U.S. Pat. No. 7,612,181, incorporated by
reference
in its entirety.
[196] In some embodiments, the monospecific or multispecific ABP comprises
a DART.
DARTs TM are described in Moore et al., Blood, 2011, 117:454-451, incorporated
by
reference in its entirety.
[197] In some embodiments, the multispecific ABP comprises a DuoBody .
DuoBodies
are described in Labrijn et al., Proc. Natl. Acad. Sci. USA, 2013, 110:5145-
5150; Gramer
et al., mAbs, 2013, 5:962-972; and Labrijn et al., Nature Protocols, 2014,
9:2450-2463;
each of which is incorporated by reference in its entirety.
[198] In some embodiments, the monospecific or multispecific ABP disclosed
herein
comprises an antibody fragment attached to another antibody or fragment. The
attachment
can be covalent or non-covalent. When the attachment is covalent, it may be in
the form of
a fusion protein or via a chemical linker. Illustrative examples of
multispecific ABPs
comprising antibody fragments attached to other antibodies include tetravalent
bispecific
antibodies, where an scFv is fused to the C-terminus of the CH3 from an IgG.
See Coloma
and Morrison, Nature Biotechnol., 1997, 15:159-163. Other examples include
antibodies in
which a Fab molecule is attached to the constant region of an immunoglobulin.
See Miler
et al., I Immunol., 2003, 170:4854-4861, incorporated by reference in its
entirety. Any
suitable fragment may be used, including any of the fragments described herein
or known
in the art.
[199] In some embodiments, the monospecific or multispecific ABP comprises
a CovX-
Body. CovX-Bodies are described, for example, in Doppalapudi et al., Proc.
Natl. Acad.
Sci. USA, 2010, 107:22611-22616, incorporated by reference in its entirety.
[200] In some embodiments, the monospecific or multispecific ABP comprises
an Fcab
antibody, where one or more antigen-binding domains are introduced into an Fc
region.
Fcab antibodies are described in Wozniak-Knopp et al., Protein Eng. Des. Se.,
2010,
23:289-297, incorporated by reference in its entirety.
[201] In some embodiments, the monospecific or multispecific ABP comprises
an TandAb
antibody. TandAb antibodies are described in Kipriyanov et al., I Mol. Biol.,
1999,
293:41-56 and Zhukovsky et al., Blood, 2013, 122:5116, each of which is
incorporated by
reference in its entirety.
[202] In some embodiments, the multispecific ABP comprises a tandem Fab.
Tandem Fabs
are described in WO 2015/103072, incorporated by reference in its entirety.
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[203] In some embodiments, the multispecific ABP comprises a ZybodyTm.
ZybodiesTm are
described in LaFleur et al., mAbs, 2013, 5:208-218, incorporated by reference
in its
entirety.
1.3. Antigen-Binding Proteins Multimerizing GITR
[204] In some embodiments, the ABPs provided herein multimerize GITR
expressed on the
surface of a target cell. The ABPs provided herein may be designed, based on
their valency
and specificity, to multimerize any suitable number of GITR molecules.
[205] In some embodiments, the ABPs provided herein multimerize two GITR
molecules.
In some embodiments, the ABPs provided herein multimerize three GITR
molecules. In
some embodiments, the ABPs provided herein multimerize four GITR molecules. In
some
embodiments, the ABPs provided herein multimerize five GITR molecules. In some
embodiments, the ABPs provided herein multimerize six GITR molecules. In some
embodiments, the ABPs provided herein multimerize seven GITR molecules. In
some
embodiments, the ABPs provided herein multimerize eight GITR molecules. In
some
embodiments, the ABPs provided herein multimerize nine GITR molecules. In some
embodiments, the ABPs provided herein multimerize ten GITR molecules. In some
embodiments, the ABPs provided herein multimerize eleven GITR molecules. In
some
embodiments, the ABPs provided herein multimerize twelve GITR molecules.
[206] In some embodiments, the ABPs provided herein multimerize at least
two GITR
molecules. In some embodiments, the ABPs provided herein multimerize at least
three
GITR molecules. In some embodiments, the ABPs provided herein multimerize at
least
four GITR molecules. In some embodiments, the ABPs provided herein multimerize
at
least five GITR molecules. In some embodiments, the ABPs provided herein
multimerize
at least six GITR molecules. In some embodiments, the ABPs provided herein
multimerize
at least seven GITR molecules. In some embodiments, the ABPs provided herein
multimerize at least eight GITR molecules. In some embodiments, the ABPs
provided
herein multimerize at least nine GITR molecules. In some embodiments, the ABPs
provided herein multimerize at least ten GITR molecules. In some embodiments,
the ABPs
provided herein multimerize at least eleven GITR molecules. In some
embodiments, the
ABPs provided herein multimerize at least twelve GITR molecules.
[207] In some embodiments, the ABPs provided herein multimerize two to twelve
GITR
molecules. In some embodiments, the ABPs provided herein multimerize three to
ten
GITR molecules. In some embodiments, the ABPs provided herein multimerize
three to
six GITR molecules. In some embodiments, the ABPs provided herein multimerize
three
to five GITR molecules. In some embodiments, the ABPs provided herein
multimerize
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three to four GITR molecules.
1.4. GITR Agonism
[208] In some embodiments, the ABPs provided herein agonize GITR upon
binding. Such
agonism can result from the multimerization of GITR by the ABPs as described
elsewhere
in this disclosure. See FIG. 1.
[209] In some embodiments, agonism of GITR by an ABP provided herein results
in
modulation of NF-KB activity in a target cell. See U.S. Pat. No. 7,812,135,
herein
incorporated by reference in its entirety. In some aspects, agonism of GITR
results in
modulation of fkB activity or stability in a target cell.
[210] In some embodiments, agonism of GITR by an ABP provided herein
results in
activation of the MAPK pathway in a target cell. In some aspects, the
components of the
MAPK pathway that are activated by an ABP provided herein include one or more
of p38,
JNK, and ERK. See Nocentini et al., Proc. Natl. Acad. Sci. USA, 1997, 94:6216-
6221;
Ronchetti et al., Eur. I Immunol., 2004, 34:613-622; and Esparza et al., I
Immunol., 2005,
174:7869-7874; each of which is incorporated by reference in its entirety.
[211] In some embodiments, agonism of GITR by an ABP provided herein
results in
increased secretion of IL-2Ra, IL-2, IL-8, and/or IFNy by a target cell. See
Ronchetti et al.,
Eur. I Immunol., 2004, 34:613-622, incorporated by reference in its entirety.
[212] In some embodiments, agonism of GITR by an ABP provided herein increases
the
proliferation, survival, and/or function of an effector T cell. In some
aspects the effector T
cell is a CD4+ effector T cell. In some aspects, the effector T cell is a CD8+
effector T
cell.
[213] In some embodiments, agonism of GITR by an ABP provided herein abrogates
suppression of an effector T cell by a regulatory T cell. In some aspects, the
regulatory T
cell is a CD4+CD25+Foxp3+ regulatory T cell. In some aspects, the regulatory T
cell is a
CD8+CD25+ regulatory T cell.
[214] In some embodiments, agonism of GITR by an ABP provided herein alters
the
frequency of occurrence or distribution of regulatory T cells. In some
aspects, the
frequency of regulatory T cells is decreased. In some aspects, the frequency
of regulatory
T cells is reduced in a particular tissue. In some aspects, the intratumoral
accumulation of
regulatory T cells is decreased, resulting in a more favorable ratio of
effector T cells to
regulatory T cells, and enhancing CD8+ T cell activity. See Cohen et al., PLoS
One, 2010,
5:e10436.
[215] In some embodiments, agonism of GITR by an ABP provided herein increases
the
activity of a natural killer (NK) cell. In some embodiments, agonism of GITR
by an ABP
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provided herein increases the activity of an antigen presenting cell. In some
embodiments,
agonism of GITR by an ABP provided herein increases the activity of a
dendritic cell. In
some embodiments, agonism of GITR by an ABP provided herein increases the
activity of
a B cell.
[216] In some embodiments, agonism of GITR by an ABP provided herein results
in an
enhancement of an immune response. In some embodiments, agonism of GITR by an
ABP
provided herein results in the delay of onset of a tumor. In some embodiments,
agonism of
GITR by an ABP provided herein results in the reduction of the size of a
tumor. In some
embodiments, agonism of GITR by an ABP provided herein results in a reduction
in the
number of metastases.
[217] In some embodiments, agonism of GITR by a multivalent monospecific ABP
provided herein results in a greater maximum amount of agonism than a bivalent
monospecific antibody. In some embodiments, the additional valency results in
an effect
on the EC50 that that is more than the additive effects of each of the binding
domains. In
some embodiments, a tetravalent monospecific ABP provided herein has a greater
maximum amount of agonism than a bivalent monospecific antibody.
[218] In some embodiments, the multispecific ABPs provided herein are more
potent
agonists of GITR than mixtures of the corresponding monospecific ABPs. For
example, if
a multispecific ABP provided herein comprises two different epitope
specificities (e.g., A
and B) then, in some embodiments, the agonism of GITR by such multispecific
ABP is
greater than the agonism of GITR by a mixture of two monospecific ABPs that
each
comprise one of the two specificities (e.g., A or B). In some embodiments, the
additional
specificities of a multispecific ABP provided herein yield a synergistic
(i.e., greater than
additive) increase in potency when compared to mixtures of monospecific ABPs
each
having only one of the specificities of the multispecific ABP.
1.5. Affinity of Antigen-Binding Proteins for GITR
[219] In some embodiments, the affinity of an ABP provided herein for GITR
as indicated
by KD, is less than about 10-5 M, less than about 10-6 M, less than about 10-7
M, less than
about 10-8 M, less than about 10-9 M, less than about 10-10 M, less than about
10-11M, or
less than about 10-12 M. In some embodiments, the affinity of the ABP is
between about
10-7 M and 10-12 M. In some embodiments, the affinity of the ABP is between
about 10-7 M
and 10-11 M. In some embodiments, the affinity of the ABP is between about 10-
7 M and
10-10 M. In some embodiments, the affinity of the ABP is between about 10-7 M
and 10-9
M. In some embodiments, the affinity of the ABP is between about 10-7 M and 10-
8 M. In
some embodiments, the affinity of the ABP is between about 10-8 M and 10-12 M.
In some
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embodiments, the affinity of the ABP is between about 10-8 M and 10-11 M. In
some
embodiments, the affinity of the ABP is between about 10-9 M and 10-11 M. In
some
embodiments, the affinity of the ABP is between about 10-10 M and 10-11 M.
[220] In some embodiments, the ABPs provided herein specifically bind to
hGITR (SEQ ID
NO: 1) with a KD ofX and to cGITR with a KD of <10X. In some embodiments, the
ABPs
provided herein specifically bind to hGITR (SEQ ID NO: 1) with a KD ofX and to
cGITR
with a KD of <5X. In some embodiments, the ABPs provided herein specifically
bind to
hGITR (SEQ ID NO: 1) with a KD ofX and to cGITR with a KD of <2X. In some
aspects,
Xis any KD described in this disclosure. In some aspects, Xis 0.01 nM, 0.1 nM,
1 nM, 10
nM, 20 nM, 50 nM, or 100 nM.
[221] In some embodiments, KD, ka, and ka are determined using surface
plasmon resonance
(SPR). In some aspects, the SPR analysis utilizes a BIACORE instrument. In
some
aspects, the antigen is immobilized on a carboxymethylated dextran biosensor
chip (CM4
or CM5) and contacted with an ABP provided herein. Association and
dissociation rate
constants may be calculated using the BlAevaluation software and a one-to-one
Langmuir binding model. In some aspects, the assay is performed at 25 C. In
some
aspects, the assay is performed at 37 C.
[222] In some embodiments, KD, ka, and ka are determined using biolayer
interferometry
(BLI). Any suitable BLI method may be used. In some aspects, the BLI analysis
utilizes a
FORTEBIO instrument. In some aspects, an anti-human IgG Fc capture (AHC)
biosensor
is used to capture ABPs onto the surface of a sensor. Subsequently,
association of the ABP
and antigen is monitored by contacting the immobilized ABP with different
concentrations
of GITR. Dissociation of the antigen and ABP is then measured in a buffer
without GITR.
Association and dissociation rate constants are calculated using the kinetic
modules of the
FORTEBIO Analysis Software. In some aspects, the assay is performed at 30 C.
[223] In other embodiments, KD may be determined by a radiolabeled antigen-
binding
assay, as described in Chen et al. I Mol. Biol., 1999, 293:865-881,
incorporated by
reference in its entirety.
1.5.1.Glycosylation Variants
[224] In certain embodiments, an ABP provided herein may be altered to
increase, decrease
or eliminate the extent to which it is glycosylated. Glycosylation of
polypeptides is
typically either "N-linked" or "0-linked."
[225] "N-linked" glycosylation refers to the attachment of a carbohydrate
moiety to the side
chain of an asparagine residue. The tripeptide sequences asparagine-X-serine
and
asparagine-X-threonine, where X is any amino acid except proline, are the
recognition
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sequences for enzymatic attachment of the carbohydrate moiety to the
asparagine side
chain. Thus, the presence of either of these tripeptide sequences in a
polypeptide creates a
potential glycosylation site.
[226] "0-linked" glycosylation refers to the attachment of one of the
sugars N-
acetylgalactosamine, galactose, or xylose to a hydroxyamino acid, most
commonly serine
or threonine, although 5-hydroxyproline or 5-hydroxylysine may also be used.
[227] Addition or deletion of N-linked glycosylation sites to or from an
ABP provided
herein may be accomplished by altering the amino acid sequence such that one
or more of
the above-described tripeptide sequences is created or removed. Addition or
deletion of 0-
linked glycosylation sites may be accomplished by addition, deletion, or
substitution of
one or more serine or threonine residues in or to (as the case may be) the
sequence of an
ABP.
[228] In some embodiments, an ABP provided herein comprises a glycosylation
motif that
is different from a naturally occurring ABP. Any suitable naturally occurring
glycosylation
motif can be modified in the ABPs provided herein. The structural and
glycosylation
properties of immunoglobulins, for example, are known in the art and
summarized, for
example, in Schroeder and Cavacini, I Allergy Cl/n. Immunol., 2010, 125:S41-
52,
incorporated by reference in its entirety.
[229] In some embodiments, an ABP provided herein comprises an IgG1 Fc
region with
modification to the oligosaccharide attached to asparagine 297 (Asn 297).
Naturally
occurring IgG1 antibodies produced by mammalian cells typically comprise a
branched,
biantennary oligosaccharide that is generally attached by an N-linkage to Asn
297 of the
CH2 domain of the Fc region. See Wright et al., TIB TECH, 1997, 15:26-32,
incorporated by
reference in its entirety. The oligosaccharide attached to Asn 297 may include
various
carbohydrates such as mannose, N-acetyl glucosamine (G1cNAc), galactose, and
sialic
acid, as well as a fucose attached to a GlcNAc in the "stem" of the
biantennary
oligosaccharide structure.
[230] In some embodiments, the oligosaccharide attached to Asn 297 is
modified to create
ABPs having altered ADCC. In some embodiments, the oligosaccharide is altered
to
improve ADCC. In some embodiments, the oligosaccharide is altered to reduce
ADCC.
[231] In some aspects, an ABP provided herein comprises an IgG1 domain with
reduced
fucose content at position Asn 297 compared to a naturally occurring IgG1
domain. Such
Fc domains are known to have improved ADCC. See Shields et al., I Biol. Chem.,
2002,
277:26733-26740, incorporated by reference in its entirety. In some aspects,
such ABPs do
not comprise any fucose at position Asn 297. The amount of fucose may be
determined
using any suitable method, for example as described in WO 2008/077546,
incorporated by
reference in its entirety.
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[232] In some embodiments, an ABP provided herein comprises a bisected
oligosaccharide,
such as a biantennary oligosaccharide attached to the Fc region of the ABP
that is bisected
by GlcNAc. Such ABP variants may have reduced fucosylation and/or improved
ADCC
function. Examples of such ABP variants are described, for example, in WO
2003/011878;
U.S. Pat. No. 6,602,684; and U.S. Pat. Pub. No. 2005/0123546; each of which is
incorporated by reference in its entirety.
[233] Other illustrative glycosylation variants are described, for example,
in U.S. Pat. Pub.
Nos. 2003/0157108, 2004/0093621, 2003/0157108, 2003/0115614, 2002/0164328,
2004/0093621, 2004/0132140, 2004/0110704, 2004/0110282, 2004/0109865;
International
Pat. Pub. Nos. 2000/61739, 2001/29246, 2003/085119, 2003/084570, 2005/035586,
2005/035778; 2005/053742, 2002/031140; Okazaki et al., 1 Mol. Biol., 2004,
336:1239-
1249; and Yamane-Ohnuki et al., Biotech. Bioeng., 2004, 87: 614-622; each of
which is
incorporated by reference in its entirety.
[234] In some embodiments, an ABP provided herein comprises an Fc region
with at least
one galactose residue in the oligosaccharide attached to the Fc region. Such
ABP variants
may have improved CDC function. Examples of such ABP variants are described,
for
example, in WO 1997/30087; WO 1998/58964; and WO 1999/22764; each of which his
incorporated by reference in its entirety.
[235] Examples of cell lines capable of producing defucosylated ABPs
include Lec13 CHO
cells, which are deficient in protein fucosylation (see Ripka et al., Arch.
Biochem.
Biophys., 1986, 249:533-545; U.S. Pat. Pub. No. 2003/0157108; WO 2004/056312;
each
of which is incorporated by reference in its entirety), and knockout cell
lines, such as
alpha-1,6-fucosyltransferase gene or FUT8 knockout CHO cells (see Yamane-
Ohnuki et
al., Biotech. Bioeng., 2004, 87: 614-622; Kanda et al., Biotechnol. Bioeng.,
2006, 94:680-
688; and WO 2003/085107; each of which is incorporated by reference in its
entirety).
[236] In some embodiments, an ABP provided herein is an aglycosylated ABP.
An
aglycosylated ABP can be produced using any method known in the art or
described
herein. In some aspects, an aglycosylated ABP is produced by modifying the ABP
to
remove all glycosylation sites. In some aspects, the glycosylation sites are
removed only
from the Fc region of the ABP. In some aspects, an aglycosylated ABP is
produced by
expressing the ABP in an organism that is not capable of glycosylation, such
as E. coil, or
by expressing the ABP in a cell-free reaction mixture.
[237] In some embodiments, an ABP provided herein has a constant region
with reduced
effector function compared to a native IgG1 antibody. In some embodiments, the
affinity
of a constant region of an Fc region of an ABP provided herein for Fc receptor
is less than
the affinity of a native IgG1 constant region for such Fc receptor.
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1.6. Fc Region Amino Acid Sequence Variants
[238] In certain embodiments, an ABP provided herein comprises an Fc region
with one or
more amino acid substitutions, insertions, or deletions in comparison to a
naturally
occurring Fc region. In some aspects, such substitutions, insertions, or
deletions yield
ABPs with altered stability, glycosylation, or other characteristics. In some
aspects, such
substitutions, insertions, or deletions yield aglycosylated ABPs.
[239] In some aspects, the Fc region of an ABP provided herein is modified
to yield an
ABP with altered affinity for an Fc receptor, or an ABP that is more
immunologically
inert. In some embodiments, the ABP variants provided herein possess some, but
not all,
effector functions. Such ABPs may be useful, for example, when the half-life
of the ABP
is important in vivo, but when certain effector functions (e.g., complement
activation and
ADCC) are unnecessary or deleterious.
[240] In some embodiments, the Fc region of an ABP provided herein is a human
IgG4 Fc
region comprising the hinge stabilizing mutation 5228P or L235E. In some
embodiment,
the IgG4 Fc region comprises the hinge stabilizing mutations 5228P and L235E.
See
Aalberse et al., Immunology, 2002, 105:9-19, incorporated by reference in its
entirety. In
some embodiments, the IgG4 Fc region comprises one or more of the following
mutations:
E233P, F234V, and L235A. See Armour et al., Mol. Immunol., 2003, 40:585-593,
incorporated by reference in its entirety. In some embodiments, the IgG4 Fc
region
comprises a deletion at position G236.
[241] In some embodiments, the Fc region of an ABP provided herein is a
human IgG1 Fc
region comprising one or more mutations to reduce Fc receptor binding. In some
aspects,
the one or more mutations are in residues selected from S228 (e.g., 5228A),
L234 (e.g.,
L234A), L235 (e.g., L235A), D265 (e.g., D265A), and N297 (e.g., N297A). In
some
aspects, the ABP comprises a PVA236 mutation. PVA236 means that the amino acid
sequence ELLG, from amino acid position 233 to 236 of IgG1 or EFLG of IgG4, is
replaced by PVA. See U.S. Pat. Pub. No. 2013/0065277, incorporated by
reference in its
entirety.
[242] In some embodiments, the Fc region of an ABP provided herein is
modified as
described in Armour et al., Eur. I Immunol., 1999, 29:2613-2624; WO
1999/058572;
and/or U.K. Pat. App. No. 98099518; each of which is incorporated by reference
in its
entirety.
[243] In some embodiments, the Fc region of an ABP provided herein is a human
IgG2 Fc
region comprising one or more of mutations A3 30S and P33 1S.
[244] In some embodiments, the Fc region of an ABP provided herein has an
amino acid
substitution at one or more positions selected from 238, 265, 269, 270, 297,
327 and 329.
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See U.S. Pat. No. 6,737,056, incorporated by reference in its entirety. Such
Fc mutants
include Fe mutants with substitutions at two or more of amino acid positions
265, 269,
270, 297 and 327, including the so-called "DANA" Fe mutant with substitution
of residues
265 and 297 with alanine. See U.S. Pat. No. 7,332,581, incorporated by
reference in its
entirety. In some embodiments, the ABP comprises an alanine at amino acid
position 265.
In some embodiments, the ABP comprises an alanine at amino acid position 297.
[245] In certain embodiments, an ABP provided herein comprises an Fe region
with one or
more amino acid substitutions which improve ADCC, such as a substitution at
one or more
of positions 298, 333, and 334 of the Fe region. In some embodiments, an ABP
provided
herein comprises an Fe region with one or more amino acid substitutions at
positions 239,
332, and 330, as described in Lazar et al., Proc. Natl. Acad. Sci. USA,
2006,103:4005-
4010, incorporated by reference in its entirety.
[246] In some embodiments, an ABP provided herein comprises one or more
alterations
that improves or diminishes Clq binding and/or CDC. See U.S. Pat. No.
6,194,551; WO
99/51642; and Idusogie et al., I Immunol., 2000, 164:4178-4184; each of which
is
incorporated by reference in its entirety.
[247] In some embodiments, an ABP provided herein comprises one or more
alterations to
increase half-life. ABPs with increased half-lives and improved binding to the
neonatal Fe
receptor (FcRn) are described, for example, in Hinton et al., I Immunol.,
2006, 176:346-
356; and U.S. Pat. Pub. No. 2005/0014934; each of which is incorporated by
reference in
its entirety. Such Fe variants include those with substitutions at one or more
of Fe region
residues: 238, 250, 256, 265, 272, 286, 303, 305, 307, 311, 312, 314, 317,
340, 356, 360,
362, 376, 378, 380, 382, 413, 424, 428, and 434 of an IgG.
[248] In some embodiments, an ABP provided herein comprises one or more Fe
region
variants as described in U.S. Pat. Nos. 7,371,826 5,648,260, and 5,624,821;
Duncan and
Winter, Nature, 1988, 322:738-740; and WO 94/29351; each of which is
incorporated by
reference in its entirety.
1.7. Cysteine Engineered Antigen-Binding Protein Variants
[249] In certain embodiments, provided herein are cysteine engineered ABPs,
also known
as "thioMAbs," in which one or more residues of the ABP are substituted with
cysteine
residues. In particular embodiments, the substituted residues occur at solvent
accessible
sites of the ABP. By substituting such residues with cysteine, reactive thiol
groups are
introduced at solvent accessible sites of the ABP and may be used to conjugate
the ABP to
other moieties, such as drug moieties or linker-drug moieties, for example, to
create an
immunoconjugate.
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[250] In certain embodiments, any one or more of the following residues may
be substituted
with cysteine: V205 of the light chain; A118 of the heavy chain Fc region; and
S400 of the
heavy chain Fc region. Cysteine engineered ABPs may be generated as described,
for
example, in U.S. Pat. No. 7,521,541, which is incorporated by reference in its
entirety.
1.7.1.Immunoconjugates
1.7.1.1. Antigen-Binding Protein-Polymer Conjugates
[251] In some embodiments, an ABP provided herein is derivatized by
conjugation with a
polymer. Any suitable polymer may be conjugated to the ABP.
[252] In some embodiments, the polymer is a water soluble polymer.
Illustrative examples
of water soluble polymers include polyethylene glycol (PEG), copolymers of
ethylene
glycol/propylene glycol, carboxymethylcellulose, dextran, polyvinyl alcohol,
polyvinyl
pyrrolidone, poly-1,3-dioxolane, poly-1,3,6-trioxane, ethylene/maleic
anhydride
copolymer, polyaminoacids (either homopolymers or random copolymers), poly(n-
vinyl
pyrrolidone)-co-polyethylene glycol, propropylene glycol homopolymers,
polypropylene
oxide/ethylene oxide co-polymers, polyoxyethylated polyols (e.g., glycerol),
polyvinyl
alcohol, and mixtures thereof In some aspects, polyethylene glycol
propionaldehyde may
have advantages in manufacturing due to its stability in water.
[253] The polymer may be of any molecular weight, and may be branched or
unbranched.
The number of polymers attached to each ABP may vary, and if more than one
polymer is
attached, they may be the same polymer or different polymers. In general, the
number
and/or type of polymers used for derivatization can be determined based on
considerations
including the particular properties or functions of the ABP to be improved and
the
intended use of the ABP.
1.7.1.2. Antigen-Binding Protein-Drug Conjugates
[254] In some embodiments, the ABPs provided herein are conjugated to one
or more
therapeutic agents. Any suitable therapeutic agent may be conjugated to the
ABP.
Exemplary therapeutic agents include cytokines, chemokines, and other agents
that induce
a desired T cell activity, such as GITRL, OX4OL, 4-1BBL, TNF-alpha, IL-2, IL-
15 fusion,
CXCL9, CXCL10, IL-10 trap, IL-27 trap, and IL-35 trap. Cytokine traps and
their use are
known in the art and described, for example, in Economides et al., Nature
Medicine, 2003,
9:47-52, incorporated by reference in its entirety.
Methods of Making GITR Antigen-Binding Proteins
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1.8. GITR Antigen Preparation
[255] The GITR antigen used for isolation of the ABPs provided herein may
be intact GITR
or a fragment of GITR. The GITR antigen may be in the form of an isolated
protein or a
protein expressed on the surface of a cell.
[256] In some embodiments, the GITR antigen is a non-naturally occurring
variant of
GITR, such as a GITR protein having an amino acid sequence or post-
translational
modification that does not occur in nature.
[257] In some embodiments, the GITR antigen is truncated by removal of, for
example,
intracellular or membrane-spanning sequences, or signal sequences. In some
embodiments,
the GITR antigen is fused at its C-terminus to a human IgG1 Fc domain or a
polyhistidine
tag.
1.9. Methods of Making Monoclonal Antibodies
[258] Monoclonal antibodies may be obtained, for example, using the
hybridoma method
first described by Kohler et al., Nature, 1975, 256:495-497 (incorporated by
reference in
its entirety), and/or by recombinant DNA methods (see e.g., U.S. Patent No.
4,816,567,
incorporated by reference in its entirety). Monoclonal antibodies may also be
obtained, for
example, using phage or yeast-based libraries. See e.g., U.S. Patent Nos.
8,258,082 and
8,691,730, each of which is incorporated by reference in its entirety.
[259] In the hybridoma method, a mouse or other appropriate host animal is
immunized to
elicit lymphocytes that produce or are capable of producing antibodies that
will
specifically bind to the protein used for immunization. Alternatively,
lymphocytes may be
immunized in vitro. Lymphocytes are then fused with myeloma cells using a
suitable
fusing agent, such as polyethylene glycol, to form a hybridoma cell. See
Goding J.W.,
Monoclonal Antibodies: Principles and Practice 3rd ed. (1986) Academic Press,
San
Diego, CA, incorporated by reference in its entirety.
[260] The hybridoma cells are seeded and grown in a suitable culture medium
that contains
one or more substances that inhibit the growth or survival of the unfused,
parental
myeloma cells. For example, if the parental myeloma cells lack the enzyme
hypoxanthine
guanine phosphoribosyl transferase (HGPRT or HPRT), the culture medium for the
hybridomas typically will include hypoxanthine, aminopterin, and thymidine
(HAT
medium), which substances prevent the growth of HGPRT-deficient cells.
[261] Useful myeloma cells are those that fuse efficiently, support stable
high-level
production of antibody by the selected antibody-producing cells, and are
sensitive media
conditions, such as the presence or absence of HAT medium. Among these,
preferred
myeloma cell lines are murine myeloma lines, such as those derived from MOP-21
and
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MC-11 mouse tumors (available from the Salk Institute Cell Distribution
Center, San
Diego, CA), and SP-2 or X63-Ag8-653 cells (available from the American Type
Culture
Collection, Rockville, MD). Human myeloma and mouse-human heteromyeloma cell
lines
also have been described for the production of human monoclonal antibodies.
See e.g.,
Kozbor, I Immunol., 1984, 133:3001, incorporated by reference in its entirety.
[262] After the identification of hybridoma cells that produce antibodies
of the desired
specificity, affinity, and/or biological activity, selected clones may be
subcloned by
limiting dilution procedures and grown by standard methods. See Goding, supra.
Suitable
culture media for this purpose include, for example, D-MEM or RPMI-1640
medium. In
addition, the hybridoma cells may be grown in vivo as ascites tumors in an
animal.
[263] DNA encoding the monoclonal antibodies may be readily isolated and
sequenced
using conventional procedures (e.g., by using oligonucleotide probes that are
capable of
binding specifically to genes encoding the heavy and light chains of the
monoclonal
antibodies). Thus, the hybridoma cells can serve as a useful source of DNA
encoding
antibodies with the desired properties. Once isolated, the DNA may be placed
into
expression vectors, which are then transfected into host cells such as
bacteria (e.g., E.
coil), yeast (e.g., Saccharomyces or Pichia sp.), COS cells, Chinese hamster
ovary (CHO)
cells, or myeloma cells that do not otherwise produce antibody, to produce the
monoclonal
antibodies.
1.10. Methods of Making Chimeric Antibodies
[264] Illustrative methods of making chimeric antibodies are described, for
example, in
U.S. Pat. No. 4,816,567; and Morrison et al., Proc. Natl. Acad. Sci. USA,
1984, 81:6851-
6855; each of which is incorporated by reference in its entirety. In some
embodiments, a
chimeric antibody is made by using recombinant techniques to combine a non-
human
variable region (e.g., a variable region derived from a mouse, rat, hamster,
rabbit, or non-
human primate, such as a monkey) with a human constant region.
1.11. Methods of Making Humanized Antibodies
[265] Humanized antibodies may be generated by replacing most, or all, of
the structural
portions of a non-human monoclonal antibody with corresponding human antibody
sequences. Consequently, a hybrid molecule is generated in which only the
antigen-
specific variable, or CDR, is composed of non-human sequence. Methods to
obtain
humanized antibodies include those described in, for example, Winter and
Milstein,
Nature, 1991, 349:293-299; Rader et al., Proc. Nat. Acad. Sci. USA., 1998,
95:8910-
8915; Steinberger et al., I Biol. Chem., 2000, 275:36073-36078; Queen et al.,
Proc. Natl.
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Acad. Sci. USA., 1989, 86:10029-10033; and U.S. Patent Nos. 5,585,089,
5,693,761,
5,693,762, and 6,180,370; each of which is incorporated by reference in its
entirety.
1.12. Methods of making Human Antibodies
[266] Human antibodies can be generated by a variety of techniques known in
the art, for
example by using transgenic animals (e.g., humanized mice). See, e.g.,
Jakobovits et al.,
Proc. Natl. Acad. Sci. USA., 1993, 90:2551; Jakobovits et al., Nature, 1993,
362:255-258;
Bruggermann et al., Year in Immuno., 1993, 7:33; and U.S. Patent Nos.
5,591,669,
5,589,369 and 5,545,807; each of which is incorporated by reference in its
entirety. Human
antibodies can also be derived from phage-display libraries (see e.g.,
Hoogenboom et al., I
Mol. Biol., 1991, 227:381-388; Marks et al., I Mol. Biol., 1991, 222:581-597;
and U.S.
Pat. Nos. 5,565,332 and 5,573,905; each of which is incorporated by reference
in its
entirety). Human antibodies may also be generated by in vitro activated B
cells (see e.g.,
U.S. Patent. Nos. 5,567,610 and 5,229,275, each of which is incorporated by
reference in
its entirety). Human antibodies may also be derived from yeast-based libraries
(see e.g.,
U.S. Patent No. 8,691,730, incorporated by reference in its entirety).
1.13. Methods of Making Antibody Fragments
[267] The antibody fragments provided herein may be made by any suitable
method,
including the illustrative methods described herein or those known in the art.
Suitable
methods include recombinant techniques and proteolytic digestion of whole
antibodies.
Illustrative methods of making antibody fragments are described, for example,
in Hudson
et al., Nat. Med., 2003, 9:129-134, incorporated by reference in its entirety.
Methods of
making scFy antibodies are described, for example, in Pliickthun, in The
Pharmacology of
Monoclonal Antibodies, vol. 113, Rosenburg and Moore eds., Springer-Verlag,
New York,
pp. 269-315 (1994); WO 93/16185; and U.S. Pat. Nos. 5,571,894 and 5,587,458;
each of
which is incorporated by reference in its entirety.
1.14. Methods of Making Alternative Scaffolds
[268] The alternative scaffolds provided herein may be made by any suitable
method,
including the illustrative methods described herein or those known in the art.
For example,
methods of preparing AdnectinsTm are described in Emanuel et al., mAbs, 2011,
3:38-48,
incorporated by reference in its entirety. Methods of preparing iMabs are
described in U.S.
Pat. Pub. No. 2003/0215914, incorporated by reference in its entirety. Methods
of
preparing Anticalins are described in Vogt and Skerra, Chem. Biochem., 2004,
5:191-199,
incorporated by reference in its entirety. Methods of preparing Kunitz domains
are
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described in Wagner etal., Biochem. & Biophys. Res. Comm., 1992, 186:118-1145,
incorporated by reference in its entirety. Methods of preparing thioredoxin
peptide
aptamers are provided in Geyer and Brent, Meth. Enzymol., 2000, 328:171-208,
incorporated by reference in its entirety. Methods of preparing Affibodies are
provided in
Fernandez, Curr. Opinion in Biotech., 2004, 15:364-373, incorporated by
reference in its
entirety. Methods of preparing DARPins are provided in Zahnd etal., I Mol.
Biol., 2007,
369:1015-1028, incorporated by reference in its entirety. Methods of preparing
Affilins are
provided in Ebersbach etal., I Mol. Biol., 2007, 372:172-185, incorporated by
reference
in its entirety. Methods of preparing Tetranectins are provided in Graversen
et al., I Biol.
Chem., 2000, 275:37390-37396, incorporated by reference in its entirety.
Methods of
preparing Avimers are provided in Silverman etal., Nature Biotech., 2005,
23:1556-1561,
incorporated by reference in its entirety. Methods of preparing Fynomers are
provided in
Silacci etal., I Biol. Chem., 2014, 289:14392-14398, incorporated by reference
in its
entirety.
[269] Further information on alternative scaffolds is provided in Binz et
al., Nat.
Biotechnol., 2005 23:1257-1268; and Skerra, Current Opin. in Biotech., 2007
18:295-304,
each of which is incorporated by reference in its entirety.
1.15. Methods of Making Multispecific ABPs
[270] The multispecific or multivalent monospecific ABPs provided herein
may be made
by any suitable method, including the illustrative methods described herein or
those known
in the art. Methods of making common light chain antibodies are described in
Merchant et
al., Nature Biotechnol., 1998, 16:677-681, incorporated by reference in its
entirety.
Methods of making tetravalent bispecific antibodies are described in Coloma
and
Morrison, Nature Biotechnol., 1997, 15:159-163, incorporated by reference in
its entirety.
Methods of making hybrid immunoglobulins are described in Milstein and Cuello,
Nature,
1983, 305:537-540; and Staerz and Bevan, Proc. Natl. Acad. Sci. USA, 1986,
83:1453-
1457; each of which is incorporated by reference in its entirety. Methods of
making
immunoglobulins with knobs-into-holes modification are described in U.S. Pat.
No.
5,731,168, incorporated by reference in its entirety. Methods of making
immunoglobulins
with electrostatic modifications are provided in WO 2009/089004, incorporated
by
reference in its entirety. Methods of making multivalent (e.g., tetravalent)
mono specific
antibodies are described in Miller et al., 2003, U.S. Patent No. 8,722,859,
incorporated by
reference in its entirety. Methods of making bispecific single chain
antibodies are
described in Traunecker etal., EiVIBO 1, 1991, 10:3655-3659; and Gruber etal.,
I
Immunol., 1994, 152:5368-5374; each of which is incorporated by reference in
its entirety.
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Methods of making single-chain antibodies, whose linker length may be varied,
are
described in U.S. Pat. Nos. 4,946,778 and 5,132,405, each of which is
incorporated by
reference in its entirety. Methods of making diabodies are described in
Hollinger et al.,
Proc. Natl. Acad. Sci. USA, 1993, 90:6444-6448, incorporated by reference in
its entirety.
Methods of making triabodies and tetrabodies are described in Todorovska et
al., I
Immunol. Methods, 2001, 248:47-66, incorporated by reference in its entirety.
Methods of
making trispecific F(ab')3 derivatives are described in Tuft et al. I
Immunol., 1991,
147:60-69, incorporated by reference in its entirety. Methods of making cross-
linked
antibodies are described in U.S. Patent No. 4,676,980; Brennan et al.,
Science, 1985,
229:81-83; Staerz, et al. Nature, 1985, 314:628-631; and EP 0453082; each of
which is
incorporated by reference in its entirety. Methods of making antigen-binding
domains
assembled by leucine zippers are described in Kostelny et al., I Immunol.,
1992,
148:1547-1553, incorporated by reference in its entirety. Methods of making
ABPs via the
DNL approach are described in U.S. Pat. Nos. 7,521,056; 7,550,143; 7,534,866;
and
7,527,787; each of which is incorporated by reference in its entirety. Methods
of making
hybrids of antibody and non-antibody molecules are described in WO 93/08829,
incorporated by reference in its entirety, for examples of such ABPs. Methods
of making
DAF antibodies are described in U.S. Pat. Pub. No. 2008/0069820, incorporated
by
reference in its entirety. Methods of making ABPs via reduction and oxidation
are
described in Carlring et al., PLoS One, 2011, 6:e22533, incorporated by
reference in its
entirety. Methods of making DVD-IgsTM are described in U.S. Pat. No.
7,612,181,
incorporated by reference in its entirety. Methods of making DARTsTm are
described in
Moore et al., Blood, 2011, 117:454-451, incorporated by reference in its
entirety. Methods
of making DuoBodies are described in Labrijn et al., Proc. Natl. Acad. Sci.
USA, 2013,
110:5145-5150; Gramer et al., mAbs, 2013, 5:962-972; and Labrijn et al.,
Nature
Protocols, 2014, 9:2450-2463; each of which is incorporated by reference in
its entirety.
Methods of making antibodies comprising scFvs fused to the C-terminus of the
CH3 from
an IgG are described in Coloma and Morrison, Nature Biotechnol., 1997, 15:159-
163,
incorporated by reference in its entirety. Methods of making antibodies in
which a Fab
molecule is attached to the constant region of an immunoglobulin are described
in Miler et
al., I Immunol., 2003, 170:4854-4861, incorporated by reference in its
entirety. Methods
of making CovX-Bodies are described in Doppalapudi et al., Proc. Natl. Acad.
Sci. USA,
2010, 107:22611-22616, incorporated by reference in its entirety. Methods of
making Fcab
antibodies are described in Wozniak-Knopp et al., Protein Eng. Des. Se!.,
2010, 23:289-
297, incorporated by reference in its entirety. Methods of making TandAb
antibodies are
described in Kipriyanov et al., I Mol. Biol., 1999, 293:41-56 and Zhukovsky et
al., Blood,
2013, 122:5116, each of which is incorporated by reference in its entirety.
Methods of
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making tandem Fabs are described in WO 2015/103072, incorporated by reference
in its
entirety. Methods of making ZybodiesTm are described in LaFleur etal., mAbs,
2013,
5:208-218, incorporated by reference in its entirety.
[271] In another aspect is provided a method for producing an anti-human
GITR antibody
or an antigen-binding fragment thereof, comprising culturing host cell(s)
selected from the
group consisting of (a) to (c) below to express a tetravalent anti-human GITR
antibody or
an antigen-binding fragment thereof: (a) a host cell transformed with an
expression vector
comprising a polynucleotide comprising a base sequence encoding the heavy
chain
variable region of the anti-human GITR antibody or the antigen-binding
fragment thereof
provided herein and a polynucleotide comprising a base sequence encoding the
light chain
variable region of the antibody or the antigen-binding fragment thereof; (b) a
host cell
transformed with an expression vector comprising a polynucleotide comprising a
base
sequence encoding the heavy chain variable region of the anti-human GITR
antibody or
the antigen-binding fragment thereof provided herein and an expression vector
comprising
a polynucleotide comprising a base sequence encoding the light chain variable
region of
the antibody or the antigen-binding fragment thereof; and (c) a host cell
transformed with
an expression vector comprising a polynucleotide comprising a base sequence
encoding
the heavy chain variable region of the anti-human GITR antibody or the antigen-
binding
fragment thereof provided herein and a host cell transformed with an
expression vector
comprising a polynucleotide comprising a base sequence encoding the light
chain variable
region of the antibody or the antigen-binding fragment thereof.
[272] In another aspect is provided a method for producing an anti-human
GITR antibody,
comprising culturing host cell(s) selected from the group consisting of (a) to
(c) below to
express an anti-human GITR antibody: (a) a host cell transformed with an
expression
vector comprising a polynucleotide comprising a base sequence encoding the
heavy chain
of the anti-human GITR antibody provided herein and a polynucleotide
comprising a base
sequence encoding the light chain of the antibody; (b) a host cell transformed
with an
expression vector comprising a polynucleotide comprising a base sequence
encoding the
heavy chain of the anti-human GITR antibody provided herein and an expression
vector
comprising a polynucleotide comprising a base sequence encoding the light
chain of the
antibody; and (c) a host cell transformed with an expression vector
comprising a
polynucleotide comprising a base sequence encoding the heavy chain of the anti-
human
GITR antibody provided herein and a host cell transformed with an expression
vector
comprising a polynucleotide comprising a base sequence encoding the light
chain of the
antibody.
1.16. Methods of Making Variants
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[273] In some embodiments, an ABP provided herein is an affinity matured
variant of a
parent ABP, which may be generated, for example, using phage display-based
affinity
maturation techniques. Briefly, one or more CDR residues may be mutated and
the variant
ABPs, or portions thereof, displayed on phage and screened for affinity. Such
alterations
may be made in CDR "hotspots," or residues encoded by codons that undergo
mutation at
high frequency during the somatic maturation process (see Chowdhury, Methods
Mol.
Biol., 2008, 207:179-196, incorporated by reference in its entirety), and/or
residues that
contact the antigen. In some embodiments, affinity maturation can be used for
altering or
introducing species binding, i.e., an anti-mouse antibody may be engineered to
bind to
human and cynomolgus monkey versions of the same target antigen, etc.
[274] Any suitable method can be used to introduce variability into a
polynucleotide
sequence(s) encoding an ABP, including error-prone PCR, chain shuffling, and
oligonucleotide-directed mutagenesis such as trinucleotide-directed
mutagenesis (TRIM).
In some aspects, several CDR residues (e.g., 4-6 residues at a time) are
randomized. CDR
residues involved in antigen binding may be specifically identified, for
example, using
alanine scanning mutagenesis or modeling. CDR-H3 and CDR-L3 in particular are
often
targeted for mutation.
[275] The introduction of diversity into the variable regions and/or CDRs
can be used to
produce a secondary library. The secondary library is then screened to
identify ABP
variants with improved affinity. Affinity maturation by constructing and
reselecting from
secondary libraries has been described, for example, in Hoogenboom et al.,
Methods in
Molecular Biology, 2001, 178:1-37, incorporated by reference in its entirety.
1.17. Vectors, Host Cells, and Recombinant Methods
[276] Also provided are isolated nucleic acids encoding GITR ABPs, vectors
comprising
the nucleic acids, and host cells comprising the vectors and nucleic acids, as
well as
recombinant techniques for the production of the ABPs.
[277] In another aspect is provided a polynucleotide comprising a base
sequence encoding
the heavy chain variable region of the anti-human GITR antibody or the antigen-
binding
fragment thereof provided herein. In another aspect is provided a
polynucleotide
comprising a base sequence encoding the light chain variable region of the
anti-human
GTIR antibody or the antigen-binding fragment thereof provided herein.
[278] In another aspect is provided a polynucleotide comprising a base
sequence encoding
the heavy chain of the anti-human GITR antibody provided herein. In another
aspect is
provided a polynucleotide comprising a base sequence encoding the light chain
of the anti-
human GTIR antibody provided herein.
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[279] For recombinant production of an ABP, the nucleic acid(s) encoding it
may be
isolated and inserted into a replicable vector for further cloning (i.e.,
amplification of the
DNA) or expression. In some aspects, the nucleic acid may be produced by
homologous
recombination, for example as described in U.S. Patent No. 5,204,244,
incorporated by
reference in its entirety.
[280] In another aspect is provided an expression vector comprising (a) a
polynucleotide
comprising a base sequence encoding the heavy chain variable region of the
anti-human
GITR antibody or the antigen-binding fragment thereof provided herein and/or
(b) a
polynucleotide comprising a base sequence encoding the light chain variable
region of the
anti-human GITR antibody or the antigen-binding fragment thereof
[281] In another aspect is provided an expression vector comprising (a) a
polynucleotide
comprising a base sequence encoding the heavy chain of the anti-human GITR
antibody
provided herein and/or (b) a polynucleotide comprising a base sequence
encoding the light
chain of the anti-human GITR antibody.
[282] Many different vectors are known in the art. The vector components
generally include
one or more of the following: a signal sequence, an origin of replication, one
or more
marker genes, an enhancer element, a promoter, and a transcription termination
sequence,
for example as described in U.S. Patent No. 5,534,615, incorporated by
reference in its
entirety.
[283] Illustrative examples of suitable host cells are provided below.
These host cells are
not meant to be limiting, and any suitable host cell may be used to produce
the ABPs
provided herein.
[284] In another aspect is provided a host cell transformed with an
expression vector
selected from the group consisting of (a) to (d): (a) a host cell transformed
with an
expression vector comprising a polynucleotide comprising a base sequence
encoding the
heavy chain variable region of the anti-human GITR antibody or the antigen-
binding
fragment thereof provided herein, and a polynucleotide comprising a base
sequence
encoding the light chain variable region of the antibody or the antigen-
binding fragment
thereof; (b) a host cell transformed with an expression vector comprising a
polynucleotide
comprising a base sequence encoding the heavy chain variable region of the
anti-human
GITR antibody or the antigen-biding fragment thereof provided herein and an
expression
vector comprising a polynucleotide comprising a base sequence encoding the
light chain
variable region of the antibody or the antigen-binding fragment thereof, (c) a
host cell
transformed with an expression vector comprising a polynucleotide comprising a
base
sequence encoding the heavy chain variable region of the anti-human GITR
antibody or
the antigen-binding fragment thereof provided herein; and (d) a host cell
transformed with
an expression vector comprising a polynucleotide comprising a base sequence
encoding
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the light chain variable region of the anti-human GITR antibody or the antigen-
binding
fragment thereof provided herein.
[285] In another aspect is provided host cell transformed with an
expression vector selected
from the group consisting of (a) to (d): (a) a host cell transformed with an
expression
vector comprising a polynucleotide comprising a base sequence encoding the
heavy chain
of the anti-human GITR antibody provided herein and a polynucleotide
comprising a base
sequence encoding the light chain of the antibody; (b) a host cell transformed
with an
expression vector comprising a polynucleotide comprising a base sequence
encoding the
heavy chain of the anti-human GITR antibody provided herein and an expression
vector
comprising a polynucleotide comprising a base sequence encoding the light
chain of the
antibody; (c) a host cell transformed with an expression vector comprising a
polynucleotide comprising a base sequence encoding the heavy chain of the anti-
human
GITR antibody provided herein; and (d) a host cell transformed with an
expression vector
comprising a polynucleotide comprising a base sequence encoding the light
chain of the
anti-human GITR antibody provided herein.
[286] Suitable host cells include any prokaryotic (e.g., bacterial), lower
eukaryotic (e.g.,
yeast), or higher eukaryotic (e.g., mammalian) cells. Suitable prokaryotes
include
eubacteria, such as Gram-negative or Gram-positive organisms, for example,
Enterobacteriaceae such as Escherichia (E. coil), Enterobacter, , Erwinia,
Klebsiella,
Proteus, Salmonella (S. typhimurium), Serratia (S. marcescans), Shigella,
Bacilli (B.
subtilis and B. licheniformis), Pseudomonas P. aeruginosa), and Streptomyces.
One useful
E. coil cloning host is E. coil 294, although other strains such as E. coil B,
E. coil X1776,
and E. coil W3110 are also suitable.
[287] In addition to prokaryotes, eukaryotic microbes such as filamentous
fungi or yeast are
also suitable cloning or expression hosts for GITR ABP -encoding vectors.
Saccharomyces
cerevisiae, or common baker's yeast, is a commonly used lower eukaryotic host
microorganism. However, a number of other genera, species, and strains are
available and
useful, such as Schizosaccharomyces pombe, Kluyveromyces (K lactis, K
fragilis, K
bulgaricus K wickeramii, K waltii, K drosophilarum, K thermotolerans, and K
marxianus), Yarrowia, Pichia pastor's, Candida (C. albicans), Trichoderma
reesia,
Neurospora crassa, Schwanniomyces (S. occidentalis), and filamentous fungi
such as, for
example Penicillium, Tolypocladium, and Aspergillus (A. nidulans and A.
niger).
[288] Useful mammalian host cells include COS-7 cells, HEK293 cells; baby
hamster
kidney (BHK) cells; Chinese hamster ovary (CHO); mouse sertoli cells; African
green
monkey kidney cells (VERO-76), and the like.
[289] The host cells used to produce the GITR ABP of this invention may be
cultured in a
variety of media. Commercially available media such as, for example, Ham's
F10,
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Minimal Essential Medium (MEM), RPMI-1640, and Dulbecco's Modified Eagle's
Medium (DMEM) are suitable for culturing the host cells. In addition, any of
the media
described in Ham et al., Meth. Enz., 1979, 58:44; Barnes et al., Anal.
Biochem., 1980,
102:255; and U.S. Patent Nos. 4,767,704, 4,657,866, 4,927,762, 4,560,655, and
5,122,469;
or WO 90/03430 and WO 87/00195 may be used. Each of the foregoing references
is
incorporated by reference in its entirety.
[290] Any of these media may be supplemented as necessary with hormones and/or
other
growth factors (such as insulin, transferrin, or epidermal growth factor),
salts (such as
sodium chloride, calcium, magnesium, and phosphate), buffers (such as HEPES),
nucleotides (such as adenosine and thymidine), antibiotics, trace elements
(defined as
inorganic compounds usually present at final concentrations in the micromolar
range), and
glucose or an equivalent energy source. Any other necessary supplements may
also be
included at appropriate concentrations that would be known to those skilled in
the art.
[291] The culture conditions, such as temperature, pH, and the like, are
those previously
used with the host cell selected for expression, and will be apparent to the
ordinarily
skilled artisan.
[292] When using recombinant techniques, the ABP can be produced
intracellularly, in the
periplasmic space, or directly secreted into the medium. If the ABP is
produced
intracellularly, as a first step, the particulate debris, either host cells or
lysed fragments, is
removed, for example, by centrifugation or ultrafiltration. For example,
Carter et al.
(Bio/Technology, 1992, 10:163-167, incorporated by reference in its entirety)
describes a
procedure for isolating ABPs which are secreted to the periplasmic space of E.
coli.
Briefly, cell paste is thawed in the presence of sodium acetate (pH 3.5),
EDTA, and
phenylmethylsulfonylfluoride (PMSF) over about 30 min. Cell debris can be
removed by
centrifugation.
[293] In some embodiments, the ABP is produced in a cell-free system. In
some aspects,
the cell-free system is an in vitro transcription and translation system as
described in Yin et
al., mAbs, 2012, 4:217-225, incorporated by reference in its entirety. In some
aspects, the
cell-free system utilizes a cell-free extract from a eukaryotic cell or from a
prokaryotic
cell. In some aspects, the prokaryotic cell is E. coli. Cell-free expression
of the ABP may
be useful, for example, where the ABP accumulates in a cell as an insoluble
aggregate, or
where yields from periplasmic expression are low.
[294] Where the ABP is secreted into the medium, supernatants from such
expression
systems are generally first concentrated using a commercially available
protein
concentration filter, for example, an Amicon or Millipore Pellcon
ultrafiltration unit. A
protease inhibitor such as PMSF may be included in any of the foregoing steps
to inhibit
proteolysis and antibiotics may be included to prevent the growth of
adventitious
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contaminants.
[295] The ABP composition prepared from the cells can be purified using,
for example,
hydroxylapatite chromatography, gel electrophoresis, dialysis, and affinity
chromatography, with affinity chromatography being a particularly useful
purification
technique. The suitability of protein A as an affinity ligand depends on the
species and
isotype of any immunoglobulin Fc domain that is present in the ABP. Protein A
can be
used to purify ABPs that comprise human yl, y2, or y4 heavy chains (Lindmark
et al.,
Immunol. Meth., 1983, 62:1-13, incorporated by reference in its entirety).
Protein G is
useful for all mouse isotypes and for human y3 (Guss et al., EiVIBO 1, 1986,
5:1567-1575,
incorporated by reference in its entirety).
[296] The matrix to which the affinity ligand is attached is most often
agarose, but other
matrices are available. Mechanically stable matrices such as controlled pore
glass or
poly(styrenedivinyl)benzene allow for faster flow rates and shorter processing
times than
can be achieved with agarose. Where the ABP comprises a CH3 domain, the
BakerBond
ABX resin is useful for purification.
[297] Other techniques for protein purification, such as fractionation on
an ion-exchange
column, ethanol precipitation, Reverse Phase HPLC, chromatography on silica,
chromatography on heparin Sepharose , chromatofocusing, SDS-PAGE, and ammonium
sulfate precipitation are also available, and can be applied by one of skill
in the art.
[298] Following any preliminary purification step(s), the mixture
comprising the ABP of
interest and contaminants may be subjected to low pH hydrophobic interaction
chromatography using an elution buffer at a pH between about 2.5 to about 4.5,
generally
performed at low salt concentrations (e.g., from about 0 to about 0.25 M
salt).
Assays
[299] A variety of assays known in the art may be used to identify and
characterize the
GITR ABPs provided herein.
1.18. Binding, Competition, and Epitope Mapping Assays
[300] Specific antigen-binding activity of the ABPs provided herein may be
evaluated by
any suitable method, including using SPR, BLI, and RIA, as described elsewhere
in this
disclosure. Additionally, antigen-binding activity may be evaluated by ELISA
assays and
Western blot assays.
[301] Assays for measuring competition between two ABPs, or an ABP and another
molecule (e.g., GITRL) are described elsewhere in this disclosure and, for
example, in
Harlow and Lane, Antibodies: A Laboratory Manual ch.14, 1988, Cold Spring
Harbor
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Laboratory, Cold Spring Harbor, N.Y, incorporated by reference in its
entirety.
[302] Assays for mapping the epitopes to which the ABPs provided herein
bind are
described, for example, in Morris "Epitope Mapping Protocols," in Methods in
Molecular
Biology vol. 66, 1996, Humana Press, Totowa, N.J., incorporated by reference
in its
entirety. In some embodiments, the epitope is determined by peptide
competition. In some
embodiments, the epitope is determined by mass spectrometry. In some
embodiments, the
epitope is determined by crystallography.
1.19. GITR Agonism Assays
[303] In some embodiments, the ABPs provided herein are screened to
identify or
characterize ABPs with agonistic activity against GITR. Any suitable assay may
be used to
identify or characterize such ABPs. In some aspects, the assay measures the
amount of a
cytokine secreted by an effector T cell after contacting the effector T cell
with an ABP
provided herein. In some aspects, the cytokine is selected from IL-2Ra, IL-2,
IL-8, IFNy,
and combinations thereof In some aspects, the cytokine is selected from
sCD40L, VEGF,
TNF-I3, TNF-a, TGF-a, RANTES, PDGF-AB/BB, PDGF-AA, MIP-113, MIP-la, MDC
(CCL22), MCP-3, MCP-1, IP-10, IL-17A, IL-15, IL-13, IL-12 (p70), IL-12 (p40),
IL-10,
IL-9, IL-8, IL-7, IL-6, IL-5, IL-4, IL-3, IL-2, IL-2Ra, IL-1RA, IL-113, IL-la,
IFNy, IFNa2,
GRO, GM-CSF, G-CSF, fractalkine, Flt-3 ligand, FGF-2, eotaxin, EGF, and
combinations
thereof
[304] In some embodiments, the effector cells are co-stimulated with an
agonist of CD3, to
promote the secretion of cytokines by the effector cell. In some aspects, the
CD3 agonist is
provided at a submaximal level.
[305] In some embodiments, functional assays are used such as the HT1080 or
Jurkat cell-
based assays described in more detail in Example 2. Additional assays are
described in
Wyzgol et al., J Immunol 2009; 183:1851-1861, incorporated herein by
reference.
[306] In some aspects, such assays may measure the proliferation of an
effector T cell after
contacting the effector T cell with an ABP provided herein. In some aspects,
proliferation
of the effector T cell is measured by dilution of a dye (e.g.,
carboxyfluorescein diacetate
succinimidyl ester; CFSE), by tritiated thymidine uptake, by luminescent cell
viability
assays, or by other assays known in the art.
[307] In some aspects, such assays may measure the differentiation,
cytokine production,
viability (e.g., survival), proliferation, or suppressive activity of a
regulatory T cell after
contacting the regulatory T cell with an ABP provided herein.
[308] In some aspects, such assays may measure the cytotoxic activity of an
NK cell after
contacting the NK cell with an ABP provided herein. In some aspects, the
cytotoxic
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activity of the NK cell is measured using a cytotoxicity assay that quantifies
NK-mediated
killing of target cells (e.g., a K562 cell line). See Jang et al., Ann. Cl/n.
Lab. Sci., 2012,
42:42-49, incorporated by reference in its entirety.
[309] Additional assays for measuring GITR agonism are described elsewhere
in this
disclosure, including in the Examples, and known in the art. A skilled person
can readily
select an appropriate assay for evaluating GITR agonism.
1.20. Assays for Effector Functions
[310] Effector function of the ABPs provided herein may be evaluated using
a variety of in
vitro and in vivo assays known in the art, including those described in
Ravetch and Kinet,
Annu. Rev. Immunol., 1991, 9:457-492; U.S. Pat. Nos. 5,500,362, 5,821,337;
Hellstrom et
al., Proc. Nat'l Acad. Sci. USA, 1986, 83:7059-7063; Hellstrom et al., Proc.
Nat'l Acad.
Sci. USA, 1985, 82:1499-1502; Bruggemann et al., 1 Exp. Med., 1987, 166:1351-
1361;
Clynes et al., Proc. Nat'l Acad. Sci. USA, 1998, 95:652-656; WO 2006/029879;
WO
2005/100402; Gazzano-Santoro et al., I Immunol. Methods, 1996, 202:163-171;
Cragg et
al., Blood, 2003, 101:1045-1052; Cragg et al. Blood, 2004, 103:2738-2743; and
Petkova et
al., Intl Immunol., 2006, 18:1759-1769; each of which is incorporated by
reference in its
entirety.
Pharmaceutical Compositions
[311] The ABPs provided herein can be formulated in any appropriate
pharmaceutical
composition and administered by any suitable route of administration. Suitable
routes of
administration include, but are not limited to, the intraarterial,
intradermal, intramuscular,
intraperitoneal, intravenous, nasal, parenteral, pulmonary, and subcutaneous
routes.
[312] In another aspect is provided a pharmaceutical composition comprising
plural kinds
of anti-human GITR antibodies or antigen-binding fragments thereof provided
herein. For
example, the pharmaceutical composition comprises an antibody or an antigen-
binding
fragment thereof, which does not undergo posttranslational modification and an
antibody
or an antigen-binding fragment thereof derived from posttranslational
modification of the
antibody or the antigen-binding fragment thereof.
[313] In one embodiment, the pharmaceutical composition comprises at least
two kinds of
anti-human GITR antibodies selected from (1) to (4): (1) an anti-human GITR
antibody
comprises two heavy chains consisting of SEQ ID NO: 7 and four light chains
consisting
of SEQ ID NO: 8; (2) an anti-human GITR antibody comprises two heavy chains
consisting of SEQ ID NO: 7, wherein the Q at position 1 is modified to
pyroglutamate and
four light chains consisting of SEQ ID NO: 8; (3) an anti-human GITR antibody
comprises
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two heavy chains consisting of the amino acid sequence ranging from Q at
position 1 to G
at position 686 of SEQ ID NO: 7, and four light chains consisting of SEQ ID
NO: 8; and
(4) an anti-human GITR antibody comprises two heavy chains consisting of the
amino acid
sequence ranging from Q at position 1 to G at position 686 of SEQ ID NO: 7,
wherein the
Q at position 1 is modified to pyroglutamate and four light chains of SEQ ID
NO: 8.
[314] In one embodiment, the pharmaceutical composition comprises an anti-
human GITR
antibody comprises two heavy chains consisting of SEQ ID NO: 7 and four light
chains
consisting of SEQ ID NO: 8; and an anti-human GITR antibody comprises two
heavy
chains consisting of the amino acid sequence ranging from Q at position 1 to G
at position
686 of SEQ ID NO: 7, wherein the Q at position 1 is modified to pyroglutamate
and four
light chains of SEQ ID NO: 8, and a pharmaceutically acceptable excipient.
[315] The pharmaceutical composition may comprise one or more
pharmaceutical
excipients. Any suitable pharmaceutical excipient may be used, and one of
ordinary skill in
the art is capable of selecting suitable pharmaceutical excipients.
Accordingly, the
pharmaceutical excipients provided below are intended to be illustrative, and
not limiting.
Additional pharmaceutical excipients include, for example, those described in
the
Handbook of Pharmaceutical Excipients, Rowe et al. (Eds.) 6th Ed. (2009),
incorporated
by reference in its entirety.
[316] In some embodiments, the pharmaceutical composition comprises an anti-
foaming
agent. Any suitable anti-foaming agent may be used. In some aspects, the anti-
foaming
agent is selected from an alcohol, an ether, an oil, a wax, a silicone, a
surfactant, and
combinations thereof In some aspects, the anti-foaming agent is selected from
a mineral
oil, a vegetable oil, ethylene bis stearamide, a paraffin wax, an ester wax, a
fatty alcohol
wax, a long chain fatty alcohol, a fatty acid soap, a fatty acid ester, a
silicon glycol, a
fluorosilicone, a polyethylene glycol-polypropylene glycol copolymer,
polydimethylsiloxane-silicon dioxide, ether, octyl alcohol, capryl alcohol,
sorbitan
trioleate, ethyl alcohol, 2-ethyl-hexanol, dimethicone, oleyl alcohol,
simethicone, and
combinations thereof
[317] In some embodiments, the pharmaceutical composition comprises a
cosolvent.
Illustrative examples of cosolvents include ethanol, poly(ethylene) glycol,
butylene glycol,
dimethylacetamide, glycerin, propylene glycol, and combinations thereof
[318] In some embodiments, the pharmaceutical composition comprises a
buffer.
Illustrative examples of buffers include acetate, borate, carbonate, lactate,
malate,
phosphate, citrate, hydroxide, diethanolamine, monoethanolamine, glycine,
methionine,
guar gum, monosodium glutamate, and combinations thereof.
[319] In some embodiments, the pharmaceutical composition comprises a
carrier or filler.
Illustrative examples of carriers or fillers include lactose, maltodextrin,
mannitol, sorbitol,
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chitosan, stearic acid, xanthan gum, guar gum, and combinations thereof
[320] In some embodiments, the pharmaceutical composition comprises a
surfactant.
Illustrative examples of surfactants include d-alpha tocopherol, benzalkonium
chloride,
benzethonium chloride, cetrimide, cetylpyridinium chloride, docusate sodium,
glyceryl
behenate, glyceryl monooleate, lauric acid, macrogol 15 hydroxystearate,
myristyl alcohol,
phospholipids, polyoxyethylene alkyl ethers, polyoxyethylene sorbitan fatty
acid esters,
polyoxyethylene stearates, polyoxylglycerides, sodium lauryl sulfate, sorbitan
esters,
vitamin E polyethylene(glycol) succinate, and combinations thereof.
[321] In some embodiments, the pharmaceutical composition comprises an anti-
caking
agent. Illustrative examples of anti-caking agents include calcium phosphate
(tribasic),
hydroxymethyl cellulose, hydroxypropyl cellulose, magnesium oxide, and
combinations
thereof
[322] Other excipients that may be used with the pharmaceutical
compositions include, for
example, albumin, antioxidants, antibacterial agents, antifungal agents,
bioabsorbable
polymers, chelating agents, controlled release agents, diluents, dispersing
agents,
dissolution enhancers, emulsifying agents, gelling agents, ointment bases,
penetration
enhancers, preservatives, solubilizing agents, solvents, stabilizing agents,
sugars, and
combinations thereof Specific examples of each of these agents are described,
for
example, in the Handbook of Pharmaceutical Excipients, Rowe et al. (Eds.) 6th
Ed.
(2009), The Pharmaceutical Press, incorporated by reference in its entirety.
[323] In some embodiments, the pharmaceutical composition comprises a
solvent. In some
aspects, the solvent is saline solution, such as a sterile isotonic saline
solution or dextrose
solution. In some aspects, the solvent is water for injection.
[324] In some embodiments, the pharmaceutical compositions are in a
particulate form,
such as a microparticle or a nanoparticle. Microparticles and nanoparticles
may be formed
from any suitable material, such as a polymer or a lipid. In some aspects, the
microparticles or nanoparticles are micelles, liposomes, or polymersomes.
[325] Further provided herein are anhydrous pharmaceutical compositions and
dosage
forms comprising an ABP, since water can facilitate the degradation of some
ABPs.
[326] Anhydrous pharmaceutical compositions and dosage forms provided herein
can be
prepared using anhydrous or low moisture containing ingredients and low
moisture or low
humidity conditions. Pharmaceutical compositions and dosage forms that
comprise lactose
and at least one active ingredient that comprises a primary or secondary amine
can be
anhydrous if substantial contact with moisture and/or humidity during
manufacturing,
packaging, and/or storage is expected.
[327] An anhydrous pharmaceutical composition should be prepared and stored
such that its
anhydrous nature is maintained. Accordingly, anhydrous compositions can be
packaged
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using materials known to prevent exposure to water such that they can be
included in
suitable formulary kits. Examples of suitable packaging include, but are not
limited to,
hermetically sealed foils, plastics, unit dose containers (e.g., vials),
blister packs, and strip
packs.
1.21. Parenteral Dosage Forms
[328] In certain embodiments, the ABPs provided herein are formulated as
parenteral
dosage forms. Parenteral dosage forms can be administered to subjects by
various routes
including, but not limited to, subcutaneous, intravenous (including infusions
and bolus
injections), intramuscular, and intraarterial. Because their administration
typically
bypasses subjects' natural defenses against contaminants, parenteral dosage
forms are
typically, sterile or capable of being sterilized prior to administration to a
subject.
Examples of parenteral dosage forms include, but are not limited to, solutions
ready for
injection, dry (e.g., lyophilized) products ready to be dissolved or suspended
in a
pharmaceutically acceptable vehicle for injection, suspensions ready for
injection, and
emulsions.
[329] Suitable vehicles that can be used to provide parenteral dosage forms
are well known
to those skilled in the art. Examples include, but are not limited to: Water
for Injection
USP; aqueous vehicles such as, but not limited to, Sodium Chloride Injection,
Ringer's
Injection, Dextrose Injection, Dextrose and Sodium Chloride Injection, and
Lactated
Ringer's Injection; water miscible vehicles such as, but not limited to, ethyl
alcohol,
polyethylene glycol, and polypropylene glycol; and non-aqueous vehicles such
as, but not
limited to, corn oil, cottonseed oil, peanut oil, sesame oil, ethyl oleate,
isopropyl myristate,
and benzyl benzoate.
[330] Excipients that increase the solubility of one or more of the ABPs
disclosed herein
can also be incorporated into the parenteral dosage forms.
[331] In some embodiments, the parenteral dosage form is lyophilized.
Exemplary
lyophilized formulations are described, for example, in U.S. Pat. Nos.
6,267,958 and
6,171,586; and WO 2006/044908; each of which is incorporated by reference in
its
entirety.
Dosage and Unit Dosage Forms
[332] In human therapeutics, the doctor will determine the posology which
he considers
most appropriate according to a preventive or curative treatment and according
to the age,
weight, condition and other factors specific to the subject to be treated.
[333] In certain embodiments, a composition provided herein is a
pharmaceutical
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composition or a single unit dosage form. Pharmaceutical compositions and
single unit
dosage forms provided herein comprise a prophylactically or therapeutically
effective
amount of one or more prophylactic or therapeutic ABPs.
[334] The amount of the ABP or composition which will be effective in the
prevention or
treatment of a disorder or one or more symptoms thereof will vary with the
nature and
severity of the disease or condition, and the route by which the ABP is
administered. The
frequency and dosage will also vary according to factors specific for each
subject
depending on the specific therapy (e.g., therapeutic or prophylactic agents)
administered,
the severity of the disorder, disease, or condition, the route of
administration, as well as
age, body, weight, response, and the past medical history of the subject.
Effective doses
may be extrapolated from dose-response curves derived from in vitro or animal
model test
systems.
[335] In certain embodiments, exemplary doses of a composition include
milligram or
microgram amounts of the ABP per kilogram of subject or sample weight (e.g.,
about 100
micrograms per kilogram to about 25 milligrams per kilogram, or about 100
micrograms
per kilogram to about 10 milligrams per kilogram). In certain embodiment, the
dosage of
the ABP provided herein, based on weight of the ABP, administered to prevent,
treat,
manage, or ameliorate a disorder, or one or more symptoms thereof in a subject
is 0.1
mg/kg, 1 mg/kg, 2 mg/kg, 3 mg/kg, 4 mg/kg, 5 mg/kg, 6 mg/kg, 10 mg/kg, 15
mg/kg, 20
mg/kg, 25 mg/kg, 30 mg/kg, or 40 mg/kg or more of a subject's body weight.
[336] The dose can be administered according to a suitable schedule, for
example, weekly,
once every two weeks, once every three weeks, or once every four weeks. In
some
embodiments, an antibody to be administered once every three or four weeks may
be
administered at a higher dose than the antibody administered every one or two
weeks. In
some embodiments, a loading dose is administered that is higher than the
maintenance
dose administered thereafter. It may be necessary to use dosages of the ABP
outside the
ranges disclosed herein in some cases, as will be apparent to those of
ordinary skill in the
art. Furthermore, it is noted that the clinician or treating physician will
know how and
when to interrupt, adjust, or terminate therapy in conjunction with subject
response.
[337] Different therapeutically effective amounts may be applicable for
different diseases
and conditions, as will be readily known by those of ordinary skill in the
art. Similarly,
amounts sufficient to prevent, manage, treat or ameliorate such disorders, but
insufficient
to cause, or sufficient to reduce, adverse effects associated with the ABPs
provided herein
are also encompassed by the dosage amounts and dose frequency schedules
provided
herein. Further, when a subject is administered multiple dosages of a
composition provided
herein, not all of the dosages need be the same. For example, the dosage
administered to
the subject may be increased to improve the prophylactic or therapeutic effect
of the
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composition or it may be decreased to reduce one or more side effects that a
particular
subject is experiencing.
[338] In certain embodiments, treatment or prevention can be initiated with
one or more
loading doses of an ABP or composition provided herein followed by one or more
maintenance doses.
[339] In certain embodiments, a dose of an ABP or composition provided
herein can be
administered to achieve a steady-state concentration of the ABP in blood or
serum of the
subject. The steady-state concentration can be determined by measurement
according to
techniques available to those of skill or can be based on the physical
characteristics of the
subject such as height, weight and age.
[340] In certain embodiments, administration of the same composition may be
repeated and
the administrations may be separated by at least 1 day, 2 days, 3 days, 5
days, 10 days, 15
days, 30 days, 45 days, 2 months, 75 days, 3 months, or 6 months.
[341] As discussed in more detail elsewhere in this disclosure, an ABP
provided herein may
optionally be administered with one or more additional agents useful to
prevent or treat a
disease or disorder. The effective amount of such additional agents may depend
on the
amount of ABP present in the formulation, the type of disorder or treatment,
and the other
factors known in the art or described herein.
Therapeutic Applications
[342] For therapeutic applications, the ABPs of the invention are
administered to a
mammal, generally a human, in a pharmaceutically acceptable dosage form such
as those
known in the art and those discussed above. For example, the ABPs of the
invention may
be administered to a human intravenously as a bolus or by continuous infusion
over a
period of time, by intramuscular, intraperitoneal, intra-cerebrospinal,
subcutaneous, intra-
articular, intrasynovial, intrathecal, or intratumoral routes. The ABPs also
are suitably
administered by peritumoral, intralesional, or perilesional routes, to exert
local as well as
systemic therapeutic effects. The intraperitoneal route may be particularly
useful, for
example, in the treatment of ovarian tumors.
[343] The ABPs provided herein may be useful for the treatment of any
disease or
condition involving GITR. In some embodiments, the disease or condition is a
disease or
condition that can benefit from treatment with an anti-GITR ABP. In some
embodiments,
the disease or condition is a tumor. In some embodiments, the disease or
condition is a cell
proliferative disorder. In some embodiments, the disease or condition is a
cancer.
[344] In some embodiments, the ABPs provided herein are provided for use as
a
medicament. In some embodiments, the ABPs provided herein are provided for use
in the
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manufacture or preparation of a medicament. In some embodiments, the
medicament is for
the treatment of a disease or condition that can benefit from an anti-GITR
ABP. In some
embodiments, the disease or condition is a tumor. In some embodiments, the
disease or
condition is a cell proliferative disorder. In some embodiments, the disease
or condition is
a cancer.
[345] Any suitable cancer may be treated with the ABPs provided herein.
Illustrative
suitable cancers include, for example, acute lymphoblastic leukemia (ALL),
acute myeloid
leukemia (AML), adrenocortical carcinoma, anal cancer, appendix cancer,
astrocytoma,
basal cell carcinoma, brain tumor, bile duct cancer, bladder cancer, bone
cancer, breast
cancer, bronchial tumor, carcinoma of unknown primary origin, cardiac tumor,
cervical
cancer, chordoma, colon cancer, colorectal cancer, craniopharyngioma, ductal
carcinoma,
embryonal tumor, endometrial cancer, ependymoma, esophageal cancer,
esthesioneuroblastoma, fibrous histiocytoma, Ewing sarcoma, eye cancer, germ
cell tumor,
gallbladder cancer, gastric cancer, gastrointestinal carcinoid tumor,
gastrointestinal stromal
tumor, gestational trophoblastic disease, glioma, head and neck cancer,
hepatocellular
cancer, histiocytosis, Hodgkin lymphoma, hypopharyngeal cancer, intraocular
melanoma,
islet cell tumor, Kaposi sarcoma, kidney cancer, Langerhans cell
histiocytosis, laryngeal
cancer, lip and oral cavity cancer, liver cancer, lobular carcinoma in situ,
lung cancer,
macroglobulinemia, malignant fibrous histiocytoma, melanoma, Merkel cell
carcinoma,
mesothelioma, metastatic squamous neck cancer with occult primary, midline
tract
carcinoma involving NUT gene, mouth cancer, multiple endocrine neoplasia
syndrome,
multiple myeloma, mycosis fungoides, myelodysplastic syndrome,
myelodysplastic/myeloproliferative neoplasm, nasal cavity and par nasal sinus
cancer,
nasopharyngeal cancer, neuroblastoma, non-small cell lung cancer,
oropharyngeal cancer,
osteosarcoma, ovarian cancer, pancreatic cancer, papillomatosis,
paraganglioma,
parathyroid cancer, penile cancer, pharyngeal cancer, pheochromocytomas,
pituitary
tumor, pleuropulmonary blastoma, primary central nervous system lymphoma,
prostate
cancer, rectal cancer, renal cell cancer, renal pelvis and ureter cancer,
retinoblastoma,
rhabdoid tumor, salivary gland cancer, Sezary syndrome, skin cancer, small
cell lung
cancer, small intestine cancer, soft tissue sarcoma, spinal cord tumor,
stomach cancer, T-
cell lymphoma, teratoid tumor, testicular cancer, throat cancer, thymoma and
thymic
carcinoma, thyroid cancer, urethral cancer, uterine cancer, vaginal cancer,
vulvar cancer,
and Wilms tumor.
[346] In some embodiments, provided herein is a method of treating a
disease or condition
in a subject in need thereof by administering an effective amount of an ABP
provided
herein to the subject. In some aspects, the disease or condition is a cancer.
[347] In some embodiments, provided herein is a method of multimerizing
GITR in a target
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cell of a subject in need thereof by administering an effective amount of an
ABP provided
herein to the subject.
[348] In some embodiments, provided herein is a method of agonizing GITR in
a target cell
of a subject in need thereof by administering an effective amount of an ABP
provided
herein to the subject. In some aspects, agonism of GITR by an ABP provided
herein results
in increased secretion of IL-2Ra, IL-2, IL-8, and/or IFNy by a target cell.
[349] In some embodiments, provided herein is a method of modulating NF-KB
activity in a
target cell of a subject in need thereof by administering an effective amount
of an ABP
provided herein to the subject.
[350] In some embodiments, provided herein is a method of modulating
degradation of IKB
in a target cell of a subject in need thereof by administering an effective
amount of an ABP
provided herein to the subject.
[351] In some embodiments, provided herein is a method of activating the MAPK
pathway
in a target cell of a subject in need thereof by administering an effective
amount of an ABP
provided herein to the subject. In some aspects, the components of the MAPK
pathway
that are activated by an ABP provided herein include one or more of p38, JNK,
and ERK.
[352] In some embodiments, provided herein is a method of increasing the
proliferation,
survival, and/or function of an effector T cell in a subject in need thereof
by administering
an effective amount of an ABP provided herein to the subject. In some aspects
the effector
T cell is a CD4+ effector T cell. In some aspects, the effector T cell is a
CD8+ effector T
cell.
[353] In some embodiments, provided herein is a method of abrogating
suppression of an
effector T cell by a regulatory T cell in a subject in need thereof by
administering an
effective amount of an ABP provided herein to the subject. In some aspects,
the regulatory
T cell is a CD4+CD25+Foxp3+ regulatory T cell. In some aspects, the regulatory
T cell is
a CD8+CD25+ regulatory T cell.
[354] In some embodiments, provided herein is a method of altering the
frequency of
occurrence or distribution or regulatory T cells in a subject in need thereof
by
administering an effective amount of an ABP provided herein to the subject. In
some
aspects, the frequency of regulatory T cells is decreased. In some aspects,
the frequency of
regulatory T cells is reduced in a particular tissue. In some aspects, the
intratumoral
accumulation of regulatory T cells is decreased, resulting in a more favorable
ratio of
effector T cells to regulatory T cells, and enhancing CD8+ T cell activity.
[355] In some embodiments, provided herein is a method of increasing the
activity of a
natural killer (NK) cell in a subject in need thereof by administering an
effective amount of
an ABP provided herein to the subject.
[356] In some embodiments, provided herein is a method of increasing the
activity of a
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dendritic cell in a subject in need thereof by administering an effective
amount of an ABP
provided herein to the subject.
[357] In some embodiments, provided herein is a method of increasing the
activity of a B
cell in a subject in need thereof by administering an effective amount of an
ABP provided
herein to the subject.
[358] In some embodiments, provided herein is a method of enhancing an
immune response
in a subject in need thereof by administering an effective amount of an ABP
provided
herein to the subject.
[359] In some embodiments, provided herein is a method delaying the onset
of a tumor in a
subject in need thereof by administering an effective amount of an ABP
provided herein to
the subject.
[360] In some embodiments, provided herein is a method of delaying the
onset of a cancer
in a subject in need thereof by administering an effective amount of an ABP
provided
herein to the subject.
[361] In some embodiments, provided herein is a method of reducing the size
of a tumor in
a subject in need thereof by administering an effective amount of an ABP
provided herein
to the subject.
[362] In some embodiments, provided herein is a method of reducing the
number of
metastases in a subject in need thereof by administering an effective amount
of an ABP
provided herein to the subject.
Combination Therapies
[363] In some embodiments, an ABP provided herein is administered with at
least one
additional therapeutic agent. Any suitable additional therapeutic agent may be
administered with an ABP provided herein. In some aspects, the additional
therapeutic
agent is selected from radiation, a cytotoxic agent, a chemotherapeutic agent,
a cytostatic
agent, an anti-hormonal agent, an EGFR inhibitor, an immunostimulatory agent,
an anti-
angiogenic agent, and combinations thereof
[364] In some embodiments, the additional therapeutic agent comprises an
immunostimulatory agent.
[365] In some embodiments, the immunostimulatory agent is an agent that
blocks signaling
of an inhibitory receptor of an immune cell, or a ligand thereof In some
aspects, the
inhibitory receptor or ligand is selected from CTLA-4, PD-1, PD-L1, NRP-1, LAG-
3,
Tim3, TIGIT, neuritin, BTLA, KIR, and combinations thereof In some aspects,
the agent
is selected from pembrolizumab (anti-PD-1), nivolumab (anti-PD-1),
atezolizumab (anti-
PD-L1), ipilimumab (anti-CTLA-4), and combinations thereof
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[366] In some embodiments, the immunostimulatory agent is an agonist of a
co-stimulatory
receptor of an immune cell. In some aspects, the co-stimulatory receptor is
selected from
0X40, ICOS, CD27, CD28, 4-1BB, or CD40. In some embodiments, the agonist is an
antibody.
[367] In some embodiments, the immunostimulatory agent is a cytokine. In
some aspects,
the cytokine is selected from IL-2, IL-5, IL-7, IL-12, IL-15, IL-21, and
combinations
thereof
[368] In some embodiments, the immunostimulatory agent is an oncolytic
virus. In some
aspects, the oncolytic virus is selected from a herpes simplex virus, a
vesicular stomatitis
virus, an adenovirus, a Newcastle disease virus, a vaccinia virus, and a
maraba virus.
[369] In some embodiments, the immunostimulatory agent is a T cell with a
chimeric
antigen receptor (CAR-T cell). In some embodiments, the immunostimulatory
agent is a
bi- or multi-specific T cell directed antibody. In some embodiments, the
immunostimulatory agent is an anti-TGF-B antibody. In some embodiments, the
immunostimulatory agent is a TGF-B trap.
[370] In some embodiments, the additional therapeutic agent is a vaccine to
a tumor
antigen. Any suitable antigen may be targeted by the vaccine, provided that it
is present in
a tumor treated by the methods provided herein. In some aspects, the tumor
antigen is a
tumor antigen that is overexpressed in comparison its expression levels in
normal tissue. In
some aspects, the tumor antigen is selected from cancer testis antigen,
differentiation
antigen, NY-ESO-1, MAGE-AL MART, and combinations thereof
[371] Further examples of additional therapeutic agents include a taxane
(e.g., paclitaxel or
docetaxel); a platinum agent (e.g., carboplatin, oxaliplatin, and/or
cisplatin); a
topoisomerase inhibitor (e.g., irinotecan, topotecan, etoposide, and/or
mitoxantrone);
folinic acid (e.g., leucovorin); or a nucleoside metabolic inhibitor (e.g.,
fluorouracil,
capecitabine, and/or gemcitabine). In some embodiments, the additional
therapeutic agent
is folinic acid, 5-fluorouracil, and/or oxaliplatin. In some embodiments, the
additional
therapeutic agent is 5-fluorouracil and irinotecan. In some embodiments, the
additional
therapeutic agent is a taxane and a platinum agent. In some embodiments, the
additional
therapeutic agent is paclitaxel and carboplatin. In some embodiments, the
additional
therapeutic agent is pemetrexate. In some embodiments, the additional
therapeutic agent is
a targeted therapeutic such as an EGFR, RAF or MEK-targeted agent.
[372] The additional therapeutic agent may be administered by any suitable
means. In some
embodiments, an ABP provided herein and the additional therapeutic agent are
included in
the same pharmaceutical composition. In some embodiments, an ABP provided
herein and
the additional therapeutic agent are included in different pharmaceutical
compositions.
[373] In embodiments where an ABP provided herein and the additional
therapeutic agent
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are included in different pharmaceutical compositions, administration of the
ABP can
occur prior to, simultaneously, and/or following, administration of the
additional
therapeutic agent. In some aspects, administration of an ABP provided herein
and the
additional therapeutic agent occur within about one month of each other. In
some aspects,
administration of an ABP provided herein and the additional therapeutic agent
occur
within about one week of each other. In some aspects, administration of an ABP
provided
herein and the additional therapeutic agent occur within about one day of each
other. In
some aspects, administration of an ABP provided herein and the additional
therapeutic
agent occur within about twelve hours of each other. In some aspects,
administration of an
ABP provided herein and the additional therapeutic agent occur within about
one hour of
each other.
Diagnostic Methods
[374] Also provided are methods for detecting the presence of GITR on cells
from a
subject. Such methods may be used, for example, to predict and evaluate
responsiveness to
treatment with an ABP provided herein.
[375] In some embodiments, a blood sample is obtained from a subject and
the fraction of
cells expressing GITR is determined. In some aspects, the relative amount of
GITR
expressed by such cells is determined. The fraction of cells expressing GITR
and the
relative amount of GITR expressed by such cells can be determined by any
suitable
method. In some embodiments, flow cytometry is used to make such measurements.
In
some embodiments, fluorescence assisted cell sorting (FACS) is used to make
such
measurement. See Li et al., I Autoimmunity, 2003, 21:83-92 for methods of
evaluating
expression of GITR in peripheral blood.
Kits
[376] Also provided are kits comprising the ABPs provided herein. The kits
may be used
for the treatment, prevention, and/or diagnosis of a disease or disorder, as
described herein.
[377] In some embodiments, the kit comprises a container and a label or
package insert on
or associated with the container. Suitable containers include, for example,
bottles, vials,
syringes, and IV solution bags. The containers may be formed from a variety of
materials,
such as glass or plastic. The container holds a composition that is by itself,
or when
combined with another composition, effective for treating, preventing and/or
diagnosing a
disease or disorder. The container may have a sterile access port. For
example, if the
container is an intravenous solution bag or a vial, it may have a port that
can be pierced by
a needle. At least one active agent in the composition is an ABP provided
herein. The label
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or package insert indicates that the composition is used for treating the
selected condition.
[378] In some embodiments, the kit comprises (a) a first container with a
first composition
contained therein, wherein the first composition comprises an ABP provided
herein; and
(b) a second container with a second composition contained therein, wherein
the second
composition comprises a further therapeutic agent. The kit in this embodiment
of the
invention may further comprise a package insert indicating that the
compositions can be
used to treat a particular condition.
[379] Alternatively, or additionally, the kit may further comprise a second
(or third)
container comprising a pharmaceutically-acceptable excipient. In some aspects,
the
excipient is a buffer. The kit may further include other materials desirable
from a
commercial and user standpoint, including filters, needles, and syringes.
Other Illustrative Embodiments
[380] The embodiments provided below are non-limiting and provided by way
of
illustration of certain embodiments and aspects of the invention, in addition
to those
described throughout this disclosure.
[381] Embodiment 1: An ABP that binds specifically to a human GITR and/or a
human
GITR complex and is capable of at least one of the following: a) cross-
competes with
GITRL for binding to GITR; b) can be internalized into a human cell; c)
inhibits
suppression of an effector T cell; d) inhibits regulatory T cell inhibition of
effector T cells;
e) decreases the number of regulatory T cells in tissues or in circulation; 0
activates an
effector T cell; g) associates GITR into a GITR complex; h) modulates an
activity of a
human GITR and/or a GITR complex.
[382] Embodiment 2: The ABP of embodiment 1, wherein the ABP has one or more
of the
following characteristics: a) is a monoclonal antibody; b) is a human
antibody, a
humanized antibody, or a chimeric antibody; c) is a multispecific or
multivalent antibody,
e.g., a tetravalent antibody; d) comprises a at least one Fab at the N-
terminus or the C-
terminus; e) is of the IgGl, IgG2, IgG3, or the IgG4 type; f) is an antigen-
binding antibody
fragment; g) is a Fab fragment, a Fab' fragment, a F(ab')2 fragment, or an Fv
fragment; h)
is a bispecific antibody, a diabody, a single chain antibody, a single domain
antibody, a VH
domain antibody, or a nanobody.
[383] Embodiment 3: The ABP of embodiment 1, wherein the ABP has one or more
of the
following characteristics: a) binds to a human GITR polypeptide of SEQ ID NOs:
1-2 or a
variant thereof, or as otherwise provided herein with a KD of less than about
20 nM; or b)
binds to a cyno GITR polypeptide of SEQ ID NO: 3 or a variant thereof, or as
otherwise
provided herein, with a KD of less than about 200 nM.
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[384] Embodiment 4: The ABP of embodiment 1, comprising a first antigen-
binding
domain that specifically binds to a first antibody recognition domain on human
GITR and
a second antigen-binding domain that specifically binds to a second antibody
recognition
domain on human GITR, wherein the first antibody recognition domain and the
second
antibody recognition domain are not identical.
[385] Embodiment 5: The ABP of embodiment 4, comprising a bispecific
binding protein
in a format selected from the group consisting of a DVD-IgTM molecule, a BiTe
molecule,
a DART molecule, molecule, a DuoBody molecule, a scFv/diabody-IgG molecule, a
cross-over
multispecific molecule, a 2-in-1 bispecific molecule, a knob-in-hole
multispecific
molecule, a Fab+IgG molecule, a CovX-Body molecule, an affibody molecule, a
scFv/diabody-CH2/CH3 bispecific molecule, a IgG-non-Ig protein scaffold-based
multispecific molecule, a Fynomer molecule, a FcabTM molecule, a TandAbO, a
ZybodyTM, and a scFV/diabody linked to normal human protein like human serum
albumin-bispecific molecule.
[386] Embodiment 6: The ABP of embodiment 4, wherein the first antigen-
binding
domain has a KD of less than about 20 nM and is capable of agonizing human
GITR, and
wherein the second antigen-binding domain has a KD of less than about 100 nM.
[387] Embodiment 7: An ABP that competes or is capable of competing for
binding to
human GITR with a reference ABP, wherein the reference ABP is the ABP of
embodiment
1.
[388] Embodiment 8: The ABP of embodiment 7, wherein the ABP and the reference
antibody cross-compete or are capable of cross-competing for binding to a
human GITR.
[389] Embodiment 9: The ABP of embodiment 1, comprising a heavy chain constant
region comprising a human heavy chain constant region or fragment or a variant
thereof,
wherein the constant region variant comprises up to 20 modified amino acid
substitutions,
wherein from 0 to up to 20 modified amino acid substitutions are conservative
amino acid
substitutions.
[390] Embodiment 10: The ABP of embodiment 1, that competes or is capable of
competing for binding to human GITRL with a GITR protein.
[391] Embodiment 11: The ABP of embodiment 1, that is capable of activating
GITR
signaling in a ligand-independent manner.
[392] Embodiment 12: The ABP of embodiment 1, that is capable of enhancing
ligand-
dependent binding of GITR and GITRL.
[393] Embodiment 13: A pharmaceutical composition comprising an ABP of any one
of
embodiments 1-12 in a pharmaceutically acceptable carrier.
[394] Embodiment 14: A bispecific antibody comprising a first antigen-
binding domain
that specifically binds to a first antibody recognition domain on human GITR
and a second
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antigen-binding domain that specifically binds to a second antibody
recognition domain on
human GITR, wherein the first antibody recognition domain and the second
antibody
recognition domain are not identical.
[395] Embodiment 15: The bispecific antibody of embodiment 14, wherein the
first
antibody recognition domain and the second antibody recognition domain are
present in
the extracellular domain of human GITR.
[396] Embodiment 16: The bispecific antibody of embodiment 14, wherein the
first
antibody recognition domain and the second antibody recognition domain are
capable of
associating at least two human GITR proteins into a functional complex.
[397] Embodiment 17: A complexing ABP comprising a first antigen-binding
domain that
specifically binds to a first antibody recognition domain on a human GITR
protein or a
human GITR complex comprising at least two GITR proteins, and is capable of at
least
one of the following: a) cross-competes with GITRL for binding to GITR; b) can
be
internalized into a human cell; c) inhibits suppression of an effector T cell;
d) inhibits
regulatory T cell inhibition of effector T cells; e) decreases the number of
regulatory T
cells in tissues or in circulation; f) activates an effector T cell; g)
associates GITR into a
GITR complex; h) modulates an activity of a human GITR and/or a GITR complex.
[398] Embodiment 18: An isolated nucleic acid encoding an ABP according to
any one of
embodiments 1 to 12 or a bispecific antibody according to any one of
embodiments 14-16
or the complexing ABP of embodiment 17.
[399] Embodiment 19: An expression vector comprising the nucleic acid
according to
embodiment 18.
[400] Embodiment 20: A prokaryotic or eukaryotic host cell comprising a
vector of
embodiment 19.
[401] Embodiment 21: A method for the production of a recombinant protein
comprising
the steps of expressing a nucleic acid according to embodiment 18 in a
prokaryotic or
eukaryotic host cell and recovering said protein from said cell or the cell
culture
supernatant.
[402] Embodiment 22: A method for treatment of a subject suffering from cancer
or from
an inflammatory disease, comprising the step of administering to the subject a
pharmaceutical composition comprising an ABP according to any one of
embodiments 1
to 12 or a bispecific antibody according to any one of embodiments 14-16 or
the
complexing ABP of embodiment 17.
[403] Embodiment 23: A method for inducing or enhancing an immune response in
a
subject, comprising the step of administering to the subject a pharmaceutical
composition
comprising an ABP according to any one of embodiments 1 to 12 or a bispecific
antibody
according to any one of embodiments 14-16 or the complexing ABP of embodiment
17,
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wherein the immune response is generated against a tumor antigen.
[404] Embodiment 24: The method of embodiment 23, wherein the ABP, bispecific
antibody or the complexing ABP is administered in an amount sufficient to
achieve one or
more of the following in the subject: a) reduce regulatory T cells suppression
of activity of
effector T cells; b) decrease levels of regulatory T cells; c) activation of
effector T cells; d)
induce or enhance effector T cell proliferation; e) inhibit tumor growth; and
f) induce
tumor regression.
[405] Embodiment 25: The method of embodiment 22, wherein the cancer is a
solid
cancer.
[406] Embodiment 26: The method of embodiment 22, wherein the cancer is a
hematological cancer.
[407] Embodiment 27: The method of any one of embodiments 22-26, wherein the
method
further comprises one or more of the following a) administering chemotherapy;
b)
administering radiation therapy; or c) administering one or more additional
therapeutic
agents.
[408] Embodiment 28: The method of embodiment 27, wherein the additional
therapeutic
agent comprises an immunostimulatory agent.
[409] Embodiment 29: The method of embodiment 27, wherein the
immunostimulatory
agent comprises an antagonist to an inhibitory receptor of an immune cell.
[410] Embodiment 30: The method of embodiment 29, wherein the inhibitory
receptor
comprises CTLA-4, PD-1, PD-L1, LAG-3, Tim3, TIGIT, neuritin, BTLA or KIR, or a
functional fragment thereof
[411] Embodiment 31: The method of embodiment 27, wherein the
immunostimulatory
agent comprises an agonist of co-stimulatory receptor of an immune cell, or a
functional
fragment thereof
[412] Embodiment 32: The method of embodiment 31, wherein the co-
stimulatory
receptor comprises 0X40, ICOS, CD27, CD28, 4-1BB, or CD40.
[413] Embodiment 33: The method of embodiment 27, wherein the
immunostimulatory
agent comprises a cytokine.
[414] Embodiment 34: The method of embodiment 27, wherein the cytokine
comprises
IL-2, IL-5, IL-7, IL-12, IL-15 or IL-21.
[415] Embodiment 35: The method of embodiment 27, wherein the
immunostimulatory
agent comprises an oncolytic virus.
[416] Embodiment 36: The method of embodiment 35, wherein the oncolytic
virus
comprises a Herpes simplex virus, a Vesicular stomatitis virus, an adenovirus,
a Newcastle
disease virus, a vaccinia virus, or a maraba virus.
[417] Embodiment 37: The method of embodiment 27, wherein the
immunostimulatory
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agent comprises a chimeric antigen engineered T cell.
[418] Embodiment 38: The method of embodiment 27, wherein the
immunostimulatory
agent comprises a bi- or multi-specific T cell directed antibody.
[419] Embodiment 39: The method of embodiment 27, wherein the additional
therapeutic
agent comprises an anti-TGF-B antibody or a TGF-B receptor trap.
[420] Embodiment 40: The method of any one of embodiments 22-39, wherein
administration of the pharmaceutical composition results in induction or
enhancement of
proliferation of a T-effector cell, or modulation of I-kappaB and/or NF-KB in
the T cell, or
modulation of GITR activity in the T cell, or T cell receptor induced
signaling in a T-
effector cell, or a combination thereof
[421] Embodiment 41: A method of screening for test compounds comprising an
ABP
according to any one of embodiments 1 to 12 that are capable of inhibiting the
interaction
of GITRL with GITR complex comprising the steps of: contacting a sample
containing
GITRL and GITR complex with the compound; and determining whether the
interaction of
GITRL with GITR complex in the sample is decreased relative to the interaction
of GITRL
with GITR complex in a sample not contacted with the compound, whereby a
decrease in
the interaction of GITRL with GITR in the sample contacted with the compound
identifies
the compound as one that inhibits the interaction of GITRL with GITR complex.
EXAMPLES
[422] The following are examples of methods and compositions of the
invention. It is
understood that various other embodiments may be practiced, given the general
description
provided herein.
Example 1: Selection of GITR Antigen-Binding Proteins
Materials and methods
Antigens were biotinylated using the EZ-Link Sulth-NHS-Biotinylation Kit from
Pierce.
Goat F(ab):2 anti-human kappa-FITC (LC-FITC), ExtrA.vidi.n. -PE (EA-PE) and
Streptavidin-AF633
(SA.-633) were obtained from Southern Biotech, Sigma, and Molecular Probes,
respectively.
Streptavidin MicroBeads and MACS LC separation columns were purchased from
Miltenyi Biotec.
Goat anti-human IgG-PE (Human-PE) was obtained from Southern Biotech.
Naïve Discovery
Eight naïve human synthetic yeast libraries each of ¨109 diversity were
propagated as
previously described (see, e.g., Y. Xu et al, Addressing polyspecificity of
antibodies selected from an
in vitro yeast presentation system: a FACS-based, high-throughput selection
and analytical tool.
PEDS 26.10, 663-70 (2013); W02009036379; W02010105256; and W02012009568.) For
the first
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two rounds of selection, a magnetic bead sorting technique utilizing the
Miltenyi MACS system was
performed, as previously described (see, e.g., Siegel et al, High efficiency
recovery and epitope-
specific sorting of an scFv yeast display library." J Immunol Methods 286(1-
2), 141-153 (2004))
Briefly, yeast cells (-1010 cells/library) were incubated with 5 ml of 10 nM
biotinylated Fe fusion
antigen for 30 min at 30 C in wash buffer (phosphate-buffered saline
(PBS)/0.1% bovine serum
albumin (BSA)). After washing once with 40 ml ice-cold wash buffer, the cell
pellet was resuspended
in 20 mL wash buffer, and Streptavidin MicroBeadsq.1) (500 ill) were added to
the yeast and incubated
for 15 min at 4 C. Next, the yeast were pelleted, resuspended in 20 mL wash
buffer, and loaded onto
a Miltenyi LS column. After the 20 mL were loaded, the column was washed 3
times with 3 ml wash
buffer. The column was then removed from the magnetic field, and the yeast
were eluted with 5 mL
of growth media and then grown overnight. The following rounds of selection
were performed using
flow cytometry. Approximately 2x107 yeast were pelleted, washed three times
with wash buffer, and
incubated at 30 C with either decreasing concentrations of biotinylated Fe
fusion antigen (10 to 1 nM)
under equilibrium conditions, 10 nM biotinylated Fe fusion antigens of
different species in order to
obtain species cross-reactivity, or with a poly-specificity depletion reagent
(PSR) to remove non-
specific antibodies from the selection. For the PSR depletion, the libraries
were incubated with a 1:10
dilution of biotinylated PSR reagent as previously described (see, e.g., Y. Xu
et al, Addressing
polyspecificity of antibodies selected from an in vitro yeast presentation
system: a FACS-based, high-
throughput selection and analytical tool. PEDS 26.10, 663-70 (2013)) Yeast
were then washed twice
with wash buffer and stained with LC-FITC (diluted 1:100) and either SA-633
(diluted 1:500) or
EAPE (diluted 1:50) secondary reagents for 15 min at 4 C. After washing twice
with wash buffer, the
cell pellets were resuspended in 0.3 mL wash buffer and transferred to
strainer-capped sort tubes.
Sorting was performed using a FACS ARIA sorter (BD Biosciences) and sort gates
were determined
to select for antibodies with desired characteristics. Selection rounds were
repeated until a population
with all of the desired characteristics was obtained. After the final round of
sorting, yeast were plated
and individual colonies were picked for characterization.
Antibody Optimization
Optimization of antibodies was performed via a light chain diversification
protocol, and then
by introducing diversities into the heavy chain and light chain variable
regions as described below. A
combination of some of these approaches was used for each antibody.
Light chain batch diversification protocol: Heavy chain plasmids from a naive
selection
output were extracted from the yeast via smash and grab, propagated in and
subsequently purified
from E.coli, and transformed into a light chain library with a diversity of 5
x 106. Selections were
perfonned with one round of MACS and four rounds of FACS employing the same
conditions as the
naive discovery.
Light chain diversification: The heavy chain variable region of a single
antibody was
amplified via PCR and transfonned, along with the heavy chain expression
vector, into a light chain
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library with a diversity of 5 x 106. Selections were performed with one round
of MACS and three
rounds of FACS employing the same conditions as the naïve discovery. For each
FACS round the
libraries were looked at for PSR binding, species cross-reactivity, and
affinity pressure, and sorting
was performed in order to obtain a population with the desired
characteristics.
CDRH1 and CDRH2 selection: The CDRH3 of a single antibody was recombined into
a
premade library with CDRH1 and CDRH2 variants of a diversity of 1 x 108 and
selections were
performed with one round of MACS and four rounds of FACS as described in the
naive discovery.
For each FACS round, the libraries were looked at for PSR binding, species
cross-reactivity, and
affinity pressure, and sorting was performed in order to obtain a population
with the desired
characteristics. For these selections affinity pressures were applied by
preincubating the biotinylated
antigen with parental IgG for 30 minutes and then applying that precomplexed
mixture to the yeast
library for a length of time which would allow the selection to reach an
equilibrium. The higher
affinity antibodies were then able to be sorted.
CDRH3NH Mutant selection: Oligonucleotides (oligos) were ordered from IDT
which
comprised the CDRH3 as well as a flanking region on either side of the CDRH3.
Amino acid
positions in the CDRH3-encoding portion of the oligo were variegated by NNK
diversity. The
CDRH3 oligos were double-stranded using primers which annealed to the flanking
region of the
CDRH3. The remaining part of the heavy chain variable region was mutagenized
via error prone PCR
in order to introduce additional diversity in non-CDR3 regions of the heavy
chain. The library was
then created by transforming the double stranded CDRH3 oligo, the mutagenized
remainder of the
heavy chain variable region, and the heavy chain expression vector into yeast
already containing the
light chain plasmid of the parent. Selections were performed similar to
previous cycles using FACS
sorting for four rounds. For each FACS round, the libraries were looked at for
PSR binding, species
cross-reactivity, and affinity pressure, and sorting was performed in order to
obtain a population with
the desired characteristics. Affinity pressures for these selections were
performed as described above
in the CDRH1 and CDRH2 selection.
CDRL1, CDRL2, and CDRL3 selection: Oligos were ordered from IDT which
comprised the
CDRL3 as well as a flanking region on either side of the CDRL3. Amino acid
positions in the
CDRL3-encoding portion of the oligo were variegated by NNK diversity. The
CDRL3 oligos were
double-stranded using primers which annealed to the flanking region of the
CDRL3. These double-
stranded CDRL3 oligos were then recombined into a premade library with CDRL1
and CDRL2
variants of a diversity of 3 x 105 and selections were performed with one
round of MACS and four
rounds of FACS as described in the naïve discovery. For each FACS round the
libraries were looked
at for PSR binding, species cross-reactivity, and affinity pressure, and
sorting was performed in order
to obtain a population with the desired characteristics. Affinity pressures
for these selections were
performed as described above in the CDRH1 and CDRH2 selection.
Antibody production and purification
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Yeast clones were grown to saturation and then induced for 48 h at 30 C with
shaking. After
induction, yeast cells were pelleted and the supernatants were harvested for
purification. igGs were
purified using a Protein A column and eluted with acetic acid, pH 2Ø Fab
fragments were generated
by papain digestion and purified over KappaSelect (GE Healthcare
LifeSciences).
ForteBio KD measurements
ForteBio affinity measurements were performed on an Octet RED384 generally
as
previously described (see, e.g., Estep et al, High throughput solution-based
measurement of antibody-
antigen affinity and epitope binning. Alabs 5(2), 270-278 (2013)). Briefly.
ForteBio affinity
measurements were performed by loading IgGs on-line onto AHQ sensors. Sensors
were equilibrated
off-line in assay buffer for 30 min and then monitored on-line for 60 seconds
for baseline
establishment. Sensors with loaded IgGs were exposed to 100 nM antigen for 3
minutes, and
afterwards were transferred to assay buffer for 3 min for off-rate
measurement. For monovalent
affinity assessment Fabs were used instead of IgGs. For this assessment, the
unbiotinylated Fc fusion
antigen was loaded on-line onto the AHQ sensors. Sensors were equilibrated off-
line in assay buffer
for 30 min and then monitored on-line for 60 seconds for baseline
establishment. Sensors with loaded
antigen were exposed to 200 nM Fab for 3 minutes, and afterwards they were
transferred to assay
buffer for 3 min for off-rate measurement. All kinetics were analyzed using
the 1:1 binding model.
ForteBio Epitope Binning/Ligand Blocking
Epitope binning/ligand blocking was performed using a standard sandwich format
cross-
blocking assay. Control anti-target IgG was loaded onto AHQ sensors and
unoccupied Fc-binding
sites on the sensor were blocked with an irrelevant human IgG1 antibody. The
sensors were then
exposed to 100 nM target antigen followed by a second anti-target antibody or
ligand. Additional
binding by the second antibody or ligand after antigen association indicates
an unoccupied epitope
(non-competitor), while no binding indicates epitope blocking (competitor or
ligand blocking).
Cell Binding Analysis
Approximately 100,000 cells overexpressing the antigen were washed with wash
buffer and
incubated with 100 [11 100 nM IgG for 5 minutes at room temperature. Cells
were then washed twice
with wash buffer and incubated with 100u1 of 1:100 Human-PE for 15 minutes on
ice. Cells were
then washed twice with wash buffer and analyzed on a FACS Canto II analyzer
(BD Biosciences.)
Size Exclusion Chromatography
A TSKgel0 SuperSW mAb HTP column (22855) was used for fast SEC analysis of
mammalian produced mAbs at 0.4 mL/min with a cycle time of 6 min/run. 200 mM
Sodium
Phosphate and 250 mM Sodium Chloride was used as the mobile phase.
Dynamic Scanning Fluorime try
[IL of 20x Sypro Orange is added to 20 uL of 0.2-1mg/mL mAb or Fab solution.
An RT-
PCR instrument (BioRad CFX96 RT PCR) is used to ramp the sample plate
temperature from 40 to
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95 C at 0.5C increment, with 2min equilibrate at each temperature. The
negative of first derivative
for the raw data is used to extract Tm.
Example 2: Characteristics of TM Format Antibodies
Affinity of IgG1 and TM format ABPs for GITR of several species
[423] The GITR ABPs were evaluated for their ability to bind recombinant hGITR
(SEQ ID
NO: 1), hGITR-T43R (SEQ ID NO: 2), cGITR (SEQ ID NO: 3), and mGITR (SEQ ID
NO: 4) using a FORTEBIO OCTET instrument. The assay was performed at 30 C,
using
1 x Kinetics Buffer (ForteBio, Inc.) as an assay buffer. Anti-human IgG Fc
Capture (AHC)
biosensors (ForteBio, Inc.) were used to capture GITR ABPs onto the sensors.
The sensors
were equilibrated in assay buffer for 600 seconds before the assay. Baseline
was
established by dipping the sensors into lx assay buffer for 60 seconds. ABPs
were loaded
onto the sensors by dipping the sensors into ABP solution for 300 seconds.
Baseline was
established by dipping the sensors into lx assay buffer for 120 seconds. The
sensors were
quenched by dipping into 200 ug/m1 human IgG for 300 seconds to prevent non-
specific
binding of GITR-Fc antigens to the sensor. Baseline was established by dipping
the
sensors into lx assay buffer for 60 seconds. Next, association was monitored
for 300
seconds in 25 nM of recombinant antigens, and dissociation was followed for
1200
seconds in buffer alone. KD was determined using the kinetic function of the
FORTEBIO
Analysis Software using a 1:1 binding model. Results are shown in Table 5.
Table 5: ABP KD by OCTET using single concentration kinetics
KD by Octet, KD by Octet, KD by Octet,
KD by Octet,
single- single- single-
single-
concentration concentration concentration
ABP Format concentration
of human of cyno of mouse
of human
GITR T43R- GITR-Fc GITR-Fc
GITR-Fc, (nM)
Fc (nM) (nM) (nM)
1 N-terminal 0.77 0.94 0.88 No
binding
Fab TM
2 N-terminal 0.55 0.73 0.79 No
binding
Fab TM
3 N-terminal 0.58 0.71 1.1 No
binding
Fab TM
4 N-terminal 0.5 0.72 1.19 No
binding
Fab TM
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N-terminal 0.57 0.7 1.21 No binding
Fab TM
6 N-terminal 0.63 0.71 1.09 No
binding
Fab TM
7 N-terminal 0.76 1.1 1.9 No
binding
Fab TM
8 N-terminal 0.73 1.02 0.94 No
binding
Fab TM
[424] Other
GITR ABPs were evaluated for their ability to bind recombinant hGITR and
cGITR (SEQ ID NO: 3) using a FORTEBIO Octet RED384 instrument generally as
previously described (see, e.g., Estep et al, High throughput solution-based
measurement
of antibody-antigen affinity and epitope binning. Mabs 5(2), 270-278 (2013)).
AHQ
biosensors (ForteBio, Inc.) were used to capture GITR ABPs onto the sensors.
The sensors
were quenched by dipping into an unrelated human IgG1 antibody to prevent non-
specific
binding of GITR-Fc antigens to the sensor. The sensors were equilibrated in
assay buffer
for 30 minutes. Baseline was established by dipping the sensors into assay
buffer for 60
seconds. Association was monitored for 180 seconds in 100 nM of recombinant
antigens,
and dissociation was followed for 180 seconds in buffer alone. KD was
determined using
the kinetic function of the FORTEBIO Analysis Software using a 1:1 binding
model.
Results are shown in Table 6.
Table 6: ABP KD by OCTET using single concentration kinetics
MMMMMMRpql
KD by Octet, single- illi]!iosIQoggiomglog
ABP Format
concentration of human 111111rwompg410111=$;0
GITR-Fc, (nM)
35 (non-TM ABP
IgG1 N297A 1.5 5.7
corresponding to ABP1)
36 (non-TM ABP
IgG1 N297A 1.4 7.5
corresponding to ABP2)
37 (non-TM ABP
IgG1 N297A 0.7 8.1
corresponding to ABP3)
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38 (non-TM ABP
IgG1 N297A 0.6 11.1
corresponding to ABP4/5)
39 (non-TM ABP
IgG1 N297A 0.7 8.2
corresponding to ABP6)
40 (non-TM ABP
IgG1 N297A 2.2 4.9
corresponding to ABP7)
41 (non-TM ABP
IgG1 N297A 2.3 6.9
corresponding to ABP8)
19 N terminal Fab TM 5.2 12.6
20 N terminal Fab TM 4.4 5.2
21 N terminal Fab TM 4.2 5.5
22 N terminal Fab TM 4.0 4.6
23 N terminal Fab TM 3.7 No Binding
24 N terminal Fab TM 6.6 No Binding
25 C terminal Fab
TM, linker = 4.5 11.0
15mer
26 C terminal Fab
TM, linker = 5.1 5.1
15mer
27 C terminal Fab
TM, linker = 4.2 7.3
15mer
28 C terminal Fab
TM, linker = 3.2 4.3
15mer
29 C terminal Fab
TM, linker = 5.7 No Binding
15mer
30 C terminal Fab
TM, linker = 5.6 No Binding
15mer
31 C terminal Fab
7.6 No Binding
TM, linker = 5mer
32 C terminal Fab
6.4 No Binding
TM, linker = 5mer
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42 (ABPs 1-10, 23, 24, 29-
IgG1 N297A* 11.5 No Binding
32, 35-41, 43, 48, 49)
44 (ABPs 11, 12, 19, 25) IgG1 N297A# 46.5 Poor Fit
45 (ABPs 13, 20, 26, 45, IgG1 N297A#
112.2 Poor Fit
52)
46 (ABPs 14-16, 21, 27, IgG1 N297A#
59.8 110.9
53-55)
47 (ABPs 17, 18, 22, 28, IgG1 N297A#
44.3 88.1
56, 57)
48 (ABPs 10, 29, 31) IgG1 N297A# 14.8 No Binding
49 (ABPs 24, 30, 32) IgG1 N297A# 63.5 No Binding
11 IgG1 format 3.1 8.5
12 IgG1 format 2.8 7.9
13 IgG1 format 5.5 6.4
14 IgG1 format 2.8 5.9
15 IgG1 format 3.3 5.2
16 IgG1 format 2.9 5.4
17 IgG1 format 4.2 4.6
18 IgG1 format 4.0 8.4
* non-optimized, non-TM IgG1 ABPs corresponding to ABPs in parentheses
# H1H2-optimized, non-TM IgG1 ABPs corresponding to ABPs in parentheses
[425] Additional KD measurements were performed on 8 N-terminal Fab TM format
antibodies using multi-concentration kinetics. The GITR ABPs were evaluated
for their
ability to bind recombinant hGITR (SEQ ID NO: 1), hGITR-T43R (SEQ ID NO: 2),
and
cGITR (SEQ ID NO: 3) using a FORTEBIO OCTET instrument. The assay was
performed at 30 C, using lx Kinetics Buffer (ForteBio, Inc.) as an assay
buffer. Anti-
human IgG Fc Capture (AHC) biosensors (ForteBio, Inc.) were used to capture
GITR
ABPs onto the sensors. The sensors were saturated in assay buffer for 600
seconds before
the assay. Baseline was established by dipping the sensors into lx assay
buffer for 60
seconds. ABPs were loaded onto the sensors by dipping the sensors into ABP
solution for
300 seconds. Baseline was established by dipping the sensors into lx assay
buffer for 120
seconds. Next, association was monitored for 300 seconds in various
concentrations of
recombinant antigens (50 nM to 0.78 nM, 2-fold dilutions in assay buffer), and
dissociation was followed for 600 or 1200 seconds in buffer alone. KD was
determined
using the kinetic function of the FORTEBIO Analysis Software using a 1:1
binding
model. Results of determination of KD by OCTET are shown in FIG.2 and in Table
7. As
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can be seen in the Figure, the KDs for human GITR are each less than 1nM, and
the KDs
for cynomolgus GITR are within 15-fold of that of human. In addition, the TM
format
antibodies bind the T43R SNP variant of human GITR.
Table 7. N-terminal Fab TM format ABP Binding Characteristics
ABP Human GITR-Fc Human GITR-Fc T43R Cyno GITR-
Fc
KD (M), n=3 KD (M), n=1 KD (M), n=3
1 3.2E-10 5.3E-10 6.2E-10
2 2.3E-10 1.0E-10 7.1E-10
3 1.2E-10 2.7E-12 1.1E-09
4 1.1E-10 4.5E-12 1.5E-09
1.2E-10 7.3E-11 1.1E-09
6 1.5E-10 3.5E-11 1.6E-09
7 4.5E-10 7.0E-10 1.7E-09
8 5.2E-10 6.6E-10 6.6E-10
GITR Binding Assays
[426] Peripheral Blood Mononuclear Cells (PBMCs) were cultured in growth
medium in
the presence of phytohemagglutinin (PHA) for 5 days at 37 C to upregulate GITR
expression. T cells were collected and washed and then incubated at 4 C with
one of eight
TM format GITR ABPs or a human isotype control antibody. After washing, cells
were
incubated at 4 C with fluorochrome conjugated anti-human CD4 antibody and anti-
human
CD8 antibody as well as fluorochrome-conjugated anti-human IgG to detect bound
anti-
human GITR ABPs. The percentage of GITR+ CD4 and CD8 cells as well as the mean
fluorescence intensity (MFI) is determined by flow cytometry.
[427] Exemplary results are shown in FIG. 3. Figure 3A shows CD4+ cells and
Figure 3B
shows CD8+ cells. In the top row, from left to right, is shown an anti-GITR
positive
control, and anti-human IgG4 isotype control, and N-terminal Fab TM format
ABPs
ABP9, ABP2, ABP1, and ABP7. On the bottom row is shown ABP8, ABP3, ABP4,
ABP5, ABP6, ABP23, and ABP24. The percentage of IgG4+ CD4/8+ cells is
indicated.
[428] Binding to cell surface human, cynomolgus, and murine GITR is
evaluated by
transfecting HT1080, CHO, or 293T cells with the respective (human, cynomolgus
or
murine) GITR. Binding to cynomolgus GITR is further evaluated using primary
cynomolgus T cells and the HSC-F T cell line, each after stimulation with anti-
CD3
antibody, to increase GITR expression.
Activity Assays for GITR Antigen-Binding Proteins
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[429] The GITR ABPs were tested for their ability to agonize GITR. In one
assay, HT1080
cells that stably express human or cynomolgus GITR were contacted with TM
format
GITR ABPs or trimeric human GITRL. After 24 hours, the amount of IL-8 secreted
by the
cells was evaluated by an ELISA assay. Increased IL-8 secretion corresponds to
an
increase in GITR agonism. Results for N-terminal Fab TM format antibodies 1-8
are
shown in FIG. 4. Shown are the IL-8 output based on antibody or ligand
concentration for
ABP1 (FIG. 4A), ABP2 (FIG. 4B), ABP3 (FIG. 4C), ABP4 (FIG. 4D), ABP5 (FIG.
4E),
ABP6 (FIG. 4F), ABP7 (FIG. 4G), and ABP8 (FIG 4H). Antibodies shown as circles
in
the FIGs. and GITRL control is shown as squares. EC50 scores are compared to
that of
GITR ligand (GITRL).
[430] Table 8. Agonistic activity of TM Format antibodies on human and
cynomolgus
GITR on HT1080 cells
% Maximum IL-
Average EC50 % Maximum IL-
Average EC50 in 8 Induction
in human 8 Induction
cynomolgus Relative to
HT1080, Relative to
ABP n HT1080, GITRL GITRL in n
GITRL on GITRL in human
on same plate cynomolgus
same plate HT1080, GITRL
(PM) HT1080, GITRL
(PM) on same plate
on same plate
1 118 97 2 104 91 2
2 126 97 2 61 80 2
3 97 96 2 72 92 2
4 92 85 2 44 102 2
95 2 2
107 62 103
6 83 96 2 77 92 2
7 217 124 2 128 95 2
8 255 100 2 201 89 2
GITRL 120 - 16 76 - 16
[431] The same assay was conducted using IgG1 N297A non-TM versions of ABPs 1-
8
(see FIG. 4) and the data are summarized in Table 9.
Table 9. Activity of IgG1 N297A antibodies on human and cynomolgus GITR on
HT1080 cells
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ABP Average ECso
(Corresponding Average ECso in
TM ABP) in human cynomolgus
HT1080, HT1080,
GITRL on 1 GITRL on 1
separate plate separate plate
(nM) n (nM)
35(1) 0.88 1 2 1
36 (2) 0.58 1 8.8 1
37(3) 0.19 1 1.7 1
38 (4) 0.33 1 14.3 1
39 (6) 0.44 1 2.3 1
40(7) 0.34 1 1.1 1
41(8) 0.44 1 2 1
GITRL 0.43 2 0.17 2
[432] The assay was again used for testing the activity of additional N-
terminal Fab and C-
terminal Fab TM format antibodies as well the IgG1 N297A versions of those
antibodies.
The data are summarized in Table 10 and Table 11 and shown in FIG. 5. FIG. 5A
shows
ABP43 (squares), ABP23 (circles), ABP24 (triangles), and ABP29 (open circles),
ABP30
(open triangles), ABP31 (open circles), and ABP32 (open triangles), all in
comparison to
GITRL (+ sign). FIG. 5B shows ABP19 (N-terminal Fab, triangles) and ABP25 (C
terminal Fab, upside down triangles). GITRL is shown as plus signs. FIG. 5C
shows
ABP21 (N-terminal Fab, triangles) and ABP27 (C terminal Fab, upside down
triangles).
GITRL is shown as plus signs. FIG. 5D shows ABP20 (N-terminal Fab, triangles)
and
ABP26 (C terminal Fab, upside down triangles). GITRL is shown as plus signs.
FIG. 5E
shows ABP22 (N-terminal Fab, triangles) and ABP28 (C terminal Fab, upside down
triangles). GITRL is shown as plus signs. As shown in the FIG., the N-terminal
Fab
format ABPs tended to induce more IL-8 than their C-terminal Fab counterparts.
[433] FIG. 10 shows an additional comparison of ABP43 (non-optimized IgG4
format)
with two corresponding TM format ABPs where the IgG1 Fab counterpart of ABP43
is
added to the N or C terminus of the ABP43. As can be seen in the Figure, the N-
terminal
Fab TM format ABP9 (squares) induced the most IL-8 (and had the best EC50) in
comparison to the C-terminal Fab TM format ABP10 (circles) and the non-TM
format
ABP43 (diamonds). Both TM format ABPs had better activity than the non-TM
format
parental ABP as well as GITRL. IL-8 induction by GITRL (positive control) is
shown as a
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single data point (star) IL-8 production is shown in pg/mL.
Table 10. Agonistic activity of N-terminal Fab and C-terminal Fab TM format
antibodies on human
GITR on HT1080 cells
Average EC50 in Human
ABP Format HT1080,
GITRL on 1 n
separate plate (nM)
GITRL -- 0.37 1
9 N terminal Fab TM 0.18 1
19 N terminal Fab TM 0.25 1
20 N terminal Fab TM 0.63 1
21 N terminal Fab TM 0.18 1
22 N terminal Fab TM 0.30 1
23 N terminal Fab TM 0.30 1
24 N terminal Fab TM 0.17 1
25 C terminal Fab TM, linker = 15mer 0.46
26 C terminal Fab TM, linker = 15mer 0.63 1
27 C terminal Fab TM, linker = 15mer 0.35 1
28 C terminal Fab TM, linker = 15mer 0.63 1
29 C terminal Fab TM, linker = 15mer 0.27 1
30 C terminal Fab TM, linker = 15mer 0.31 1
31 C terminal Fab TM, linker = 5mer 0.18 1
32 C terminal Fab TM, linker = 5mer 0.30 1
Table 11. Agonistic activity of IgG1 N297A antibodies format antibodies on
human GITR on
HT1080 cells
EC50 in human EC50
in cynomolgus
HT1080, GITRL on
HT1080, GITRL on
ABP Format
separate plate (nM), n
separate plate (nM), n
=2 =1
42 IgG1 N297A 17.8 no activity
44 IgG1 N297A 4.2 28.0
45 IgG1 N297A 10.5 15.1
46 IgG1 N297A 1.2 6.9
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47 IgG1 N297A 4.1 11.8
48 IgG1 N297A 0.8 minimal activity
49 IgG1 N297A 2.7 65.8
[434] Additional IgG1 N297A versions of H1H2NH optimized antibodies are
provided and
shown below in Table 12 and their agonistic activity was determined in the
HT1080 assay,
as described above, is indicated. These antibodies are suitable to be made
into TM format.
Table 12. Agonistic activity of IgG1 N297A antibodies format antibodies on
human GITR on
HT1080 cells
EC50 in human EC50
in cynomolgus
ABP Format HT1080, GITRL on
HT1080, GITRL on
separate plate (nM)
separate plate (nM)
11 IgG1 N297A 3.9 21.6
12 IgG1 N297A 3.5 28.4
13 IgG1 N297A 7.8 15.0
14 IgG1 N297A 0.6 4.1
15 IgG1 N297A 0.6 4.5
16 IgG1 N297A 0.6 3.8
17 IgG1 N297A 5.3 8.4
18 IgG1 N297A 2.1 13.2
GITRL 0.4 0.2
[435] A
similar assay was performed in Jurkat cells, a T-cell based cell line,
engineered to
express human GITR and an NF-KB luciferase reporter (Promega0). Eight
optimized, N-
terminal Fab TM format antibodies, ABPs 1-8, were compared to GITRL for their
ability
to induce IL-8 in T cells. As shown in FIG. 7A-H (for ABPs 1-8) and Table 13,
all eight
ABPs were better than GITRL at inducing cytokine production.
Table 13. 8 Optimized TM format with N-terminal Fab are superior to GITRL
% Activity Compared to
ABP EC50 (nM)
GITRL
1 0.30 104
GITRL 0.90
2 0.21 114
GITRL 0.91
3 0.19 120
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GITRL 0.85
4 0.22 117
GITRL 1.13
0.25 104
GITRL 1.01
6 0.29 121
GITRL 0.90
7 0.20 108
GITRL 0.81
8 0.30 114
GITRL 0.89
Agonistic activity of N-Terminal Fab TM-Format Antibodies in Primary Cells
[436] In one embodiment, the GITR ABPs are further tested for their ability
to deliver a co-
stimulatory signal to CD4+CD25- effector T cells in conjunction with TCR
signaling via
anti-CD3 antibody. Increased T cell activation is measured by quantifying cell
proliferation, and/or measuring the increased production and secretion of
cytokines
including IFN-gamma using an ELISA assay.
[437] In another embodiment, the GITR ABPs are further analyzed to evaluate
their ability
to prevent the suppression of effector T cells by regulatory T cells. Human
CD4+ T cells
are isolated using a human CD4+ T cell isolation kit (Miltenyi #130-096).
Regulatory T
cells are further enriched using human CD25 MicroBeads0 II (Miltenyi #130-092-
983)
following the manufacturer's instructions. The CD25 depleted CD4+ T cells are
used as
effector T cells in the suppression assay. Beads conjugated to anti-human CD2,
CD3, and
CD28 antibodies are used to activate the effector T cells in the assay (Treg
Suppression
Inspector; a.k.a. T cell activation beads, Miltenyi #130-092-909). In a 96-
well plate,
50,000 each of regulatory T cells and effector T cells are seeded in each well
and
incubated with an equal number of T cell activation beads. The mixtures of
cells and beads
are contacted with different concentrations of GITR ABP for five days.
Proliferation of
effector T cells is measured by pulsing the cell cultures with tritiated
thymidine during the
last 16 hours of culture, washing, and measuring the amount of radioactivity
incorporated
into the cells.
Agonistic activity of N-Terminal Fab TM-Format Antibodies in T-blast Primary
Cell Assay
Pre-stimulated T cells (T-blasts) were used to assess the activity of eight N-
Terminal Fab TM
format antibodies (ABPs 1-8).
To generate T-blasts, PBMCs are stimulated with PHA for 5-7 days in order to
expand the
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cells and induce expression of GITR. Cells are washed and frozen prior to the
setup of the functional
assay.
To assess functional activity of ABPs 1-8, cells are stimulated by adding 1
mg/ml anti-CD3 +
2 mg/ml anti-CD28, 0.003 - 10 mg/ml ABPs 1-8, and 10 mg/ml GITRL as a positive
control. Cells
are incubated at 37 C and supernatants are collected after 48 hours. IL-2
production is measured by
AlphaLISA0 (Perkin Elmer ). FIG. 8A shows the percentage of GITR+ CD4+ cells
(left) and CD8+
cells (right) in cells from two human donors at various time points, +1-
stimulation with PHA.
FIGs 8B-8M show results of treatment of T-blasts from 4 different donors
treated with
controls or ABPs and the resultant IL-2 production. In each FIG. the top row,
from left to right is
FACS measurement of ABO binding to CD4+ cells, IL-2 production in cells from
Donor 1, IL-2
production in cells from Donor 2. In the bottom row, from left to right, is
FACS measurement of
ABP binding to CD8+ cells, IL-2 production in cells from Donor 3, IL-2
production in cells from
Donor 4. Shown are data as follows: 8B: IgG4 isotype control; 8C: SEC4
antibody; 8D: IgG4 TM
format control; 8E: non-TM IgG4 non-optimized lineage parent of ABPs 1-8
(ABP9). 8F: ABP1; 8G:
ABP2; 8H: ABP3; 81: ABP4; 8J: ABP5; 8K: ABP6; 8L: ABP7; 8M:ABP8.
As shown in the FIG., T-blasts from different donors varied in their response
to treatment
with ABPs 1-8; however, treatment with all eight TM format ABPs had agonistic
activity at most
concentrations tested, as measured by production of IL-2 and as compared to
control antibodies.
GITR Agonistic Activity Comparison of Optimized N-Terminal Fab TM Format ABPs
and Non-
TM Format IgG1 ABPs
[438] The same eight optimized N-terminal Fab TM format ABPs (1-8) as in
the above
Examples were compared to the non-optimized parental controls in the same IL-8-
induction assay described above and shown in FIG. 4. Each FIG. shows a
comparison of
the N-terminal Fab TM format ABPs 1-8 (IgG4 5228P with N-terminal IgG1 Fab,
"IgG4
TM") and the corresponding non-TM format IgG1 (IgG1 N297, "IgGl") ABP, as well
GITRL as a positive control and IgG4 negative control (GITRL and IgG4 run on a
separate
plate from the ABPs). The left panels show results from cells expressing hGITR
and the
right panels show results from cells expressing cGITR. IL-8 induction as a
function of
ABP concentration is shown for ABP 1 (FIG. 9A), ABP 2 (FIG. 9B), ABP3 (FIG.
9C),
ABP4 (FIG. 9D), ABP5 (FIG. 9E), ABP6 (FIG. 9F), ABP7 (FIG. 9G), and ABP8 (FIG.
9H). Induction of IL-8 by GITRL is shown as circles, by TM format antibodies
as
triangles, by parental IgG1 antibodies as diamonds, and by IgG4 control
antibodies as
squares. A table of EC50 values is shown on the bottom of each panel of each
FIG.
[439] The results shown in Figure 9 demonstrate that the eight optimized
ABPs are able to
induce IL-8 production in cells expressing human or cynomolgus GITR to a much
greater
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extent as N-terminal Fab TM format antibodies when compared to bivalent IgG1
N297A.
The N-terminal Fab TM format antibodies had smaller EC50 values and greater
maximum
production of IL-8 as compared to the bivalent IgG1 N297A format. In addition,
all of the
TM antibodies had greater affinity for GITR than the non-TM form of the
parental
antibody (compare Table 5 and 7 with Table 6).
Evaluation of GITRL Blockade
[440] In another embodiment, GITR blockade was evaluated using a FORTEBIO
OCTET
instrument. The analysis was performed at 30 C using lx Kinetics Buffer
(ForteBio, Inc.)
as assay buffer. Anti-human IgG Fc Capture (AHC) biosensors (ForteBio, Inc.)
are used to
capture human GITR ABPs onto the sensors. Sensors are saturated in assay
buffer for 300
seconds before the assay. ABPs were loaded onto sensors by dipping the sensors
into ABP
supernatant solution for 300 seconds. Baseline was established by dipping the
sensors into
lx assay buffer for 200 seconds. Next, association of recombinant GITR was
monitored
for 180 seconds. The ability of recombinant GITRL to then associate with the
ABP /GITR
complex was the determined by dipping the sensors in GITRL for 180 seconds.
[441] GITR blockade was evaluated according to the methods above using ABPs
1-8. All
eight ABPs were capable of blocking GITRL binding.
[442] In one embodiment, to evaluate whether anti-human GITR ABPs block GITR
ligand
(GITRL, R&D Systems #6987-GL-CF) binding to GITR, various concentrations of
unlabeled GITR ABP are incubated with human pan T cells at 4 C in staining
buffer (e.g.,
phosphate buffered saline with 0.5% bovine serum albumin) for 10 min. Without
washing,
4 nM of HA-tagged human GITRL is added and incubated at 4 C for another 30
min. The
cells are then washed twice with staining buffer, and incubated with anti-HA
PE antibody
(Miltenyi #130-092-257) in staining buffer at 4 C for 30 min. The cells are
then washed
twice with staining buffer and fixed with 2% paraformaldehyde in PBS for flow
cytometry
analysis. A decrease in the amount of PE staining indicates that the ABP
blocks the GITR-
GITRL interaction.
Example 3: Multispecific Antigen-Binding Proteins
[443] One embodiment of multispecific GITR agonistic ABPs comprises a
common light
chain antibody. ABPs were identified that bind two distinct epitopes on GITR.
These two
ABPs share the same light chain. ABP61 is an agonistic antibody and ABP59 has
less
agonistic activity but does not compete with ABP61 for binding to GITR.
Multivalent
multispecific antibodies with common light chains were produced by
transfecting a HEK-
293 host cell with vectors encoding two heavy chains and the single common
light chain.
The first exemplary ABP disclosed herein with a common light chain (SEQ ID
NO:125) is
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ABP33, which has an IgG4 heavy chain (ABP61) with N-terminal Fab (from ABP58
(IgG1 counterpart of ABP59)), and the full-length sequence SEQ ID NO:124 (HC)
and
SEQ ID NO:125 (LC). The second exemplary ABP disclosed herein with a common
light
chain is ABP34, which has a IgG4 heavy chain (ABP61) with C-terminal Fab (from
ABP58 (IgG1 counterpart of ABP59), and the full-length sequence SEQ ID NO:136
(HC)
and SEQ ID NO:125 (LC).
[444] Each multispecific ABP binds two distinct epitopes on GITR. The ABPs
are
characterized as described in the Examples above. Other multispecific ABPs
described
herein are produced and similarly characterized. FIG. 6 shows the results of
EC50
determination for ABP33 and ABP34 in the HT1080 assay as described above in
Example
2. ABP33 (tetravalent version combining the IgG4 ABP61 with the IgG1 Fab of
ABP58
[IgG1 counterpart of ABP591 on the N-terminus) and ABP34 (tetravalent version
combining the IgG4 of ABP61 with the IgG1 Fab of ABP58 [IgG1 counterpart of
ABP591
on the C-terminus) were compared to bivalent ABPs 59 and 61 (IgG4 5228P). As
shown
in the FIG., the tetravalent antibodies both had superior EC50, as measured by
IL-8
induction, when compared to their bivalent counterparts.
Example 4: Comparison of TM-Format Antibodies to Benchmark anti-GITR
antibodies
[445] The activity of N-terminal Fab TM-format antibodies was tested
against two known
anti-GITR antibodies, SEC4 and SEC9 in HT1080 cells that were engineered to
stably
express human GITR. Cultured cells were treated for six hours with a range of
concentrations of two benchmark agonist antibodies SEC4 ("35E6" formatted to
have
mouse variable regions with human IgG4 S228P/kappa regions, orange diamonds)
and
SEC9 (humanized 6C8 N62Q IgG1 N297A). SEC4 is described, e.g., in U.S. Patent
No.
8,709,424; variable regions are found in SEQ ID NOs 1 and 12. SEC9 is
described, e.g.,
in U.S. Patent No. 7,812,135; full length sequences are set forth in SEQ ID
NOs 58 and 63.
A bivalent antibody dose of 1pg/m1 is equivalent to 6.67nM; a tetravalent
antibody dose of
1 pg/m1 is equivalent to 4 nM.
[446] Induction of IL-8 was measured by ELISA and the EC50 was calculated.
FIG. 11A
shows a comparison of hGITRL to SEC4 (diamonds) and SEC9 (circles), as well as
an
IgG4 negative control (closed triangles), an IgG1 negative control (open
triangles), and
trimeric human GITR ligand ("hGITRL", squares) as a positive control. GITRL
had a
better EC50 (inset) and max induction compared to both SEC4 and SEC9.
[447] Also shown are comparisons to GITRL with ABP1 (FIG. 11B), ABP2 (FIG.
11C),
ABP3 (FIG. 11D), ABP4 (FIG. 11E), ABP5 (FIG. 11F), ABP6 (FIG. 11G), ABP7 (FIG.
11H), and ABP8 (FIG. 11I). As shown in the Figure, all eight N-Terminal Fab TM
format
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antibodies had a more favorable EC50 than did either SEC4 or SEC9. All eight N-
Terminal
Fab TM format antibodies also had a greater induction of IL-8 production than
either
SEC4 or SEC9.
[448] SEC4 was also tested as described above in the T-blast primary cell
assay. As shown
in FIG. 8, induction of IL-2 in primary cells by SEC4 was less for all four
donors than that
of the TM format antibodies.
Example 5: Anti-GITR ABPs increase production of cytokines in patient Tumor
Infiltrating
Lymphocytes and GITR+ T cells
Materials and Methods
[449] Dissociated human tumor samples were purchased from Conversant Bio.
Samples
were NSCLC adenocarcinoma isolated from a 75-year-old male, a previous smoker,
with
Stage Ia disease, prior to any treatment. The sample was quickly thawed, then
restimulated with several conditions, with and without immunotherapies as
follows: cells
were either unstimulated controls, or were stimulated with lug/mL aCD3
(Soluble) +2
ug/mL aCD28 (Soluble) + IL-2 (50ng/mL); cells either received no immunotherapy
treatment (for assessment of checkpoint protein levels), pembrolizumab (10
g/mL), TM
format ABP control (2 g/mL), ABP1 (2 g/mL), or ABP1 + pembrolizumab. Cells
were
incubated for 48 hours before supernatants were collected and stained for
expression of
checkpoints. In some samples, brefeldin A (an inhibitor of cytokine secretion)
was added
to the last five hours of stimulation for detection of cytokines by
intracellular cytokine
staining.
Results
[450] Cells were gated on GITR-positive T cells and the results are shown
in Figure 12A
(TNF production) and Figure 12B (IFNy production). As shown in the Figures,
anti-GITR
(ABP1) single agent treatment resulted in an increase in cytokine production.
There was
not a significant increase in this assay of cytokine production in cells
receiving the
combination ABP1 + pembrolizumab treatment.
Example 6: Mutational analysis for epitope determination: Alanine Scanning
[451] To identify the epitope for ABP1 binding to human GITR, single point
mutations
were made in the human GITR extra cellular domain to determine if APB1 binding
was
reduced. Either alanine substitutions or murine specific residues were used
(ABP1 does
not bind mouse GITR). Proteins were expressed in HEK-293 cells with an Fc tag,
secreted
as soluble protein, purified on MabSelectO Sure Lx resin, and characterized
by SDS-
PAGE. Binding was assessed by Bio-Layer Interferometry (BLI) using the Octet
platform.
Wild type human GITR-Fc or GITR-Fc mutant was captured on anti-human Fc
sensors,
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washed, and exposed to ABP1 Fab. Residues considered part of the binding
epitope
demonstrated reduced or no binding (e.g., a KD more than 3-fold poorer than
that of
binding to wild type human GITR). Alanine substitution at residues C58, R59,
Y61, E64,
C67 and an aspartic acid substitution at position C66 resulted in no binding,
while alanine
substitution at residues R56, D60, P62 or E65 resulted in reduced binding.
Example 7: In Vivo Evaluation of GITR Antigen-Binding Proteins
[452] In vivo studies are performed to demonstrate the capability of the
ABPs to decrease
circulating regulatory T cell numbers in a humanized NSG mouse model. Neonate
NSG
mice are transplanted with CD34+ human fetal liver cells by retro-orbital
injection. Over
16 weeks, the animals develop a diverse human immune cell repertoire including
CD4+
and CD8+ effector T cells, and regulatory T cells. A single intraperitoneal
dose of 25
mg/kg anti-human GITR ABP or human isotype control is given to the mice and
the
percent of circulating human CD4+ T cells expressing the regulatory T cell
marker FoxP3
is determined by flow cytometry on day 4 after dosing.
Example 8. Incubation of activated T cells with TM Format ABP1 or its
conventional format
parent ABP35
Preparation of activated T blasts
[453] Activated T blasts were generated by stimulating freshly isolated
PBMCs from two
healthy donors for 7 days with PHA (final conc. 10 g/ml) and IL-2 (final
conc. 4 ng/ml,
added only during last 24h) at 37 C & 5% CO2.
Internalization of GITR after antibody binding
[454] Activated CD4+/CD8+ T blasts were seeded at 2x105 per well in 96-well
U bottom
plates. Wells were treated with medium alone, recombinant GITR-Ligand (10
ug/ml, R&D
Systems, Cat# 6987-GL-025/CF), ABP1 (-250 kDa), hIgG4 TM Format isotype
control,
ABP35 (-150kDa), or hIgG1 standard format isotype control at nine doses each:
10
ug/mL, 2 ug/mL, 0.4 ug/mL, 80 ng/mL, 16 ng/mL, 3.2 ng/mL, 0.64 ng/mL, 0.13
ng/mL,
and 0.026 ng/mL.
Internalization of GITR after antibody binding
[455] Antibody-mediated clustering promotes endocytosis and signaling of
TNF receptor
superfamily members such as GITR. The ability of ABP1 and ABP35 (as well as
the
isotype controls) to mediate clustering and internalization was measured.
[456] The activated T blasts were first stained for FACS sorting. Fc
receptors were blocked
with human TruStain0 FcX for 10 minutes at room temperature. Cells were
incubated
with fluorescent-conjugated antibodies for 30 minutes at 4 C as in Table 14.
Cells were
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then washed 2X with FACS buffer, fixed in paraformaldehyde for 30 minutes in
the dark,
washed again, and resuspended in 200 ul FACS buffer and acquired on a BD
Fortessa
instrument.
[457] GITR
internalization was measured in CD4+ cells (Figures 13A-E) and CD8+ cells
(Figures 13F-J) from either Donor 1 (Figures 13A-B, 13F-G) or Donor 2 (Figures
13C-D,
13H-I). Cells were treated with either ABP1 TM format antibody or ABP35
standard
bivalent antibody. As can be observed, incubation with either ABP1 or ABP35
inhibits the
subsequent staining with ABP1Dylight650 (Figures 13A, 13C, 13F, 13H) but only
incubation with ABP1 induces GITR internalization as measured by staining with
non-
competitive clone 108-17 (Figures 13B, 13D, 13G, 131). A determination of the
EC50 of
ABP1 on cells from both donors is shown in Figure 13E (CD4+ cells) and Figure
131
(CD8+ cells).
Table 14. Fluorochrome labeling
Antigen Clone Isotype control Fluorochrome
CD3 UCHT-1 PE/Cy 7
CD4 OKT4 BV421
CD8 RPA-T8 FITC
GITR "ABP1" hIgG4 TM format Dylight-650
GITR 108-17 mIgG2a BV605
Cytokine production in activated T cells treated with ABP1 or ABP35
[458] Activated CD4+/CD8+ T blasts were seeded at 5x104 per well in 96-well U
bottom plates.
Wells were treated with medium alone, recombinant GITR-Ligand (10 ug/ml, R&D
Systems,
Cat# 6987-GL-025/CF), ABP1, hIgG4 TM Format isotype control, ABP35, or hIgG1
standard
format isotype control at nine doses each: 10 ug/mL, 2 ug/mL, 0.4 ug/mL, 80
ng/mL, 16 ng/mL,
3.2 ng/mL, 0.64 ng/mL, 0.13 ng/mL, and 0.026 ng/mL.
[459] Cells were stimulated by adding 1 ug/m1 anti-CD3 antibodies (mouse
anti-human,
clone UCHT-1), R&D Systems, Cat# MAB100) and 2 ug/m1 anti-CD28 (mouse anti-
human, clone 37407), R&D Systems, Cat# MAB342) antibodies. Assays were
incubated at
37 C & 5% CO2 for 48h. IL-2 production was measured by AlphaLISA0 (Perkin
Elmer).
[460] Incubation of T blasts with the TM format antibody ABP1, but not the
same antibody
in standard bivalent format (ABP35) or either of the isotype controls, leads
to
internalization of GITR by activated CD4+ and CD8+ T cells (FIG. 13).
[461] In addition, superclustering of the GITR receptor by ABP1, but not
binding by a
conventional bivalent ABP, promotes IL-2 secretion by activated T blasts in a
dose-
dependent manner (FIG. 14). Together, these data show that T blasts that don't
respond to
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conventional anti-GITR therapy will respond to the corresponding TM format
antibody.
Example 9: Cytokine production in activated T cells treated with ABP1 in
comparison with
two benchmark antibodies, SEC4 and SEC9
[462] Activated T blasts were prepared as described in Example 8, and used to
compare the activity
of ABP1 with benchmark anti-GITR antibodies SEC4 and SEC9. Activated T blasts
were
generated by stimulating freshly isolated PBMCs from two healthy donors for 7
days with PHA
(final conc. 10 g/ml) and IL-2 (final conc. 4 ng/ml, added only during last
24h) at 37 C & 5%
CO2.
[463] Activated CD4+/CD8+ T blasts were seeded at 5x104 per well in 96-well U
bottom plates.
Wells were treated with medium alone, recombinant GITR-Ligand, ABP1, SEC4,
SEC9, hIgG4
TM Format isotype control ("IsoTM"), hIgG1 standard format isotype control,
and hIgG4
standard format isotype control, at nine doses each: 10 ug/mL, 2 ug/mL, 0.4
ug/mL, 80 ng/mL,
16 ng/mL, 3.2 ng/mL, 0.64 ng/mL, 0.13 ng/mL, and 0.026 ng/mL.
Cytokine production in activated T cells treated with ABP1, SEC4, and SEC9
[464] Cells were stimulated by adding 1 ug/m1 anti-CD3 antibodies and 2 ug/m1
anti-CD28
antibodies. Cells were incubated at 37 C & 5% CO2 for 48h. IL-2 production was
measured by
AlphaLISA0 (Perkin Elmer). As seen in Example 8, TM format ABP1 promotes IL-2
secretion
by activated T blasts in a dose-dependent manner. IL-2 production from
activated T cells is
shown in Figure 15A (Donor 1) and 15B (Donor 2). The corresponding EC50 is
shown in
Figure 15C (Donor 1) and 15D (Donor 2).
[465] As can be seen in the Figures, SEC4 and SEC9 don't induce IL-2
efficiently in a dose
dependent manner, ABP1 did induce IL-2 production to a greater extent than
SEC4 and SEC9 in
a clear dose dependent manner. Together, these data support that T blasts that
don't respond to
conventional anti-GITR therapy will respond to the corresponding TM format
antibody.
EQUIVALENTS
[466] The disclosure set forth above may encompass multiple distinct
inventions with
independent utility. Although each of these inventions has been disclosed in
its preferred
form(s), the specific embodiments thereof as disclosed and illustrated herein
are not to be
considered in a limiting sense, because numerous variations are possible. The
subject
matter of the inventions includes all novel and nonobvious combinations and
subcombinations of the various elements, features, functions, and/or
properties disclosed
herein. The following claims particularly point out certain combinations and
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subcombinations regarded as novel and nonobvious. Inventions embodied in other
combinations and subcombinations of features, functions, elements, and/or
properties may
be claimed in this application, in applications claiming priority from this
application, or in
related applications. Such claims, whether directed to a different invention
or to the same
invention, and whether broader, narrower, equal, or different in scope in
comparison to the
original claims, also are regarded as included within the subject matter of
the inventions of
the present disclosure.
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APPENDIX A: SEQUENCES
SEQ Molecule Description SEQUENCE
ID
NO
1 Human NP_004186.1 MAQHGAMGAFRALCGLALLCALSLGQRPTGGPGCGPGRLLL
GITR (ref seq) or GTGTDARCCRVHTTRCCRDYPGEECCSEWDCMCVQPEFHCG
Q9Y5U5 DPCCTTCRHHPCPPGQGVQSQGKFSFGFQCIDCASGTFSGGHE
(UniProt) GHCKPWTDCTQFGFLTVFPGNKTHNAVCVPGSPPAEPLGWL
TVVLLAVAACVLLLTSAQLGLHIWQLRSQCMWPRETQLLLE
VPPSTEDARSCQFPEEERGERSAEEKGRLGDLWV
2 Human MAQHGAMGAFRALCGLALLCALSLGQRPTGGPGCGPGRLLL
GITR GRGTDARCCRVHTTRCCRDYPGEECCSEWDCMCVQPEFHCG
T43R DPCCTTCRHHPCPPGQGVQSQGKFSFGFQCIDCASGTFSGGHE
SNP GHCKPWTDCTQFGFLTVFPGNKTHNAVCVPGSPPAEPLGWL
variant TVVLLAVAACVLLLTSAQLGLHIWQLRSQCMWPRETQLLLE
VPPSTEDARSCQFPEEERGERSAEEKGRLGDLWV
3 Cynomol XP_00554518 MCACGTLCCLALLCAASLGQRPTGGPGCGPGRLLLGTGKDA
gus 0.1 RCCRVHPTRCCRDYQSEECCSEWDCVCVQPEFHCGNPCCTTC
monkey QHHPCPSGQGVQPQGKFSFGFRCVDCALGTFSRGHDGHCKP
GITR WTDCTQFGFLTVFPGNKTHNAVCVPGSPPAEPPGWLTIVLLA
VAACVLLLTSAQLGLHIWQLGSQPTGPRETQLLLEVPPS lEDA
SSCQFPEEERGERLAEEKGRLGDLWV
4 Mouse MGAWAMLYGVSMLCVLDLGQPSVVEEPGCGPGKVQNGSGN
GITR NTRCCSLYAPGKEDCPKERCICVTPEYHCGDPQCKICKHYPC
QPGQRVESQGDIVFGFRCVACAMGTFSAGRDGHCRLWTNCS
QFGFLTMFPGNKTHNAVCIPEPLPTEQYGHLTVIFLVMAACIF
FLTTVQLGLHIWQLRRQHMCPRETQPFAEVQLSAEDACSFQF
PEEERGEQ lEEKCHLGGRWP
Linker 1 GGGGS
6 Linker 2 GGGGSGGGGSGGGGS
7 ABP1 Heavy Chain QVQLQESGPGLVKPSETLSLTCAVSGYSISSGLGWGWIRQPPG
Full, heavy KGLEWIGGIYESGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
variable 1 + AADTAVYYCAHERVRGYGDYGGHHAFDIWGQGTMVTVSSA
IgG1 CH1 + STKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG
GGGGS + ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
heavy variable KPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLSLT
1 + IgG4 CAVSGYSISSGLGWGWIRQPPGKGLEWIGGIYESGSTYYNPSL
5228P (with KSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAHERVRGYGD
YGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTSEST
98
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or without C AALGCLVKDYFPEPVTVSWN S GALT S GVHTFPAVLQ SSGLYS
terminal Lys) L S SVVTVPS S SLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC
PP CPAPEFL GGP S VFLFPPKPKDTLMI SRTPEVTCVVVD VSQE
DPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLH
QDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLP
PSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT
PPVLD SD GSFFLYSRL TVDK SRWQEGNVF S C S VMHEALHNH
YTQKSL SL SLGK
17 ABP2
Heavy Chain QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVVVSWIRQHP
Full, heavy GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSS
variable 1 + VTAADTAVYYCADENVRGYGDYGGHHAFDIWGQGTMVTVS
IgG1 CH1 + SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
GGGGS + S GAL T
S GVHTFP AVLQ S SGLYSL S SVVTVPS S SL GTQTYICNV
heavy variable NHKP SNTKVDKRVEPK S C GGGG S QVQL QE S GP GLVKP SQTL S
1 + IgG4 LT CTVS GGS IS S GGAVVVSWIRQHPGKGLEWIGGIAYSGSTYY
S228P (with NP SLK SRVTI S VDT SKNQF SLKL S SVTAADTAVYYCADENVR
or without C GYGDYGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRST
terminal Lys) SESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQS S
GLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKY
GPPCPPCPAPEFL GGP S VFLFPPKPKD TLMI SRTPEVT CVVVD V
SQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVY
TLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLD SD GSFFLYSRL TVDK SRWQEGNVF S C S VMHEALH
NHYTQKSL SL SL GK
24 ABP3
Heavy Chain QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
Full, heavy
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
variable 1 + AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
IgG1 CH1 + STKGP S VFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
GGGGS + ALT S
GVHTFPAVL Q S SGLYSL S S VVTVP S S SL GTQTYICNVNH
heavy variable KPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLSLT
1 + IgG4
CAVSGYSIS SGAGWGWIRQPPGKGLEWIGLIVHSGSTYYNPSL
S228P (with KSRVTISVDTSKNQFSLKLSSVTAADTAVYYCVLESVRGYGD
or without C YGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTSEST
terminal Lys) AAL G CLVKDYFPEPVTVS WN S GALT S GVHTFPAVLQ S SGLYS
L S SVVTVPS S SLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC
PP CPAPEFL GGP S VFLFPPKPKDTLMI SRTPEVTCVVVD VSQE
DPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLH
QDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLP
PSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT
99
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PPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALHNH
YTQKSL SL SLGK
32 ABP 4
Heavy Chain QVQLQES GP GLVKP SETL SLTCAVSGYSIS SGAGWGWIRQPPG
Full, heavy
KGPEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
variable 1 + AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
IgG1 CH1 + STKGP S VFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
GGGGS + ALT S
GVHTFPAVL Q S SGLYSL S S VVTVP S S SL GTQTYICNVNH
heavy variable KPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLSLT
1 + IgG4
CAVSGYSIS SGAGWGWIRQPPGKGPEWIGLIVH SGSTYYNPSL
S228P (with KSRVTIS VDT SKNQF SLKL S S VTAADTAVYYCVLESVRGYGD
or without C YGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTSEST
terminal Lys) AAL G CLVKDYFPEPVTVS WN S GALT S GVHTFPAVLQ S SGLYS
L S SVVTVPS S SLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC
PP CPAPEFL GGP S VFLFPPKPKDTLMI SRTPEVTCVVVD VSQE
DPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLH
QDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLP
PSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT
PPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALHNH
YTQKSL SL SLGK
37 ABP5
Heavy Chain QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
Full, heavy
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
variable 1 + AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
IgG1 CH1 + STKGP S VFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
GGGGS + ALT S
GVHTFPAVL Q S SGLYSL S S VVTVP S S SL GTQTYICNVNH
heavy variable KPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLSLT
1 + IgG4
CAVSGYSIS SGAGWGWIRQPPGKGLEWIGLIVHSGSTYYNPSL
S228P (with KSRVTISVDTSKNQFSLKLSSVTAADTAVYYCVLESVRGYGD
or without C YGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTSEST
terminal Lys) AAL G CLVKDYFPEPVTVS WN S GALT S GVHTFPAVLQ S SGLYS
L S SVVTVPS S SLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC
PP CPAPEFL GGP S VFLFPPKPKDTLMI SRTPEVTCVVVD VSQE
DPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLH
QDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLP
PSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT
PPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALHNH
YTQKSL SL SLGK
38 ABP 6
Heavy Chain QVQLQES GP GLVKP SETL SLTCAVSGYSIS SGAGWGWIRQPPG
Full, heavy
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
variable 1 + AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
IgG1 CH1 + STKGP S VFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
100
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GGGGS + ALT S GVHTFPAVL Q S SGLYSL S SVVTVP S S SL GTQTYICNVNH
heavy variable KPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLSLT
1 + IgG4 CAVSGYSIS SGAGWGWIRQPPGKGLEWIGLIVHSGSTYYNPSL
S228P (with KSRVTISVDTSKNQFSLKLSSVTAADTAVYYCVLESVRGYGD
or without C YGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTSEST
terminal Lys) AAL G CLVKDYFPEPVTVS WN S GALT S GVHTFPAVLQ S SGLYS
L S SVVTVPS S SLGTKTYTCNVDHKPSNTKVDKRVESKYGPPC
PP CPAPEFL GGP S VFLFPPKPKDTLMI SRTPEVTCVVVD VSQE
DPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLH
QDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLP
PSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTT
PPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALHNH
YTQKSL SL SLGK
42 ABP7 Heavy Chain QVQLQESGPGLVKPSETLSLTCAVSGYSISSEYMVVGWIRQPP
Full, heavy GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
variable 1 + VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
IgG1 CH1 + SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
GGGGS + S GAL TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SL
GTQTYICNV
heavy variable NHKPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLS
1 + IgG4 LTCAVSGYSIS SEYMWGWIRQPPGKGLEWIGLIYHSGKTYYN
S228P (with PSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREADRG
or without C YGDYGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTS
terminal Lys) ESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQ S SG
LYSL S SVVTVPS S SL GTKTYTCNVDHKPSNTKVDKRVESKYG
PP CPP CPAPEFL GGP S VFLFPPKPKDTLMI SRTPEVTCVVVD VS
QEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTV
LHQDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYT
LPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYK
TTPPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALHN
HYTQKSLSLSLGK
51 ABP 8 Heavy Chain QVQLQES GP GLVKP SETL SLTCAVSGYSIS SEYMVVGWIRQPP
Full, heavy GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
variable 1 + VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
IgG1 CH1 + SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
GGGGS + S GAL TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SL
GTQTYICNV
heavy variable NHKPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLS
1 + IgG4 LTCAVSGYSIS SEYMWGWIRQPPGKGLEWIGLIYHSGKTYYN
S228P (with PSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREADRG
or without C YGDYGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTS
terminal Lys) ESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQ S SG
101
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LYSL S SVVTVPS S SL GTKTYTCNVDHKPSNTKVDKRVESKYG
PP CPP CPAPEFL GGP S VFLFPPKPKDTLMI SRTPEVTCVVVD VS
QEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTV
LHQDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYT
LPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYK
TTPPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALHN
HYTQKSLSLSLGK
57 ABP9 Heavy Chain QLQLQESGPGLVKPSETLSLTCTVSGGSISSSSYAWGWIRQPP
Full, heavy
GKGLEWIGSIYYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSV
variable 1 + TAADTAVYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS S
IgG1 CH1 + ASTKGP S VFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNS
GGGGS + GALT S
GVHTFPAVL Q S SGLYSL S SVVTVPS S SL GTQTYICNVN
heavy variable HKPSNTKVDKRVEPKSCGGGGSQLQLQESGPGLVKPSETLSL
1 + IgG4 TCTVS
GGSIS S S SYAWGWIRQPPGKGLEWIGSIYYSGSTYYNP
S 228P (with SLK SRVTI S VD T SKNQF SLKL S SVTAADTAVYYCARD SGRGY
or without C GDYGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTSE
terminal Lys) STAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGL
YSL S SVVTVPS S SL GTKTYTCNVDHKPSNTKVDKRVESKYGP
PCPPCPAPEFLGGPSVFLFPPKPKDTLM I SRTPEVTCVVVDVSQ
EDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVL
HQDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTL
PPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKT
TPPVLD SD GSFFLYSRLTVDKSRWQEGNVF S C S VMHEALHNH
YTQKSL SL SLGK
57 ABP10 Heavy Chain QLQLQESGPGLVKPSETLSLTCTVSGGSISSSSYAWGWIRQPP
Full, heavy
GKGLEWIGSIYYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSV
variable 1 + TAADTAVYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS S
IgG1 CH1 + ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNS
GGGGS + GALT S
GVHTFPAVL Q S SGLYSL S SVVTVPS S SL GTKTYTCNVD
heavy variable HKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKD
1 + IgG4
TLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTK
S228P (with PREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSI
or without C EKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPS
terminal Lys) DIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSL SL SL GKGGGGSQLQLQE
S GP GL VKP SETL SLT CTVS GG SI S S S SYAWGWIRQPPGKGLEW
IGSIYYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVTAADTA
VYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS S A S TKGP
S VFPL AP S SK S T S GGTAAL GCLVKDYFPEP VTVS WN S GALT S G
102
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VHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNT
KVDKRVEPKSC
171 ABP11
Heavy Chain QVQLVQSGAEVKRPGS SVKVS CKASGGTFS SYYISWVRQVPG
Full (IgG1)
QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
SLRSEDTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVS
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
S GAL TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SLGTQTYICNV
NHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLV
KGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSL SLSPGK
172 ABP12
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYYISWVRQAPG
Full (IgG1)
QRLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
SLRSEDTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVS
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
S GAL TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SLGTQTYICNV
NHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLV
KGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSL SLSPGK
173 ABP13 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSRGAISWVRQAP
Full (IgG1) GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTGTAYMEL
S SLR SED TAVYYCARQ SDYGLPRGMD VVVGQ GTTVTVS SAS T
KGPSVFPLAPS SKSTS GGTAAL GCLVKDYFPEPVTVSWNS GA
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
PSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPK
DTLMISRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKT
KPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPA
PIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFY
PSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKLTVDKSR
WQQGNVFSCSVMHEALHNHYTQKSL SLSPGK
174 ABP14
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
Full (IgG1)
QGLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYMEL S S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS SA STKGP
S VFPL AP S SK S T S GGTAAL GCLVKDYFPEP VTVS WNS GALT S G
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNT
103
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KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
ISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPRE
EQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFSCSVMHEALHNHYTQKSLSLSPGK
175 ABP15
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
Full (IgG1)
QRLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYMELSS
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
ISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPRE
EQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFSCSVMHEALHNHYTQKSLSLSPGK
176 ABP16
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
Full (IgG1)
QGLEWMGGIIPISGFTNYAQKFQGKVTITADESTSTAYMEL SS
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
ISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPRE
EQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFSCSVMHEALHNHYTQKSLSLSPGK
177 ABP17
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
Full (IgG1)
GQGLEWMGGIIPIFGEAQYAQRFQGRVTITADESTSTAYMELS
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
ISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPRE
EQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFSCSVMHEALHNHYTQKSLSLSPGK
104
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178 ABP18
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
Full (IgG1) GQ
GLEWMGGIIP IF GEAQYAQKFRGRATITADE S T S TAYMEL S
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVS SASTKGP
S VFPL AP S SK S T S GGTAAL GCLVKDYFPEP VTVS WN S GALT S G
VHTFPAVLQS SGLYSLS SVVTVPS S SL GTQTYI CNVNHKP S NT
KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
I SRTPEVTCVVVD VSHEDPEVKFNWYVD GVEVHNAKTKPRE
EQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPP SRDELTKNQVSL TCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFS CSVMHEALHNHYTQKSL SL SP GK
106 ABP19
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYYISWVRQAPG
Full, heavy
QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
variable 1 + SLRSEDTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVS
IgG1 CH1 + SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
GGGGS + S GAL
TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SL GTQTYICNV
heavy variable NHKPSNTKVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSV
1 + IgG4
KVSCKASGGTFS SYYISWVRQAPGQGLEWMGGIIPVPGTANY
S228P (with AQKFQGRVTITADESTSTAYMELS SLRSEDTAVYYCARAGGG
or without C YARGDHYYGMDVVVGQGTTVTVSSASTKGPSVFPLAPCSRST
terminal Lys) SESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQS S
GLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKY
GPPCPPCPAPEFL GGP S VFLFPPKPKD TLMI SRTPEVTCVVVD V
SQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVY
TLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALH
NHYTQKSLSL SL G
108 ABP20
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSRGAISWVRQAP
Full, heavy
GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTSTAYMEL
variable 1 + S SLRSEDTAVYYCARQSDYGLPRGMDVVVGQGTTVTVS SAS T
IgG1 CH1 + KGPSVFPLAPS SKSTS GGTAAL GCLVKDYFPEPVTVSWNS GA
GGGGS +
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
heavy variable PSNTKVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVS
1 + IgG4
CKASGGTFSRGAISWVRQAPGQGLEWMGGIIPIEGTAYYAQK
S228P (with FQGRVTITADESTSTAYMEL S SLRSEDTAVYYCARQSDYGLP
or without C RGMDVVVGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALG
terminal Lys) CLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSV
VT VP S S SL GTKTYTCNVDHKP SNTKVDKRVE SKYGPP CPP CP
APEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEV
105
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QFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDW
LNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLPPSQE
EMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVL
D SD GSFFLYSRL TVDK SRWQEGNVF S C S VMHEALHNHYTQK
SLSLSLG
110 ABP21
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
Full, heavy
QGLEWMGGIIPISGFANYAQKFQGRVTITADESTSTAYMELSS
variable 1 + LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
IgG1 CH1 + S VFPL AP S SKST S GGTAAL GCLVKDYFPEP VTVS WNS GALT S G
GGGGS +
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
heavy variable KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
1 + IgG4 GGTF S
SYAISWVRQAPGQGLEWMGGIIPISGFANYAQKFQGR
S 228P (with VTITADESTSTAYMEL S SLR SED TAVYYCAREGGHYY S GWPY
or without C WGQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKD
terminal Lys) YFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSVVTVPSS
SLGTKTYTCNVDHKP SNTKVDKRVESKYGPP CPP CP APEFL G
GP S VFLFPPKPKDTLMI SRTPEVT CVVVD VSQEDPEVQFNWY
VDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKE
YKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLPPSQEEMTKN
QVSLT CLVKGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GS
FFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSL SL SL
G
112 ABP22
Heavy Chain QVQLVQ S GAEVKKP GS SVKVSCKASGGTFVRYAISWVRQAP
Full, heavy
GQGLEWMGGIIPIFGEAQYAQKFQGRVTITADESTSTAYMELS
variable 1 + SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
IgG1 CH1 + S VFPL AP S SKST S GGTAAL GCLVKDYFPEP VTVS WNS GALT S G
GGGGS +
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
heavy variable KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
1 + IgG4
GGTFVRYAISWVRQAPGQGLEWMGGIIPIFGEAQYAQKFQGR
S 228P (with VTITADESTSTAYMEL S SLR SED TAVYYCARE GYYYGALPYW
or without C GQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYF
terminal Lys) PEP VTVS WNSGALTSGVHTFP AVLQ S SGLY SL SSVVTVPSSSL
GTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFL GGP
SVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD
GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKC
KVSNKGLPS SIEKTISKAKGQPREPQVYTLPP SQEEMTKNQVS
LT CLVKGFYP SDIAVEWESNGQPENNYKTTPPVLD SD GSFFLY
SRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSL SL SLG
114 ABP23
Heavy Chain QVQLQES GP GLVKP SETL SLTCAVSGYSIS SEYMVVGWIRQPP
Full, heavy
GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
106
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variable 1 + VTAADTAVYYCARDSGRGYGDYGGHHAFDIWGQGTMVTVS
IgG1 CH1 + SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
GGGGS + S GAL
TS GVHTFP AVLQ S SGLYSL S SVVTVPS S SL GTQTYICNV
heavy variable NHKPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLS
1 + IgG4
LTCAVSGYSIS SEYMWGWIRQPPGKGLEWIGLIYHSGKTYYN
S228P (with PSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCARDSGRG
or without C YGDYGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRSTS
terminal Lys) ESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQS SG
LYSL S SVVTVPS S SL GTKTYTCNVDHKPSNTKVDKRVESKYG
PP CPP CPAPEFL GGP S VFLFPPKPKDTLMI SRTPEVTCVVVD VS
QEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTV
LHQDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYT
LPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYK
TTPPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALHN
HYTQKSL SL SLG
115 ABP24
Heavy Chain QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVVVSWIRQHP
Full, heavy GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSS
variable 1 + VTAADTAVYYCARDSVRGYGDYGGHHAFDIWGQGTMVTVS
IgG1 CH1 + SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
GGGGS + S GAL
TS GVHTFP AVLQ S SGLYSL S SVVTVPS S SL GTQTYICNV
heavy variable NHKP SNTKVDKRVEPK S C GGGG S QVQL QE S GP GLVKP SQTL S
1 + IgG4 LTCTVS GGS IS S GGAVVVSWIRQHPGKGLEWIGGIAYSGSTYY
S228P (with NPSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCARDSVR
or without C GYGDYGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPCSRST
terminal Lys) SESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQS S
GLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKY
GPPCPPCPAPEFL GGP S VFLFPPKPKD TLMI SRTPEVTCVVVD V
SQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVY
TLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLD SD GSFFLYSRL TVDKSRWQEGNVF S C S VMHEALH
NHYTQKSL SL SL G
116 ABP25
Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYYISWVRQAPG
Full, heavy
QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
variable 1 + SLRSEDTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVS
IgG4 5228P SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
(with or S GAL
TS GVHTFP AVLQ S SGLYSL S SVVTVPS S SL GTKTYTCNV
without C
DHKPSNTKVDKRVESKYGPPCPPCPAPEFL GGPSVFLFPPKPK
terminal Lys) DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
+
KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
107
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GGGGSGGG SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
GS GGGGS + PSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLY SRL TVDK SR
heavy variable WQEGNVFSCSVMHEALHNHYTQKSLSLSLGGGGGSGGGGSG
1+ IgG1 CH1 GGGSQVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYYISWVR
QAPGQGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAY
MEL S SLR SED TAVYYCARAGGGYARGDHYYGMD VVVGQ GT
TVTVS SASTKGPSVFPLAPS SKSTSGGTAAL GCLVKDYFPEPV
TVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQT
YICNVNHKPSNTKVDKRVEPKS C
117 ABP26 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSRGAISWVRQAP
Full, heavy GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTSTAYMEL
variable 1 + S SLRSEDTAVYYCARQSDYGLPRGMDVVVGQGTTVTVS SAS T
IgG4 S228P KGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGAL
(with or TSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKP
without C SNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLM
terminal Lys) ISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPRE
+ EQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKT
GGGGSGGG ISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIA
GS GGGGS + VEWESNGQPENNYKTTPPVLD SD GSFFLY SRL TVDKSRWQEG
heavy variable NVFSCSVMHEALHNHYTQKSLSLSLGGGGGSGGGGSGGGGS
1 + IgG1 CH1 QVQLVQSGAEVKKPGSSVKVSCKASGGTFSRGAISWVRQAP
GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTSTAYMEL
S SLR SED TAVYYCARQ SDYGLPRGMD VVVGQ GTTVTVS SAS T
KGPSVFPLAPS SKSTSGGTAAL GCLVKDYFPEPVTVSWNS GA
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
PSNTKVDKRVEPKSC
118 ABP27 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
Full, heavy QGLEWMGGIIPISGFANYAQKFQGRVTITADESTSTAYMELSS
variable 1 + LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
IgG4 S228P SVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSG
(with or VHTFPAVLQS SGLYSL S SVVTVPS S SLGTKTYTCNVDHKPSNT
without C KVDKRVE SKYGPP CPP CPAPEFL GGP S VFLFPPKPKDTLMI SR
terminal Lys) TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
+ NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKTISK
GGGGSGGG AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYP SDIAVE
GS GGGGS + WE SNGQPENNYKTTPPVLD SD GSFFLYSRLTVDKSRWQEGN
heavy variable VFSCSVMHEALHNHYTQKSLSLSLGGGGGSGGGGSGGGGSQ
1+ IgG1 CH1 VQLVQ S GAEVKKP GS SVKVS CKASGGTFS SYAISWVRQAPGQ
GLEWMGGIIPI S GFANYAQKFQ GRVTITADE ST STAYMEL S SL
RSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS SASTKGPS
108
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VFPL AP S SKST S GGTAAL GCLVKDYFPEPVTVSWNS GALT S G
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
KVDKRVEPKSC
119 ABP28 Heavy Chain QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
Full, heavy GQGLEWMGGIIPIFGEAQYAQKFQGRVTITADESTSTAYMELS
variable 1 + SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
IgG4 S228P SVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSG
(with or VHTFPAVLQS SGLYSL S SVVTVPS S SLGTKTYTCNVDHKPSNT
without C KVDKRVE SKYGPP CPP CPAPEFL GGP S VFLFPPKPKDTLMI SR
terminal Lys) TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
+ NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKTISK
GGGGSGGG AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYP SDIAVE
GS GGGGS + WE SNGQPENNYKTTPPVLD SD GSFFLYSRLTVDKSRWQEGN
heavy variable VFSCSVMHEALHNHYTQKSLSLSLGGGGGSGGGGSGGGGSQ
1+ IgG1 CH1 VQLVQSGAEVKKPGS SVKVS CKASGGTFVRYAISWVRQAPG
QGLEWMGGIIPIFGEAQYAQKFQGRVTITADESTSTAYMEL S S
LRSEDTAVYYCAREGYYYGALPYWGQGTLVTVS S ASTK GP S
VFPL AP S SKST S GGTAAL GCLVKDYFPEPVTVSWNS GALT S G
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
KVDKRVEPKSC
120 AB P29 Heavy Chain QVQLQE S GP GLVKP S ETL SL TCAVS GY S I S
SEYMVVGWIRQPP
Full, heavy GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
variable 1 + VTAADTAVYYCARDSGRGYGDYGGHHAFDIWGQGTMVTVS
IgG4 S228P SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
(with or S GAL T S GVH TFP AVLQ S SGLYSL S SVVTVPS S SL
GTKTYTCNV
without C DHKPSNTKVDKRVESKYGPPCPPCPAPEFL GGPSVFLFPPKPK
terminal Lys) DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
+ KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
GGGGSGGG SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
GS GGGGS + PSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLY SRL TVDK SR
heavy variable WQEGNVFSCSVMHEALHNHYTQKSLSLSLGGGGGSGGGGSG
1+ IgG1 CH1 GGGSQVQLQESGPGLVKP SETLSLTCAVSGYSIS SEYMVVGWI
RQPPGKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSL
KL S SVTAADTAVYYCARD SGRGYGDYGGHHAFDIWGQGTM
VTVS S A S TKGP S VFPL AP S SKSTSGGTAALGCLVKDYFPEPVT
VSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTY
ICNVNHKPSNTKVDKRVEPKSC
121 ABP30 Heavy Chain QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVVVSWIRQHP
Full, heavy GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSS
variable 1 + VTAADTAVYYCARDSVRGYGDYGGHHAFDIWGQGTMVTVS
109
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IgG4 S228P SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
(with or S GAL T S GVH TFP AVLQ S SGLYSL S SVVTVPS S SL
GTKTYTCNV
without C DHKPSNTKVDKRVESKYGPPCPPCPAPEFL GGPSVFLFPPKPK
terminal Lys) DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
+ KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
GGGGSGGG SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
GS GGGGS + PSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLY SRL TVDK SR
heavy variable WQEGNVFSCSVMHEALHNHYTQKSLSLSLGGGGGSGGGGSG
1+ IgG1 CH1 GGGSQVQL QES GP GLVKP SQTL SLTCTVSGGSIS S GGAVVV SW
IRQHPGKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSL
KL S SVTAADTAVYYCARD SVRGYGDYGGHHAFDIWGQGTM
VTVS S A S TKGP S VFPL AP S SKSTSGGTAALGCLVKDYFPEPVT
VSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTY
ICNVNHKPSNTKVDKRVEPKSC
122 ABP31 Heavy Chain QVQLQES GP GLVKP SETL SL TCAVS GYSI S SEYMVVGWIRQPP
Full, heavy GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
variable 1 + VTAADTAVYYCARDSGRGYGDYGGHHAFDIWGQGTMVTVS
IgG4 S228P SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
(with or S GAL T S GVH TFP AVLQ S SGLYSL S SVVTVPS S SL
GTKTYTCNV
without C DHKPSNTKVDKRVESKYGPPCPPCPAPEFL GGPSVFLFPPKPK
terminal Lys) DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
+ GGGGS + KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
heavy variable SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
1 + IgG1 CH1 PSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSR
WQEGNVFSCSVMHEALHNHYTQKSL SL SLGGGGGSQVQLQE
S GP GL VKP SETL SLT CAVS GY S I S SEYMWGWIRQPPGKGLEWI
GL IYH S GKTYYNP SLK SRVTI S VDT SKNQF SLKL S SVTAADTA
VYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS S A S TKGP
S VFPL AP S SK S T S GGTAAL GCLVKDYFPEP VTVS WN S GALT S G
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
KVDKRVEPKSC
123 ABP32 Heavy Chain QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVVVSWIRQHP
Full, heavy GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSS
variable 1 + VTAADTAVYYCARDSVRGYGDYGGHHAFDIWGQGTMVTVS
IgG4 S228P SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
(with or S GAL T S GVH TFP AVLQ S SGLYSL S SVVTVPS S SL
GTKTYTCNV
without C DHKPSNTKVDKRVESKYGPPCPPCPAPEFL GGPSVFLFPPKPK
terminal Lys) DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
+ GGGGS + KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
110
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heavy variable PSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSR
1 + IgG1 CH1 WQEGNVFSCSVMHEALHNHYTQKSLSLSLGGGGGSQVQLQE
S GP GL VKP SQ TL SL TCTVS GG SI S SGGAVW SWIRQHPGKGLE
WIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVTAAD
TAVYYCARDSVRGYGDYGGHHAFDIWGQGTMVTVSSASTK
GP S VFPLAP S SK ST S GGTAAL GCL VKDYFPEPVTVS WNS GAL T
SGVHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYICNVNHKPS
NTKVDKRVEPKS C
124 ABP33
Heavy Chain QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYYMHWVRQA
Full, heavy
PGQGLEWMGIINPSGGSTTYAQKFQGRVTMTRDTSTSTVYME
variable 1 + L S SLRSEDTAVYYCARGQYGYYGGRLDVWGQGTTVTVS SAS
IgG1 CH1 + TKGPSVFPLAPS SK ST S GGTAAL GCL VKDYFPEPVTVS WNS G
GGGGS + ALT S
GVHTFPAVL Q S SGLYSL S S VVTVP S S SL GTQTYICNVNH
heavy variable KPSNTKVDKRVEPKSCGGGGSQVQLVESGGGVVQPGRSLRL
2+ IgG4 S CAA S
GFTF S SYGMHWVRQAPGKGLEWVAVISYDGSNKYYA
S228P (with DSVKGRFTISRDNSKNTLYLQMNSLRAEDTAVYYCARDGLG
or without C YMAWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALGCL
terminal Lys) VKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSL SSVVT
VP S S SL GTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAP
EFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQF
NWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLN
GKEYKCKVSNKGLPS SIEKTISKAKGQPREPQVYTLPPSQEEM
TKNQVSLTCLVKGFYP SD IAVEWE SNGQPENNYKTTPPVLD S
DGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSL
SLSLGK
136 AB P34
Heavy Chain QVQLVESGGGVVQPGRSLRL S CAA S GFTF S SYGMHWVRQ AP
Full, C GKGLEWVAVISYDGSNKYYAD SVKGRFTISRDNSKNTLYLQ
terminal Fab, MNSLRAEDTAVYYCARDGLGYMAWGQGTTVTVSSASTKGP
common light SVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSG
chain
VHTFPAVLQS SGLYSL S SVVTVPS S SLGTKTYTCNVDHKPSNT
bispecific: KVDKRVE SKYGPP CPP CPAPEFL GGP S VFLFPPKPKDTLMI SR
heavy variable TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
2+ IgG4
NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKTISK
5228P (with AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVE
or without C WESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGN
terminal Lys) VFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLVQSGAE
+ GGGGS + VKKPGASVKVSCKASGYTFTSYYMHWVRQAPGQGLEWMGII
heavy variable NPSGGSTTYAQKFQGRVTMTRDTSTSTVYMEL S SLRSEDTAV
1+ IgG1 CH1 YYCARGQYGYYGGRLDVWGQGTTVTVSSASTKGPSVFPLAP
S SK S T S GGTAAL GCL VKDYFPEPVTVSWNS GALT S GVHTFP A
111
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VLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRV
EPKSC
N TERMINAL FAB TM = HEAVY VARIABLE 1+ IGG1 CH1 + GGGGS + HEAVY VARIABLE 1+
IGG4 S228P (WITH OR WITHOUT C l'ERMINAL LYS)
8 ABP1
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
18 ABP2
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
25 ABP3
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full
PKLLIYAASSLKYGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
33 ABP4
Light Chain DIQMTQSPSSLSASVGDRVTITCRASKSIDSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
33 ABP5
Light Chain DIQMTQSPSSLSASVGDRVTITCRASKSIDSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
39 ABP6
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full
PKLLIYAADSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
43 ABP7
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSINSYLNWYQQKPGKA
Full PKLLIYAASSLDSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
112
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
VVCLLNNFYPREAKVQWKVDNALQ S GNSQES VTEQD SKD ST
YSL SSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
52 ABP 8
Light Chain DIQMTQ SP S SL SASVGDRVTITCGASQSISTYLNWYQQKPGKA
Full
PKLLIYSAS SLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYC
QQEYNTPPSFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASV
VCLLNNFYPREAKVQWKVDNALQ S GN SQESV 1EQD SKD STY
SLSSTLTLSKADYEKHKVYACEVTHQGL SSPVTKSFNRGEC
8 ABP9
Light Chain DIQMTQ SP S SL SASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAAS SLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQ S GNSQES VTEQD SKD ST
YSL SSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP10
Light Chain DIQMTQ SP S SL SASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAAS SLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQ S GNSQES VTEQD SKD ST
YSL SSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
172 ABP11
Light Chain DIVMTQ SPL SLPVTPGEPAS IS CRS SQ SLLH SNGYNYLDWYLQ
Full KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QDSKDSTYSL SSTLTLSKADYEKHKVYACEVTHQGL SSPVTK
SFNRGEC
174 ABP12
Light Chain DIVMTQ SPL SLPVTPGEPAS IS CRS SQ SLLH SNGYNYLDWYLQ
Full KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QDSKDSTYSL SSTLTLSKADYEKHKVYACEVTHQGL SSPVTK
SFNRGEC
176 ABP13
Light Chain DIQLTQ SP S SVSASVGDRVTITCRASQGIS SWLAWYQQKPGK
Full
APKLLIYGAS SLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATY
YCQQASLFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTA
SVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDS
TYSL SSTLTL SKADYEKHKVYACEVTHQGL SSPVTKSFNRGE
C
178 ABP14
Light Chain EIVLTQSPATL SLSPGERATL SCRASQSVS SYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARF S GS GS GTDFTLTIS SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQ S GNSQES VTEQD SKD ST
YSL SSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
180 ABP15
Light Chain EIVLTQSPATL SLSPGERATL SCRASQSVS SYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
182 ABP16
Light Chain EIVLTQSPATL SLSPGERATL SCRASQSVS SYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
184 ABP17
Light Chain EIVLTQSPATL SLSPGERATL SCRASQSVS SYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
186 ABP18
Light Chain EIVLTQSPATL SLSPGERATL SCRASQSVS SYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
107 ABP19
Light Chain DIVMTQ SPL SLPVTPGEPAS IS CRS SQSLLH SNGYNYLDWYLQ
Full KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKI SRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
109 ABP20
Light Chain DIQLTQ SP S SVSASVGDRVTITCRASQGIS SWLAWYQQKPGK
Full
APKLLIYGAS SLQ S GVP SRF S GS GS GTDF TLTIS SLQPEDFATY
YCQQASLFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTA
SVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD S
TY SL S STLTL SKADYEKHKVYACEVTHQGL S SPVTKSFNRGE
C
111 ABP21
Light Chain EIVLTQSPATL SLSPGERATL SCRASQSVS SYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
113 ABP22
Light Chain EIVLTQSPATL SLSPGERATL SCRASQSVS SYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
114
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP23
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP24
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
107 ABP25
Light Chain DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLDWYLQ
Full KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTK
SFNRGEC
109 ABP26
Light Chain DIQLTQSPSSVSASVGDRVTITCRASQGISSWLAWYQQKPGK
Full
APKLLIYGASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATY
YCQQASLFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTA
SVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDS
TYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGE
C
111 ABP27
Light Chain EIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113 ABP28
Light Chain EIVLTQSPATLSLSPGERATLSCRASQSVSSYLAWYQQKPGQA
Full PRLLIYDASNRATGIPARFSGSGSGTDFTLTISSLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP29
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
115
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
8 ABP30
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP31
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP32
Light Chain DIQMTQSPSSLSASVGDRVTITCRASQSISSYLNWYQQKPGKA
Full PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQDSKDST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
125 ABP33
Light Chain DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLDWYLQ
Full KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQALQTPITFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTK
SFNRGEC
125 ABP34
Light Chain DIVMTQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLDWYLQ
Full KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQALQTPITFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QDSKDSTYSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTK
SFNRGEC
9 ABP1 VH QVQLQESGPGLVKPSETLSLTCAVSGYSISSGLGWGWIRQPPG
KGLEWIGGIYESGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
AADTAVYYCAHERVRGYGDYGGHHAFDIWGQGTMVTVSS
19 ABP2 VH QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVVVSWIRQHP
GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSS
VTAADTAVYYCADENVRGYGDYGGHHAFDIWGQGTMVTVS
S
26 ABP3 VH QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSS
34 ABP4 VH QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
KGPEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSS
116
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
26 ABP 5 VH QVQLQESGPGLVKPSETLSLTCAVSGYSIS SGAGWGWIRQPPG
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKL S S VT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS S
26 ABP6 VH QVQLQESGPGLVKPSETLSLTCAVSGYSIS SGAGWGWIRQPPG
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKL S S VT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS S
44 ABP7 VH QVQLQESGPGLVKPSETLSLTCAVSGYSIS SEYMWGWIRQPP
GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
S
44 ABP 8 VH QVQLQESGPGLVKPSETLSLTCAVSGYSIS SEYMWGWIRQPP
GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
S
58 ABP 9 VH QLQLQE S GP GL VKP SETL SLT CTVS GGSIS S S SYAWGWIRQPP
GKGLEWIGSIYYSGSTYYNP SLKSRVTISVDTSKNQFSLKLS SV
TAADTAVYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS S
58 ABP10 VH QLQLQE S GP GL VKP SETL SLT CTVS GGSIS S S SYAWGWIRQPP
GKGLEWIGSIYYSGSTYYNP SLKSRVTISVDTSKNQFSLKLS SV
TAADTAVYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS S
62 ABP11 VH QVQLVQSGAEVKRPGS SVKVS CKASGGTFS SYYISWVRQVPG
QGLEWMGGIIPVPGTANYAQKFQGRVTITADEST STAYNIEL S
SLRSEDTAVYYCARAGGGYARGDHYYGM DVWGQGTTVTVS
S
70 ABP12 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYYISWVRQAPG
QRLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYNIELS
SLRSEDTAVYYCARAGGGYARGDHYYGM DVWGQGTTVTVS
S
71 ABP13 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFSRGAISWVRQAP
GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTGTAYNIEL
S SLR SED TAVYYCARQ SDYGLPRGMD VWGQ GTTVTVS S
79 ABP14 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPG
QGLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYNIEL S S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS S
87 ABP15 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPG
QRLEWMGGIIPI S GFTNYAQKFQGRVTITADES TS TAYNIEL S S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS S
88 ABP16 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPG
QGLEWMGGIIPISGFTNYAQKFQGKVTITADESTSTAYNIEL S S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS S
117
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
89 ABP17 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFVRYAISWVRQAP
GQGLEWMGGIIPIFGEAQYAQRFQGRVTITADESTSTAYIVIELS
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVS S
95 ABP18 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFVRYAISWVRQAP
GQGLEWNIGGIIPIF GEAQYAQKFRGRATITADE ST STAYIVIEL S
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVS S
97 ABP19 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYYISWVRQAPG
QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYIVIELS
SLRSEDTAVYYCARAGGGYARGDHYYGM DVWGQGTTVTVS
S
98 ABP20 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFSRGAISWVRQAP
GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADE STSTAYIVIEL
S SLR SED TAVYYCARQ SDYGLPRGMD VWGQ GTTVTVS S
99 ABP21 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPG
QGLEWMGGIIPISGFANYAQKFQGRVTITADESTSTAYIVIELS S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS S
101 ABP22 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFVRYAISWVRQAP
GQGLEWMGGIIPIFGEAQYAQKFQGRVTITADESTSTAYIVIEL S
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVS S
104 ABP23 VH QVQLQESGPGLVKPSETLSLTCAVSGYSIS SEYMWGWIRQPP
GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS
S
105 ABP24 VH QVQLQESGPGLVKPSQTLSLTCTVSGGSIS SGGAVWSWIRQHP
GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCARD SVRGYGDYGGHHAFDIWGQGTMVTVS
S
97 ABP25 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYYISWVRQAPG
QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYIVIELS
SLRSEDTAVYYCARAGGGYARGDHYYGM DVWGQGTTVTVS
S
98 ABP26 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFSRGAISWVRQAP
GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADE STSTAYIVIEL
S SLR SED TAVYYCARQ SDYGLPRGMD VWGQ GTTVTVS S
99 ABP27 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPG
QGLEWMGGIIPISGFANYAQKFQGRVTITADESTSTAYIVIELS S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS S
101 ABP28 VH QVQLVQSGAEVKKPGS SVKVSCKASGGTFVRYAISWVRQAP
GQGLEWMGGIIPIFGEAQYAQKFQGRVTITADESTSTAYIVIEL S
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVS S
118
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
104 ABP29 VH QVQLQESGPGLVKPSETLSLTCAVSGYSIS SEYMWGWIRQPP
GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS
105 ABP30 VH QVQLQESGPGLVKPSQTLSLTCTVSGGSIS SGGAVWSWIRQHP
GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCARD SVRGYGDYGGHHAFDIWGQGTMVTVS
104 ABP31 VH QVQLQESGPGLVKPSETLSLTCAVSGYSIS SEYMWGWIRQPP
GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCARD SGRGYGDYGGHHAFDIWGQGTMVTVS
105 ABP32 VH QVQLQESGPGLVKPSQTLSLTCTVSGGSIS SGGAVWSWIRQHP
GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCARD SVRGYGDYGGHHAFDIWGQGTMVTVS
126 ABP33 VH Variable 1 QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYYMHWVRQA
PGQGLEWMGIINPSGGSTTYAQKFQGRVTMTRDTSTSTVYME
LS SLRSEDTAVYYCARGQYGYYGGRLDVWGQGTTVTVS S
126 ABP34 VH Variable 1 QVQLVQSGAEVKKPGASVKVSCKASGYTFTSYYMHWVRQA
PGQGLEWMGIINPSGGSTTYAQKFQGRVTMTRDTSTSTVYME
LS SLRSEDTAVYYCARGQYGYYGGRLDVWGQGTTVTVS S
127 ABP33 VH Variable 2 QVQLVESGGGVVQPGRSLRL S CAA S GFTF S SYGMHWVRQAP
GKGLEWVAVISYDGSNKYYAD SVKGRFTISRDNSKNTLYLQ
MNSLRAEDTAVYYCARDGLGYMAWGQGTTVTVS S
127 ABP34 VH Variable 2 QVQLVESGGGVVQPGRSLRL SCAASGFTFS SYGMHWVRQAP
GKGLEWVAVISYDGSNKYYAD SVKGRFTISRDNSKNTLYLQ
MNSLRAEDTAVYYCARDGLGYMAWGQGTTVTVS S
ABP 1 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
ABP2 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIK
27 ABP3 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLKYGVP SRF S GS GS GTDFTL TIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIK
35 ABP4 VL DIQMTQ SP S SL S A S VGDRVTIT CRA SK S ID
SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIK
119
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35 ABP 5 VL DIQMTQ SP S SL S A S VGDRVTIT CRA SK S ID
SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIK
40 ABP6 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAD SLQSGVP SRF S GS GS GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIK
45 ABP7 VL DIQMTQ SP S SL SAS VGDRVTITCRASQ S INSYLNWYQQKPGKA
PKLLIYAAS SLD S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIK
53 ABP 8 VL DIQMTQ SP S SL SASVGDRVTITCGASQSISTYLNWYQQKPGKA
PKLLIYSAS SLE S GVP SRF S G S GS GTDFTL TIS SLQPEDFATYYC
QQEYNTPP SF GGGTKVEIK
ABP 9 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
10 ABP10 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
63 ABP11 VL DIVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKI SRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIK
63 ABP12 VL DIVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKI SRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIK
72 ABP13 VL DIQLTQ SP S SVSASVGDRVTITCRASQGIS SWLAWYQQKPGK
APKLLIYGAS SLQ S GVP SRF S GS G S GTDF TLTIS SLQPEDFATY
YCQQASLFPPTFGGGTKVEIK
80 ABP14 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIK
80 ABP15 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIK
80 ABP16 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIK
90 ABP17 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIK
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90 ABP18 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIK
63 ABP19 VL DIVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKI SRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIK
72 ABP20 VL DIQLTQ SP S SVSASVGDRVTITCRASQGIS SWLAWYQQKPGK
APKLLIYGAS SLQ S GVP SRF S GS G S GTDF TLTIS SLQPEDFATY
YCQQASLFPPTFGGGTKVEIK
80 ABP21 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIK
90 ABP22 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIK
ABP23 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
10 ABP24 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
63 ABP25 VL DIVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKI SRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIK
72 ABP26 VL DIQLTQ SP S SVSASVGDRVTITCRASQGIS SWLAWYQQKPGK
APKLLIYGAS SLQ S GVP SRF S GS G S GTDF TLTIS SLQPEDFATY
YCQQASLFPPTFGGGTKVEIK
80 ABP27 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIK
90 ABP28 VL EIVLTQSPATL SL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIK
10 ABP29 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
10 ABP30 VL DIQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
121
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ABP31 VL DIQMTQ SP S SL SASVGDRVTITCRASQSISSYLNWYQQKPGKA
PKLLIYAAS SLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
10 ABP32 VL DIQMTQ SP S SL SASVGDRVTITCRASQSISSYLNWYQQKPGKA
PKLLIYAAS SLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIK
128 ABP33 VL DIVNITQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLDWYLQ
KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKI SRVEAE
DVGVYYCMQALQTPITFGGGTKVEIK
128 ABP34 VL DIVNITQSPLSLPVTPGEPASISCRSSQSLLHSNGYNYLDWYLQ
KPGQ SPQLLIYLGSNRAS GVPDRF S GS GS GTDFTLKI SRVEAE
DVGVYYCMQALQTPITFGGGTKVEIK
11 ABP 1 VH CDR3 AHERVRGYGDYGGHHAFDI
21 ABP2 VH CDR3 ADENVRGYGDYGGHHAFD I
28 ABP3 VH CDR3 VLESVRGYGDYGGHHAFDI
28 ABP4 VH CDR3 VLESVRGYGDYGGHHAFDI
28 ABP5 VH CDR3 VLESVRGYGDYGGHHAFDI
28 ABP6 VH CDR3 VLESVRGYGDYGGHHAFDI
46 ABP7 VH CDR3 AREADRGYGDYGGHHAFDI
46 ABP 8 VH CDR3 AREADRGYGDYGGHHAFDI
59 ABP9 VH CDR3 ARDSGRGYGDYGGHHAFDI
59 ABP10 VH CDR3 ARDSGRGYGDYGGHHAFDI
64 ABP11 VH CDR3 ARAGGGYARGDHYYGMD V
64 ABP12 VH CDR3 ARAGGGYARGDHYYGMD V
73 ABP13 VH CDR3 ARQSDYGLPRGMDV
81 ABP14 VH CDR3 AREGGHYYSGWPY
81 ABP15 VH CDR3 AREGGHYYSGWPY
81 ABP16 VH CDR3 AREGGHYYSGWPY
91 ABP17 VH CDR3 AREGYYYGALPY
91 ABP18 VH CDR3 AREGYYYGALPY
64 ABP19 VH CDR3 ARAGGGYARGDHYYGMD V
73 ABP20 VH CDR3 ARQSDYGLPRGMDV
81 ABP21 VH CDR3 AREGGHYYSGWPY
91 ABP22 VH CDR3 AREGYYYGALPY
46 ABP23 VH CDR3 ARDSGRGYGDYGGHHAFDI
21 ABP24 VH CDR3 ARDSVRGYGDYGGHHAFDI
64 ABP25 VH CDR3 ARAGGGYARGDHYYGMD V
73 ABP26 VH CDR3 ARQSDYGLPRGMDV
81 ABP27 VH CDR3 AREGGHYYSGWPY
122
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91 ABP28 VH CDR3 AREGYYYGALPY
46 ABP29 VH CDR3 ARD SGRGYGDYGGHHAFDI
21 ABP30 VH CDR3 ARD SVRGYGDYGGHHAFDI
46 ABP31 VH CDR3 ARD SGRGYGDYGGHHAFDI
21 ABP32 VH CDR3 ARD SVRGYGDYGGHHAFDI
131 ABP33 VH CDR3 vi ARGQYGYYGGRLDV
131 ABP34 VH CDR3 vi ARGQYGYYGGRLDV
134 ABP33 VH CDR3 v2 ARDGLGYMA
134 ABP34 VH CDR3 v2 ARDGLGYMA
12 ABP 1 VH CDR2 GIYESGSTYYNP SLKS
22 ABP2 VH CDR2 GIAYS GS TYYNP SLKS
29 ABP3 VH CDR2 LIVHSGSTYYNPSLKS
29 ABP4 VH CDR2 LIVHSGSTYYNPSLKS
29 ABP5 VH CDR2 LIVHSGSTYYNPSLKS
29 ABP6 VH CDR2 LIVHSGSTYYNPSLKS
47 ABP7 VH CDR2 LIYHSGKTYYNPSLKS
47 ABP 8 VH CDR2 LIYHSGKTYYNPSLKS
60 ABP 9 VH CDR2 SIYYSGSTYYNPSLKS
60 ABP10 VH CDR2 SIYYSGSTYYNPSLKS
65 ABP11 VH CDR2 GIIPVPGTANYAQKFQG
65 ABP12 VH CDR2 GIIPVPGTANYAQKFQG
74 ABP13 VH CDR2 GIIPIEGTAYYAQKFQG
82 ABP14 VH CDR2 GIIPISGFTNYAQKFQG
82 ABP15 VH CDR2 GIIPISGFTNYAQKFQG
82 ABP16 VH CDR2 GIIPISGFTNYAQKFQG
92 ABP17 VH CDR2 GIIPIFGEAQYAQRFQG
96 ABP18 VH CDR2 GIIPIFGEAQYAQKFRG
65 ABP19 VH CDR2 GIIPVPGTANYAQKFQG
74 ABP20 VH CDR2 GIIPIEGTAYYAQKFQG
100 ABP21 VH CDR2 GIIPISGFANYAQKFQG
102 ABP22 VH CDR2 GIIPIFGEAQYAQKFQG
47 ABP23 VH CDR2 LIYHSGKTYYNPSLKS
22 ABP24 VH CDR2 GIAYS GS TYYNP SLKS
65 ABP25 VH CDR2 GIIPVPGTANYAQKFQG
74 ABP26 VH CDR2 GIIPIEGTAYYAQKFQG
100 ABP27 VH CDR2 GIIPISGFANYAQKFQG
102 ABP28 VH CDR2 GIIPIFGEAQYAQKFQG
47 ABP29 VH CDR2 LIYHSGKTYYNPSLKS
22 ABP30 VH CDR2 GIAYS GS TYYNP SLKS
123
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47 ABP31 VH CDR2 LIYHSGKTYYNPSLKS
22 ABP32 VH CDR2 GIAYS GS TYYNP SLKS
130 ABP33 VH CDR2 vi IINPSGGSTTYAQKFQG
130 ABP34 VH CDR2 vi IINPSGGSTTYAQKFQG
133 ABP33 VH CDR2 v2 VI SYD G SNKYYAD SVKG
133 ABP34 VH CDR2 v2 VI SYD GSNKYYAD SVKG
11 ABP 1 VH CDR1 YSISSGLGWG
21 ABP2 VH CDR1 GSISSGGAVWS
28 ABP3 VH CDR1 YSISSGAGWG
28 ABP4 VH CDR1 YSISSGAGWG
28 ABP5 VH CDR1 YSISSGAGWG
28 ABP6 VH CDR1 YSISSGAGWG
46 ABP7 VH CDR1 YSISSEYNIWG
46 ABP 8 VH CDR1 YSISSEYNIWG
59 ABP9 VH CD R 1 GSISSS SYAWG
59 ABP10 VH CD R 1 GSISSS SYAWG
64 ABP11 VH CDR1 GTFSSYYIS
64 ABP12 VH CDR1 GTFSSYYIS
73 ABP13 VH CDR1 GTFSRGAIS
81 ABP14 VH CDR1 GTFSSYAIS
81 ABP15 VH CDR1 GTFSSYAIS
81 ABP16 VH CDR1 GTFSSYAIS
91 ABP17 VH CDR1 GTFVRYAIS
91 ABP18 VH CDR1 GTFVRYAIS
64 ABP19 VH CDR1 GTFSSYYIS
73 ABP20 VH CDR1 GTFSRGAIS
81 ABP21 VH CDR1 GTFSSYAIS
91 ABP22 VH CDR1 GTFVRYAIS
46 ABP23 VH CDR1 YSISSEYNIWG
21 ABP24 VH CDR1 GSISSGGAVWS
64 ABP25 VH CDR1 GTFSSYYIS
73 ABP26 VH CDR1 GTFSRGAIS
81 ABP27 VH CDR1 GTFSSYAIS
91 ABP28 VH CDR1 GTFVRYAIS
46 ABP29 VH CDR1 YSISSEYNIWG
21 ABP30 VH CDR1 GSISSGGAVWS
46 ABP31 VH CDR1 YSISSEYNIWG
21 ABP32 VH CDR1 GSISSGGAVWS
129 ABP33 VH CDR1 vi YTFTSYYMH
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129 ABP34 VH CDR1 vi YTFTSYYMH
132 ABP33 VH CDR1 v2 FTF S SYGMH
132 ABP34 VH CDR1 v2 FTF S SYGMH
16 ABP 1 VI, CDR3 QQEYATPPT
17 ABP2 VI, CDR3 QQEYNTPPT
17 ABP3 VI, CDR3 QQEYNTPPT
17 ABP4 VI, CDR3 QQEYNTPPT
17 ABP5 VI, CDR3 QQEYNTPPT
17 ABP6 VI, CDR3 QQEYNTPPT
17 ABP7 VI, CDR3 QQEYNTPPT
56 ABP 8 VI, CDR3 QQEYNTPPS
16 ABP9 VI, CDR3 QQEYATPPT
16 ABP10 VI, CDR3 QQEYATPPT
69 ABP11 VI, CDR3 MQKIGTPLT
69 ABP12 VI, CDR3 MQKIGTPLT
78 ABP13 VI, CDR3 QQASLFPPT
86 ABP14 VI, CDR3 QQRLVFPPT
86 ABP15 VI, CDR3 QQRLVFPPT
86 ABP16 VI, CDR3 QQRLVFPPT
94 ABP17 VI, CDR3 QQHSVFPPT
94 ABP18 VI, CDR3 QQHSVFPPT
69 ABP19 VI, CDR3 MQKIGTPLT
78 ABP20 VI, CDR3 QQASLFPPT
86 ABP21 VI, CDR3 QQRLVFPPT
94 ABP22 VI, CDR3 QQHSVFPPT
16 ABP23 VI, CDR3 QQEYATPPT
16 ABP24 VI, CDR3 QQEYATPPT
69 ABP25 VI, CDR3 MQKIGTPLT
78 ABP26 VI, CDR3 QQASLFPPT
86 ABP27 VI, CDR3 QQRLVFPPT
94 ABP28 VI, CDR3 QQHSVFPPT
16 ABP29 VI, CDR3 QQEYATPPT
16 ABP30 VI, CDR3 QQEYATPPT
16 ABP31 VI, CDR3 QQEYATPPT
16 ABP32 VI, CDR3 QQEYATPPT
135 ABP33 VI, CDR3 MQALQTPIT
135 ABP34 VI, CDR3 MQALQTPIT
15 ABP 1 VI, CDR2 AASSLQS
15 ABP2 VI, CDR2 AASSLQS
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31 ABP3 VI, CDR2 AASSLKY
15 ABP4 VI, CDR2 AASSLQS
15 ABP5 VI, CDR2 AASSLQS
41 ABP6 VI, CDR2 AADSLQS
50 ABP7 VI, CDR2 AASSLDS
55 ABP8 VI, CDR2 SASSLES
15 ABP9 VI, CDR2 AASSLQS
15 ABP10 VI, CDR2 AASSLQS
68 ABP11 VI, CDR2 LGSNRAS
68 ABP12 VI, CDR2 LGSNRAS
77 ABP13 VI, CDR2 GASSLQS
85 ABP14 VI, CDR2 DASNRAT
85 ABP15 VI, CDR2 DASNRAT
85 ABP16 VI, CDR2 DASNRAT
85 ABP17 VI, CDR2 DASNRAT
85 ABP18 VI, CDR2 DASNRAT
68 ABP19 VI, CDR2 LGSNRAS
77 ABP20 VI, CDR2 GASSLQS
85 ABP21 VI, CDR2 DASNRAT
85 ABP22 VI, CDR2 DASNRAT
15 ABP23 VI, CDR2 AASSLQS
15 ABP24 VI, CDR2 AASSLQS
68 ABP25 VI, CDR2 LGSNRAS
77 ABP26 VI, CDR2 GASSLQS
85 ABP27 VI, CDR2 DASNRAT
85 ABP28 VI, CDR2 DASNRAT
15 ABP29 VI, CDR2 AASSLQS
15 ABP30 VI, CDR2 AASSLQS
15 ABP31 VI, CDR2 AASSLQS
15 ABP32 VI, CDR2 AASSLQS
68 ABP33 VI, CDR2 LGSNRAS
68 ABP34 VI, CDR2 LGSNRAS
14 ABP1 VI, CDR1 RASQSISSYLN
14 ABP2 VI, CDR1 RASQSISSYLN
14 ABP3 VI, CDR1 RASQSISSYLN
36 ABP4 VI, CDR1 RASKSIDSYLN
36 ABP5 VI, CDR1 RASKSIDSYLN
14 ABP6 VI, CDR1 RASQSISSYLN
49 ABP7 VI, CDR1 RASQSINSYLN
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54 ABP8 VL CDR1 GASQSISTYLN
14 ABP9 VL CDR1 RASQSISSYLN
14 ABP10 VL CDR1 RASQSISSYLN
67 ABP11 VL CDR1 RSSQSLLHSNGYNYLD
67 ABP12 VL CDR1 RSSQSLLHSNGYNYLD
76 ABP13 VL CDR1 RASQGISSWLA
84 ABP14 VL CDR1 RASQSVSSYLA
84 ABP15 VL CDR1 RASQSVSSYLA
84 ABP16 VL CDR1 RASQSVSSYLA
84 ABP17 VL CDR1 RASQSVSSYLA
84 ABP18 VL CDR1 RASQSVSSYLA
67 ABP19 VL CDR1 RSSQSLLHSNGYNYLD
76 ABP20 VL CDR1 RASQGISSWLA
84 ABP21 VL CDR1 RASQSVSSYLA
84 ABP22 VL CDR1 RASQSVSSYLA
14 ABP23 VL CDR1 RASQSISSYLN
14 ABP24 VL CDR1 RASQSISSYLN
67 ABP25 VL CDR1 RSSQSLLHSNGYNYLD
76 ABP26 VL CDR1 RASQGISSWLA
84 ABP27 VL CDR1 RASQSVSSYLA
84 ABP28 VL CDR1 RASQSVSSYLA
14 ABP29 VL CDR1 RASQSISSYLN
14 ABP30 VL CDR1 RASQSISSYLN
14 ABP31 VL CDR1 RASQSISSYLN
14 ABP32 VL CDR1 RASQSISSYLN
67 ABP33 VL CDR1 RSSQSLLHSNGYNYLD
67 ABP34 VL CDR1 RSSQSLLHSNGYNYLD
137 IgG1 Constant ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNS
CH1 Region GALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN
HKPSNTKVDKRVEPKSC
138 IgG4 Constant ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNS
S228P Region GALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVD
HKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKD
TLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTK
PREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSI
EKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPS
DIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSLSLSLGK
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139 IgG4 Constant ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNS
S228P no Region GALT S GVHTFPAVLQ S SGLYSLS SVVTVPS S SLGTKTYTCNVD
C HKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKD
terminal TLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTK
Lys PREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SI
EKTISKAKGQPREPQVYTLPP SQEEMTKNQVSLTCLVKGFYPS
DIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSL SLSLG
140 Kappa Constant RTVAAPSVFIFPPSDEQLKSGTASVVCLLNNFYPREAKVQWK
Region VDNALQSGNSQESVTEQD SKD STYSLS STLTLSKADYEKHKV
YACEVTHQGLS SPVTKSFNRGEC
141 CDRH3 Generic X1X2X3X4X5RGYGDYGGHHAFDI, WHEREIN X1 IS A OR V, X2
Sequence 1 IS H, D, L, OR R, X3 IS E OR D, X4 IS R, N, S, OR A, AND X5 IS
V, D OR G
142 CDRH2 Generic X1lX2X3SGX4TYYNPSLKS, WHEREIN X1 IS G, L, OR S, X2 IS
Y,
Sequence 1 A, OR V, X3 IS E, Y OR H, AND X4I5 S OR K
143 CDRH1 Generic X1SISSX2X3X4X5WX6, WHEREIN X1 IS Y OR G, X2 IS G, S,
OR
Sequence 1 E, X3IS L, G, S, Y, OR A, X4I5 G, A, Y, M, OR G, X5 IS V, A, OR
IS ABSENT, AND X6 IS S OR G
144 CDRL 3 Generic QQEYX1TPPX2, WHEREIN X1 IS A OR N AND X2 IS T OR S
Sequence 1
145 CDRL2 Generic X1AX2SLX3X4, WHEREIN X1 IS A OR S, X2 IS D OR S, X3 IS
Q,
Sequence 1 D, K, ORE, AND X4 IS S OR Y
146 CDRL1 Generic XiAS X25I X3X4YLN, WHEREIN X1 IS G OR R, X2 IS Q OR K,
Sequence 1 X3 IS S, D, OR N, AND X4 IS S OR T
147 CDRH2 Generic GIIPIFGEAQYAQX1FX2G, WHEREIN X1 IS K OR R, AND X2 IS
Sequence 2 Q OR R
148 ABP35 Full Heavy, QVQLQES GP GLVKP SETL SL TCAVS GYSIS
SGLGWGWIRQPPG
IgG1 N297A KGLEWIGGIYESGSTYYNPSLKSRVTISVDTSKNQFSLKLS SVT
AADTAVYYCAHERVRGYGDYGGHHAFDIWGQGTMVTVS SA
STKGP SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
ALT S GVHTFPAVLQ S SGLYSLS SVVTVP S S SLGTQTYICNVNH
KPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGF
YPSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKS
RWQQGNVFSCSVMHEALHNHYTQKSLSL SPGK
149 ABP36 Full Heavy,
QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVWSWIRQHP
IgG1 N297A GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLS S
128
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VTAADTAVYYCADENVRGYGDYGGHHAFDIWGQGTMVTVS
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
S GAL TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SL GTQTYICNV
NHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLV
KGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSL SLSPGK
150 ABP37 Full Heavy, QVQLQES
GP GLVKP SETL SL TCAVS GYSI S SGAGWGWIRQPPG
IgG1 N297A KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKL S SVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA
STKGP SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
ALT S GVHTFPAVLQ S SGLYSLS SVVTVP S S SL GTQTYICNVNH
KPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGF
YPSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKS
RWQQGNVFSCSVMHEALHNHYTQKSLSL SPGK
151 ABP38 Full Heavy, QVQLQES
GP GLVKP SETL SL TCAVS GYSI S SGAGWGWIRQPPG
IgG1 N297A KGPEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLS SVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA
STKGP SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
ALT S GVHTFPAVLQ S SGLYSLS SVVTVP S S SL GTQTYICNVNH
KPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGF
YPSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKS
RWQQGNVFSCSVMHEALHNHYTQKSLSL SPGK
152 ABP39 Full Heavy, QVQLQES
GP GLVKP SETL SL TCAVS GYSI S SGAGWGWIRQPPG
IgG1 N297A KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKL S SVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA
STKGP SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
ALT S GVHTFPAVLQ S SGLYSLS SVVTVP S S SL GTQTYICNVNH
KPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGF
129
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YPSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKS
RWQQGNVFSCSVMHEALHNHYTQKSL SL SPGK
153 AB P40 Full Heavy, QVQLQE S GP GLVKP S ETL SL TCAVS GY S I S
SEYMVVGWIRQPP
IgG1 N297A GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKL S S
VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
S GAL TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SL GTQTYICNV
NHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLV
KGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSL SL SPGK
154 ABP41 Full Heavy, QVQLQESGPGLVKPSETLSLTCAVSGYSIS SEYMVVGWIRQPP
IgG1 N297A GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKL S S
VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
S GAL TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SL GTQTYICNV
NHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLV
KGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSL SL SPGK
155 AB P42 Full Heavy, QLQLQE S GP GL VKP SETL SLTCTVSGGSIS S S
SYAWGWIRQPP
IgG1 N297A GKGLEWIGSIYYSGSTYYNP SLKSRVTISVDTSKNQFSLKL S SV
TAAD TAVYY CARD SGRGYGDYGGHHAFDIWGQGTMVTVS S
ASTKGP SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNS
GALT S GVHTFPAVLQ S SGLYSL S SVVTVPS S SL GTQTYICNVN
HKPSNTKVDKKVEPKS CDKTHTCPPCPAPELL GGPSVFLFPPK
PKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNA
KTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKAL
PAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKG
FYPSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKLTVDK
SRWQQGNVFSCSVMHEALHNHYTQKSL SL SPGK
156 ABP43 Full Heavy, QLQLQESGPGLVKPSETLSLTCTVSGGSISSSSYAWGWIRQPP
IgG4 S228P GKGLEWIGSIYYSGSTYYNP SLKSRVTISVDTSKNQFSLKL S SV
TAAD TAVYY CARD SGRGYGDYGGHHAFDIWGQGTMVTVS S
ASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNS
GALT S GVHTFPAVLQ S SGLYSL S SVVTVPS S SL GTKTYTCNVD
130
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HKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKD
TLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTK
PREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SI
EKTISKAKGQPREPQVYTLPP SQEEMTKNQVSLTCLVKGFYPS
DIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSL SL SL GK
157 ABP44 Full Heavy, QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYYISWVRQAPG
IgG1 N297A QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
SLRSEDTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVS
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
S GAL TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SL GTQTYICNV
NHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLV
KGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSL SL SPGK
158 AB P45 Full Heavy, QVQLVQSGAEVKKPGS SVKVSCKASGGTFSRGAISWVRQAP
IgG1 N297A GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTSTAYMEL
S SLR SED TAVYYCARQ SDYGLPRGMD VVVGQ GTTVTVS SAS T
KGPSVFPLAPS SKSTSGGTAAL GCLVKDYFPEPVTVSWNS GA
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
PSNTKVDKKVEPKSCDKTHTCPPCPAPELL GGPSVFLFPPKPK
DTLMISRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKT
KPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPA
PIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFY
PSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKLTVDKSR
WQQGNVFSCSVMHEALHNHYTQKSL SL SPGK
159 ABP46 Full Heavy, QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
IgG1 N297A QGLEWMGGIIPISGFANYAQKFQGRVTITADESTSTAYMEL S S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS SA STKGP
S VFPL AP S SK S T S GGTAAL GCLVKDYFPEP VTVS WN S GALT S G
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
I SRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPRE
EQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPP SRDELTKNQVSL TCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFS CSVMHEALHNHYTQKSL SL SP GK
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160 ABP47 Full Heavy,
QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
IgG1 N297A GQGLEWMGGIIPIFGEAQYAQKFQGRVTITADESTSTAYMELS
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
ISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPRE
EQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGFYPSDI
AVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQ
QGNVFSCSVMHEALHNHYTQKSLSLSPGK
161 ABP48 Full Heavy, QVQLQESGPGLVKPSETLSLTCAVSGYSISSEYMVVGWIRQPP
IgG1 N297A GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
VTAADTAVYYCARDSGRGYGDYGGHHAFDIWGQGTMVTVS
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
NHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLV
KGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK
162 ABP49 Full Heavy,
QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVVVSWIRQHP
IgG1 N297A GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSS
VTAADTAVYYCARDSVRGYGDYGGHHAFDIWGQGTMVTVS
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
NHKPSNTKVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLV
KGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGK
163 ABP50
Full Heavy, N QVQLVQSGAEVKRPGSSVKVSCKASGGTFSSYYISWVRQVPG
terminal Fab QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
TM (IgG1
SLRSEDTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVS
Fab, IgG4
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
5228P)
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
NHKPSNTKVDKRVEPKSCGGGGSQVQLVQSGAEVKRPGSSV
KVSCKASGGTFSSYYISWVRQVPGQGLEWMGGIIPVPGTANY
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AQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARAGGG
YARGDHYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRST
SESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSS
GLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKY
GPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV
SQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVY
TLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALH
NHYTQKSLSLSLGK
164 ABP51
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYYISWVRQAPG
terminal Fab QRLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
TM (IgG1
SLRSEDTAVYYCARAGGGYARGDHYYGMDVWGQGTTVTVS
Fab, IgG4
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
S228P)
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
NHKPSNTKVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSV
KVSCKASGGTFSSYYISWVRQAPGQRLEWMGGIIPVPGTANY
AQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARAGGG
YARGDHYYGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRST
SESTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSS
GLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKY
GPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV
SQEDPEVQFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVY
TLPPSQEEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLDSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALH
NHYTQKSLSLSLGK
165 ABP52 Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSRGAISWVRQAP
terminal Fab GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTGTAYMEL
TM (IgG1
SSLRSEDTAVYYCARQSDYGLPRGMDVWGQGTTVTVSSAST
Fab, IgG4
KGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGA
5228P)
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
PSNTKVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVS
CKASGGTFSRGAISWVRQAPGQGLEWMGGIIPIEGTAYYAQK
FQGRVTITADESTGTAYMELSSLRSEDTAVYYCARQSDYGLP
RGMDVWGQGTTVTVSSASTKGPSVFPLAPCSRSTSESTAALG
CLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV
VTVPSSSLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCP
APEFLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEV
QFNWYVDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDW
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LNGKEYKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQE
EMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVL
DSDGSFFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQK
SLSLSLGK
166 ABP53
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
terminal Fab QGLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYMELSS
TM (IgG1
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
Fab, IgG4
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
S228P)
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFSSYAISWVRQAPGQGLEWMGGIIPISGFTNYAQKFQGR
VTITADESTSTAYMELSSLRSEDTAVYYCAREGGHYYSGWPY
WGQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKD
YFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSS
SLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLG
GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWY
VDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKE
YKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKN
QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGS
FFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSL
GK
167 ABP54
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
terminal Fab QRLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYMELSS
TM (IgG1
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
Fab, IgG4
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
S228P)
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFSSYAISWVRQAPGQRLEWMGGIIPISGFTNYAQKFQGR
VTITADESTSTAYMELSSLRSEDTAVYYCAREGGHYYSGWPY
WGQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKD
YFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSS
SLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLG
GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWY
VDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKE
YKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKN
QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGS
FFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSL
GK
168 ABP55 Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
terminal Fab QGLEWMGGIIPISGFTNYAQKFQGKVTITADESTSTAYMELSS
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TM (IgG1
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
Fab, IgG4
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
S228P)
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFSSYAISWVRQAPGQGLEWMGGIIPISGFTNYAQKFQGK
VTITADESTSTAYMELSSLRSEDTAVYYCAREGGHYYSGWPY
WGQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKD
YFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSS
SLGTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLG
GPSVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWY
VDGVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKE
YKCKVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKN
QVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGS
FFLYSRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSL
GK
169 ABP56
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
terminal Fab GQGLEWMGGIIPIFGEAQYAQRFQGRVTITADESTSTAYMELS
TM (IgG1
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
Fab, IgG4
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
S228P)
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFVRYAISWVRQAPGQGLEWMGGIIPIFGEAQYAQRFQGR
VTITADESTSTAYMELSSLRSEDTAVYYCAREGYYYGALPYW
GQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYF
PEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSL
GTKTYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGP
SVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD
GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKC
KVSNKGLPSSIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVS
LTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLY
SRLTVDKSRWQEGNVFSCSVMHEALHNHYTQKSLSLSLGK
170 ABP57
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
terminal Fab GQGLEWMGGIIPIFGEAQYAQKFRGRATITADESTSTAYMELS
TM (IgG1
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
Fab, IgG4
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
S228P)
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFVRYAISWVRQAPGQGLEWMGGIIPIFGEAQYAQKFRGR
ATITADESTSTAYMELSSLRSEDTAVYYCAREGYYYGALPYW
GQGTLVTVSSASTKGPSVFPLAPCSRSTSESTAALGCLVKDYF
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PEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSL
GTKTYTCNVDHKP SNTKVDKRVESKYGPPCPPCPAPEFL GGP
SVFLFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVD
GVEVHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKC
KVSNKGLP S SIEKTISKAKGQPREPQVYTLPP SQEEMTKNQVS
LT CLVKGFYP SDIAVEWESNGQPENNYKTTPPVLD SD GSFFLY
SRLTVDKSRWQEGNVF SCSVMHEALHNHYTQK SL SL SLGK
179 AB P58 Full Heavy, QVQLVQ S GAEVKKP GA S VKV S CKA S GYTFT
SYYMHWVRQA
IgG1 N297A PGQGLEWMGIINPSGGSTTYAQKFQGRVTMTRDTSTSTVYME
L S SLRSEDTAVYYCARGQYGYYGGRLDVVVGQGTTVTVS SA S
TKGP SVFPLAP S SKST SGGTAALGCLVKDYFPEPVTVSWNSG
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
KP SNTKVDKKVEPKSCDKTHTCPPCPAPELLGGP S VFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSRDELTKNQVSLTCLVKGF
YP SDIAVEWESNGQPENNYKTTPPVLD SD GSFFLY SKL TVDK S
RWQQGNVF SCSVMHEALHNHYTQKSL SL SP GK
180 AB P59 Full Heavy, QVQLVQ S GAEVKKP GA S VKV S CKA S GYTFT
SYYMHWVRQA
IgG4 S228P PGQGLEWMGIINP SGGSTTYAQKFQ GRVTMTRDT STSTVYME
L S SLRSEDTAVYYCARGQYGYYGGRLDVVVGQGTTVTVS SA S
TKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGA
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHK
P SNTKVDKRVESKYGPPCPPCPAPEFLGGP SVFLFPPKPKDTL
MISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPR
EEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLP S SIEK
TISKAKGQPREPQVYTLPP SQEEMTKNQVSLTCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQE
GNVF SCSVMHEALHNHYTQKSL SL SL GK
181 AB P60 Full Heavy,
QVQLVESGGGVVQPGRSLRL S CAA S GFTF S SYGMHWVRQ AP
IgG1 N297A GKGLEWVAVISYDGSNKYYAD SVKGRFTISRDNSKNTLYLQ
MNSLRAEDTAVYYCARDGLGYMAWGQGTTVTVSSASTKGP
S VFPL AP S SKST SGGTAALGCLVKDYFPEPVTVS WN S GALT S G
VHTFPAVLQ S SGLYSL S SVVTVP S S SLGTQTYICNVNHKP S NT
KVDKKVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
ISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAKTKPRE
EQYASTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPP SRDELTKNQVSLTCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQ
QGNVF S CSVMHEALHNHYTQKSL SL SP GK
136
CA 03042727 2019-05-02
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PCT/US2017/062443
182 ABP61 Full Heavy,
QVQLVESGGGVVQPGRSLRLSCAASGFTFSSYGMHWVRQAP
IgG4 S228P GKGLEWVAVISYDGSNKYYADSVKGRFTISRDNSKNTLYLQ
MNSLRAEDTAVYYCARDGLGYMAWGQGTTVTVSSASTKGP
SVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSG
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKPSNT
KVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISR
TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISK
AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVE
WESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGN
VFSCSVMHEALHNHYTQKSLSLSLGK
183 ABP62
Full Heavy, N QVQLQESGPGLVKPSETLSLTCAVSGYSISSGLGWGWIRQPPG
terminal Fab KGLEWIGGIYESGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM (IgG1
AADTAVYYCAHERVRGYGDYGGHHAFDIWGQGTMVTVSSA
Fab, IgG1
STKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3)))
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
KPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLSLT
CAVSGYSISSGLGWGWIRQPPGKGLEWIGGIYESGSTYYNPSL
KSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAHERVRGYGD
YGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPSSKSTSGGT
AALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS
LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKT
HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV
SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY
TLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALH
NHYTQKSLSLSPGK
184 ABP63
Full Heavy, N QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVWSWIRQHP
terminal Fab GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSS
TM (IgG1
VTAADTAVYYCADENVRGYGDYGGHHAFDIWGQGTMVTVS
Fab, IgG1 SASTKGPSVFPLAPSS
(G1m(3)))
KSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVL
QSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEP
KSCGGGGSQVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAV
WSWIRQHPGKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKN
QFSLKLSSVTAADTAVYYCADENVRGYGDYGGHHAFDIWG
QGTMVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYF
PEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSL
GTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELL
137
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PCT/US2017/062443
GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW
YVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGK
EYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTK
NQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG
SFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLS
PGK
185 ABP64
Full Heavy, N QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
terminal Fab KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM (IgG1
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
Fab, IgG1
STKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3)))
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
KPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLSLT
CAVSGYSISSGAGWGWIRQPPGKGLEWIGLIVHSGSTYYNPSL
KSRVTISVDTSKNQFSLKLSSVTAADTAVYYCVLESVRGYGD
YGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPSSKSTSGGT
AALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS
LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKT
HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV
SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY
TLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALH
NHYTQKSLSLSPGK
186 ABP65
Full Heavy, N QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
terminal Fab KGPEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM (IgG1
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
Fab, IgG1
STKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3)))
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
KPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLSLT
CAVSGYSISSGAGWGWIRQPPGKGPEWIGLIVHSGSTYYNPSL
KSRVTISVDTSKNQFSLKLSSVTAADTAVYYCVLESVRGYGD
YGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPSSKSTSGGT
AALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYS
LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKT
HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV
SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY
TLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALH
NHYTQKSLSLSPGK
138
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187 ABP66
Full Heavy, N QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
terminal Fab KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM (IgG1
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA
Fab, IgG1 STKGP
SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3))) ALT S
GVHTFPAVLQ S SGLYSLS SVVTVP S S SLGTQTYICNVNH
KPSNTKVDKRVEPKS CGGGGSQVQLQESGPGLVKPSETLSLT
CAVSGYSIS SGAGWGWIRQPPGKGLEWIGLIVHSGSTYYNPSL
KSRVTISVDTSKNQFSLKLS SVTAADTAVYYCVLESVRGYGD
YGGHHAFDIWGQGTMVTVS SASTKGPSVFPLAPS SKS TS GGT
AAL G CLVKDYFPEPVTVS WN S GALT S GVHTFPAVLQ S SGLYS
LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKT
HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV
SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY
TLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLD SD G SFFLY SKL TVDK SRWQQ GNVF SCSVMHEALH
NHYTQKSLSL SP GK
188 ABP67
Full Heavy, N QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
terminal Fab KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM (IgG1
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA
Fab, IgG1 STKGP
SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3))) ALT S
GVHTFPAVLQ S SGLYSLS SVVTVP S S SLGTQTYICNVNH
KPSNTKVDKRVEPKS CGGGGSQVQLQESGPGLVKPSETLSLT
CAVSGYSIS SGAGWGWIRQPPGKGLEWIGLIVHSGSTYYNPSL
KSRVTISVDTSKNQFSLKLS SVTAADTAVYYCVLESVRGYGD
YGGHHAFDIWGQGTMVTVS SASTKGPSVFPLAPS SKS TS GGT
AAL G CLVKDYFPEPVTVS WN S GALT S GVHTFPAVLQ S SGLYS
LSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKT
HTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDV
SHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLT
VLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVY
TLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNY
KTTPPVLD SD G SFFLY SKL TVDK SRWQQ GNVF SCSVMHEALH
NHYTQKSLSL SP GK
189 ABP68 Full Heavy, N QVQLQESGPGLVKPSETLSLTCAVSGYSISSEYMVVGWIRQPP
terminal Fab GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
TM (IgG1
VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
Fab, IgG1
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3))) S GAL
TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SLGTQTYICNV
NHKPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLS
139
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LTCAVSGYSISSEYMWGWIRQPPGKGLEWIGLIYHSGKTYYN
PSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREADRG
YGDYGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPSSKSTS
GGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG
LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCD
KTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVV
DVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS
VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREP
QVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPE
NNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMH
EALHNHYTQKSLSLSPGK
190 ABP69 Full Heavy, N QVQLQESGPGLVKPSETLSLTCAVSGYSISSEYMVVGWIRQPP
terminal Fab GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
TM (IgG1
VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
Fab, IgG1
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3)))
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
NHKPSNTKVDKRVEPKSCGGGGSQVQLQESGPGLVKPSETLS
LTCAVSGYSISSEYMWGWIRQPPGKGLEWIGLIYHSGKTYYN
PSLKSRVTISVDTSKNQFSLKLSSVTAADTAVYYCAREADRG
YGDYGGHHAFDIWGQGTMVTVSSASTKGPSVFPLAPSSKSTS
GGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSG
LYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCD
KTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVV
DVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVS
VLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREP
QVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPE
NNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMH
EALHNHYTQKSLSLSPGK
191 ABP70
Full Heavy, C QVQLQESGPGLVKPSETLSLTCAVSGYSISSGLGWGWIRQPPG
terminal Fab KGLEWIGGIYESGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM, linker = AADTAVYYCAHERVRGYGDYGGHHAFDIWGQGTMVTVSSA
5mer (IgG1
STKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3)),
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
IgG1 Fab)
KPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGF
YPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKS
RWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKGGGGSQVQL
QESGPGLVKPSETLSLTCAVSGYSISSGLGWGWIRQPPGKGLE
140
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WIGGIYESGSTYYNPSLKSRVTISVDTSKNQFSLKLS SVTAAD
TAVYYCAHERVRGYGDYGGHHAFDIWGQGTMVTVS SASTK
GP SVFPLAP S SKSTS GGTAAL GCL VKDYFPEPVTVSWNS GAL T
SGVHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPS
NTKVDKRVEPKS C
192 ABP71
Full Heavy, C QVQLQESGPGLVKPSQTLSLTCTVSGGSISSGGAVVVSWIRQHP
terminal Fab GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSS
TM, linker = VTAADTAVYYCADENVRGYGDYGGHHAFDIWGQGTMVTVS
5mer (IgG1
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3)), S GAL
TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SLGTQTYICNV
IgG1 Fab)
NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLV
KGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSL SLSPGKGGGGSQ
VQLQE S GP GL VKP SQTL SLT CTVS GGS IS SGGAVWSWIRQHPG
KGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSV
TAADTAVYYCADENVRGYGDYGGHHAFDIWGQGTMVTVS S
ASTKGP SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNS
GALT S GVHTFPAVLQ S SGLYSLS SVVTVPS S SLGTQTYICNVN
HKPSNTKVDKRVEPKSC
193 ABP72
Full Heavy, C QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
terminal Fab KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM, linker = AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
5mer (IgG1 STKGP
SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3)), ALT S
GVHTFPAVLQ S SGLYSLS SVVTVP S S SLGTQTYICNVNH
IgG1 Fab)
KPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGF
YPSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKS
RWQQGNVFSCSVMHEALHNHYTQKSLSL SPGKGGGGSQVQL
QESGPGLVKPSETLSLTCAVSGYSIS SGAGWGWIRQPPGKGLE
WI GLIVH S GS TYYNP SLK SRVTI S VD T SKNQF SLKL S SVTAAD
TAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA STK
GP SVFPLAP S SKSTS GGTAAL GCL VKDYFPEPVTVSWNS GAL T
SGVHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPS
NTKVDKRVEPKS C
141
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194 ABP73
Full Heavy, C QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
terminal Fab KGPEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLS SVT
TM, linker = AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
5mer (IgG1 STKGP
SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3)),
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
IgG1 Fab)
KPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGF
YPSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKS
RWQQGNVFSCSVMHEALHNHYTQKSLSL SPGKGGGGSQVQL
QES GP GLVKP SETLSLTCAVSGYSIS SGAGWGWIRQPPGKGPE
WI GLIVH S GS TYYNP SLK SRVTI S VD T SKNQF SLKL S SVTAAD
TAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA STK
GP SVFPLAP S SKSTS GGTAAL GCL VKDYFPEPVTVSWNS GAL T
SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS
NTKVDKRVEPKS C
195 ABP74
Full Heavy, C QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
terminal Fab KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM, linker = AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
5mer (IgG1 STKGP
SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3)),
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
IgG1 Fab)
KPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGF
YPSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKS
RWQQGNVFSCSVMHEALHNHYTQKSLSL SPGKGGGGSQVQL
QESGPGLVKPSETLSLTCAVSGYSIS SGAGWGWIRQPPGKGLE
WI GLIVH S GS TYYNP SLK SRVTI S VD T SKNQF SLKL S SVTAAD
TAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA STK
GP SVFPLAP S SKSTS GGTAAL GCL VKDYFPEPVTVSWNS GAL T
SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS
NTKVDKRVEPKS C
196 ABP75
Full Heavy, C QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
terminal Fab KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
TM, linker = AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
5mer (IgG1 STKGP
SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSG
(G1m(3)),
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNH
IgG1 Fab)
KPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKP
142
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KDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHNAK
TKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALP
APIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGF
YPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTVDKS
RWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKGGGGSQVQL
QESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPGKGLE
WIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVTAAD
TAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSASTK
GPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT
SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS
NTKVDKRVEPKSC
197 ABP76 Full Heavy, C QVQLQESGPGLVKPSETLSLTCAVSGYSISSEYMWGWIRQPP
terminal Fab GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
TM, linker = VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
5mer (IgG1
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3)),
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
IgG1 Fab)
NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLV
KGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKGGGGSQ
VQLQESGPGLVKPSETLSLTCAVSGYSISSEYMWGWIRQPPG
KGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSSV
TAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVSS
ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNS
GALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN
HKPSNTKVDKRVEPKSC
198 ABP77 Full Heavy, C QVQLQESGPGLVKPSETLSLTCAVSGYSISSEYMWGWIRQPP
terminal Fab GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSS
TM, linker = VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
5mer (IgG1
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3)),
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
IgG1 Fab)
NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLV
KGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKGGGGSQ
VQLQESGPGLVKPSETLSLTCAVSGYSISSEYMWGWIRQPPG
143
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
KGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLSSV
TAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVSS
ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNS
GALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN
HKPSNTKVDKRVEPKSC
199 ABP78
Full Heavy, N QVQLVQSGAEVKRPGSSVKVSCKASGGTFSSYYISWVRQVPG
terminal Fab QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
TM (IgG1
SLRSEDTAVYYCARAGGGYARGDHYYGMDVWGQGTTVTVS
Fab, IgG1
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3)))
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
NHKPSNTKVDKRVEPKSCGGGGSQVQLVQSGAEVKRPGSSV
KVSCKASGGTFSSYYISWVRQVPGQGLEWMGGIIPVPGTANY
AQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARAGGG
YARGDHYYGMDVWGQGTTVTVSSASTKGPSVFPLAPSSKST
SGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSS
GLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSC
DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVV
VDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVV
SVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPRE
PQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQP
ENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVM
HEALHNHYTQKSLSLSPGK
200 ABP79
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYYISWVRQAPG
terminal Fab QRLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
TM (IgG1
SLRSEDTAVYYCARAGGGYARGDHYYGMDVWGQGTTVTVS
Fab, IgG1
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3)))
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
NHKPSNTKVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSV
KVSCKASGGTFSSYYISWVRQAPGQRLEWMGGIIPVPGTANY
AQKFQGRVTITADESTSTAYMELSSLRSEDTAVYYCARAGGG
YARGDHYYGMDVWGQGTTVTVSSASTKGPSVFPLAPSSKST
SGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSS
GLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSC
DKTHTCPPCPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVV
VDVSHEDPEVKFNWYVDGVEVHNAKTKPREEQYNSTYRVV
SVLTVLHQDWLNGKEYKCKVSNKALPAPIEKTISKAKGQPRE
PQVYTLPPSREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQP
ENNYKTTPPVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVM
HEALHNHYTQKSLSLSPGK
144
CA 03042727 2019-05-02
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PCT/US2017/062443
201 ABP80 Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSRGAISWVRQAP
terminal Fab GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTGTAYMEL
TM (IgG1
SSLRSEDTAVYYCARQSDYGLPRGMDVWGQGTTVTVSSAST
Fab, IgG1
KGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGA
(G1m(3)))
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
PSNTKVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVS
CKASGGTFSRGAISWVRQAPGQGLEWMGGIIPIEGTAYYAQK
FQGRVTITADESTGTAYMELSSLRSEDTAVYYCARQSDYGLP
RGMDVWGQGTTVTVSSASTKGPSVFPLAPSSKSTSGGTAALG
CLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSV
VTVPSSSLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPP
CPAPELLGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDP
EVKFNWYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQ
DWLNGKEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPP
SREEMTKNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTP
PVLDSDGSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHY
TQKSLSLSPGK
202 ABP81
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
terminal Fab QGLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYMELSS
TM (IgG1
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
Fab, IgG1
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
(G1m(3)))
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFSSYAISWVRQAPGQGLEWMGGIIPISGFTNYAQKFQGR
VTITADESTSTAYMELSSLRSEDTAVYYCAREGGHYYSGWPY
WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKD
YFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSS
SLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEL
LGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFN
WYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNG
KEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMT
KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLD SD
GSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLS
LSPGK
203 ABP82
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
terminal Fab QRLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYMELSS
TM (IgG1
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
Fab, IgG1
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
(G1m(3)))
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
145
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
GGTFSSYAISWVRQAPGQRLEWMGGIIPISGFTNYAQKFQGR
VTITADESTSTAYMELSSLRSEDTAVYYCAREGGHYYSGWPY
WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKD
YFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSS
SLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEL
LGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFN
WYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNG
KEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMT
KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLD SD
GSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLS
LSPGK
204 ABP83
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
terminal Fab QGLEWMGGIIPISGFTNYAQKFQGKVTITADESTSTAYMEL SS
TM (IgG1
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
Fab, IgG1
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
(G1m(3)))
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFSSYAISWVRQAPGQGLEWMGGIIPISGFTNYAQKFQGK
VTITADESTSTAYMELSSLRSEDTAVYYCAREGGHYYSGWPY
WGQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKD
YFPEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSS
SLGTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPEL
LGGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFN
WYVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNG
KEYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMT
KNQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLD SD
GSFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLS
LSPGK
205 ABP84
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
terminal Fab GQGLEWMGGIIPIFGEAQYAQRFQGRVTITADESTSTAYMELS
TM (IgG1
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
Fab, IgG1
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
(G1m(3)))
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFVRYAISWVRQAPGQGLEWMGGIIPIFGEAQYAQRFQGR
VTITADESTSTAYMELSSLRSEDTAVYYCAREGYYYGALPYW
GQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYF
PEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSL
GTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELL
GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW
146
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
YVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGK
EYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTK
NQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG
SFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLS
PGK
206 ABP85
Full Heavy, N QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
terminal Fab GQGLEWMGGIIPIFGEAQYAQKFRGRATITADESTSTAYMELS
TM (IgG1
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
Fab, IgG1
SVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
(G1m(3)))
VHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNT
KVDKRVEPKSCGGGGSQVQLVQSGAEVKKPGSSVKVSCKAS
GGTFVRYAISWVRQAPGQGLEWMGGIIPIFGEAQYAQKFRGR
ATITADESTSTAYMELSSLRSEDTAVYYCAREGYYYGALPYW
GQGTLVTVSSASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYF
PEPVTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSL
GTQTYICNVNHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELL
GGPSVFLFPPKPKDTLMISRTPEVTCVVVDVSHEDPEVKFNW
YVDGVEVHNAKTKPREEQYNSTYRVVSVLTVLHQDWLNGK
EYKCKVSNKALPAPIEKTISKAKGQPREPQVYTLPPSREEMTK
NQVSLTCLVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDG
SFFLYSKLTVDKSRWQQGNVFSCSVMHEALHNHYTQKSLSLS
PGK
207 ABP86
Full Heavy, C QVQLVQSGAEVKRPGSSVKVSCKASGGTFSSYYISWVRQVPG
terminal Fab QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
TM, linker = SLRSEDTAVYYCARAGGGYARGDHYYGMDVWGQGTTVTVS
5mer (IgG1
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3)),
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
IgG1 Fab)
NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLV
KGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSLSLSPGKGGGGSQ
VQLVQSGAEVKRPGSSVKVSCKASGGTFSSYYISWVRQVPGQ
GLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELSS
LRSEDTAVYYCARAGGGYARGDHYYGMDVWGQGTTVTVSS
ASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNS
GALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVN
HKPSNTKVDKRVEPKSC
147
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
208 ABP87
Full Heavy, C QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYYISWVRQAPG
terminal Fab QRLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
TM, linker = SLRSEDTAVYYCARAGGGYARGDHYYGMDVWGQGTTVTVS
5mer (IgG1
SASTKGPSVFPLAP S SKSTSGGTAALGCLVKDYFPEPVTVSWN
(G1m(3)), S GAL
TS GVH TFP AVLQ S SGLYSL S SVVTVPS S SLGTQTYICNV
IgG1 Fab)
NHKPSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPP
KPKDTLMISRTPEVTCVVVDVSHEDPEVKFNWYVDGVEVHN
AKTKPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNK
ALPAPIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLV
KGFYP SD IAVEWE SNGQPENNYKTTPPVLD SD GSFFLYSKLTV
DKSRWQQGNVFSCSVMHEALHNHYTQKSL SLSPGKGGGGSQ
VQLVQ SGAEVKKPGS SVKVS CKASGGTFS SYYISWVRQAPGQ
RLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS S
LRSEDTAVYYCARAGGGYARGDHYYGMDVWGQGTTVTVS S
ASTKGP SVFPL AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNS
GALT S GVHTFPAVLQ S SGLYSLS SVVTVPS S SLGTQTYICNVN
HKPSNTKVDKRVEPKSC
209 ABP88 Full Heavy, C QVQLVQSGAEVKKPGSSVKVSCKASGGTFSRGAISWVRQAP
terminal Fab GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTGTAYMEL
TM, linker = S SLRSEDTAVYYCARQSDYGLPRGMDVWGQGTTVTVS SAS T
5mer (IgG1
KGPSVFPLAPS SKSTS GGTAAL GCLVKDYFPEPVTVSWNS GA
(G1m(3)),
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
IgG1 Fab)
PSNTKVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPK
DTLMISRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKT
KPREEQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPA
PIEKTISKAKGQPREPQVYTLPPSREEMTKNQVSLTCLVKGFY
PSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLYSKLTVDKSR
WQQGNVFSCSVMHEALHNHYTQKSLSLSPGKGGGGSQVQL
VQSGAEVKKPGS SVKVSCKASGGTFSRGAISWVRQAPGQGLE
WMGGIIPIEGTAYYAQKFQGRVTITADESTGTAYMELS SLRSE
DTAVYYCARQSDYGLPRGMDVWGQGTTVTVS SASTKGPSVF
PLAPS SKST S GGTAALGCLVKDYFPEPVTVS WNS GAL TS GVH
TFPAVLQS SGLYSL S SVVTVPS S SLGTQTYICNVNHKP SNTKV
DKRVEPKSC
210 ABP89
Full Heavy, C QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
terminal Fab QGLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYMEL S S
TM, linker = LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
5mer (IgG1 SVFPL
AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTS G
(G1m(3)),
VHTFPAVLQS SGLYSLS SVVTVPS S SL GTQTYI CNVNHKP S NT
IgG1 Fab)
KVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
148
CA 03042727 2019-05-02
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PCT/US2017/062443
I SRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPRE
EQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TISKAKGQPREPQVYTLPP SREEMTKNQVSL TCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFS CSVMHEALHNHYTQKSL SL SP GKGGGG S QVQLVQ S
GAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPGQGLEWM
GGIIPISGFTNYAQKFQGRVTITADESTSTAYMEL S SLRSEDTA
VYYCAREGGHYYSGWPYWGQGTLVTVS SASTKGPSVFPLAP
S SKS TS GGTAAL GCL VKDYFPEPVTVSWNS GALTS GVHTFP A
VLQS SGLYSL S SVVTVPS S SL GTQTYICNVNHKPSNTKVDKRV
EPKSC
211 ABP90
Full Heavy, C QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPG
terminal Fab QRLEWMGGIIPI S GFTNYAQKFQGRVTITADES TS TAYMEL S S
TM, linker = LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
5mer (IgG1 SVFPL
AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTS G
(G1m(3)),
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
IgG1 Fab)
KVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
I SRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPRE
EQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TI SKAKGQPREPQVYTLPP SREEMTKNQVSL TCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFS CSVMHEALHNHYTQKSL SL SP GKGGGG S QVQLVQ S
GAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPGQRLEWM
GGIIPISGFTNYAQKFQGRVTITADESTSTAYMEL S SLRSEDTA
VYYCAREGGHYYSGWPYWGQGTLVTVS SASTKGPSVFPLAP
S SKS TS GGTAAL GCL VKDYFPEPVTVSWNS GALTS GVHTFP A
VLQS SGLYSL S SVVTVPS S SL GTQTYICNVNHKPSNTKVDKRV
EPKSC
212 ABP91
Full Heavy, C QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPG
terminal Fab QGLEWMGGIIPISGFTNYAQKFQGKVTITADESTSTAYMEL S S
TM, linker = LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
5mer (IgG1 SVFPL
AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTS G
(G1m(3)),
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
IgG1 Fab)
KVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
I SRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPRE
EQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TI SKAKGQPREPQVYTLPP SREEMTKNQVSL TCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFS CSVMHEALHNHYTQKSL SL SP GKGGGG S QVQLVQ S
GAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPGQGLEWM
149
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GGIIPISGFTNYAQKFQGKVTITADESTSTAYMEL S SLRSEDTA
VYYCAREGGHYYSGWPYWGQGTLVTVS SASTKGPSVFPLAP
S SKS TS GGTAAL GCL VKDYFPEPVTVSWNS GALTS GVHTFP A
VLQS SGLYSL S SVVTVPS S SL GTQTYICNVNHKPSNTKVDKRV
EPKSC
213 ABP92
Full Heavy, C QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
terminal Fab GQGLEWMGGIIPIFGEAQYAQRFQGRVTITADESTSTAYMEL S
TM, linker = SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
5mer (IgG1 SVFPL
AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTS G
(G1m(3)),
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
IgG1 Fab)
KVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
I SRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPRE
EQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TI SKAKGQPREPQVYTLPP SREEMTKNQVSL TCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFS CSVMHEALHNHYTQKSL SL SP GKGGGG S QVQLVQ S
GAEVKKPGS SVKVS CKASGGTFVRYAISWVRQAPGQGLEWM
GGIIPIFGEAQYAQRFQGRVTITADESTSTAYMEL S SLRSEDTA
VYYCAREGYYYGALPYWGQGTLVTVS S A S TKGP S VFPL AP S S
KST S GGTAAL GCLVKDYFPEPVTVSWNS GAL TS GVHTFPAVL
QS SGLYSL S SVVTVP S S SLGTQTYICNVNHKPSNTKVDKRVEP
KSC
214 ABP93
Full Heavy, C QVQLVQSGAEVKKPGSSVKVSCKASGGTFVRYAISWVRQAP
terminal Fab GQGLEWMGGIIPIF GEAQYAQKFRGRATITADE ST STAYMEL S
TM, linker = SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVSSASTKGP
5mer (IgG1 SVFPL
AP S SKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTS G
(G1m(3)),
VHTFPAVLQS SGLYSL S SVVTVPS S SL GTQTYI CNVNHKP S NT
IgG1 Fab)
KVDKRVEPKSCDKTHTCPPCPAPELLGGPSVFLFPPKPKDTLM
I SRTPEVTCVVVDVSHEDPEVKFNWYVD GVEVHNAKTKPRE
EQYNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKALPAPIEK
TI SKAKGQPREPQVYTLPP SREEMTKNQVSL TCLVKGFYP SDI
AVEWESNGQPENNYKTTPPVLD SD GSFFLYSKL TVDKSRWQ
QGNVFS CSVMHEALHNHYTQKSL SL SP GKGGGG S QVQLVQ S
GAEVKKPGS SVKVS CKASGGTFVRYAISWVRQAPGQGLEWM
GGIIPIFGEAQYAQKFRGRATITADESTSTAYMEL S SLR SED TA
VYYCAREGYYYGALPYWGQGTLVTVS S A S TKGP S VFPL AP S S
KST S GGTAAL GCLVKDYFPEPVTVSWNS GAL TS GVHTFPAVL
QS SGLYSL S SVVTVP S S SLGTQTYICNVNHKPSNTKVDKRVEP
KSC
150
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8 ABP35 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
18 ABP36 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
25 ABP37 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLKYGVP SRF S GS GS GTDFTL TIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
33 ABP38 Full Light D IQMTQ
SP S SL S A S VGDRVTITCRA SK SID SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
39 ABP39 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAD SLQSGVP SRF S GS GS GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
43 ABP40 Full Light D IQMTQ
SP S SL S A S VGDRVTITCRA S Q SINSYLNWYQQKP GKA
PKLLIYAASSLDSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
52 ABP41 Full Light D IQMTQ
SP S SL SASVGDRVTITCGASQSISTYLNWYQQKPGKA
PKLLIYSASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYC
QQEYNTPPSFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASV
VCLLNNFYPREAKVQWKVDNALQ S GN S QES V 1EQD SKD S TY
SLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP42 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
151
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
8 ABP43 Full Light D IQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
107 ABP44 Full Light D IVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
109 ABP45 Full Light D IQL TQ SP S S VS A S VGDRVTIT CRA S Q GI S
SWLAWYQQKPGK
APKLLIYGASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATY
YCQQASLFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTA
SVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD S
TY SL S STLTL SKADYEKHKVYACEVTHQGL S SPVTKSFNRGE
C
111 ABP46 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113 ABP47 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP48 Full Light D IQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP49 Full Light D IQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
107 ABP50 Full Light D IVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
152
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LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
107 ABP51 Full Light D IVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
109 ABP52 Full Light D IQL TQ SP S S VS A S VGDRVTIT CRA S Q GI S
SWLAWYQQKPGK
APKLLIYGASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATY
YCQQASLFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTA
SVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD S
TY SL S STLTL SKADYEKHKVYACEVTHQGL S SPVTKSFNRGE
C
111 ABP53 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
111 ABP54 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
111 ABP55 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113 ABP56 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113 ABP57 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
153
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
107 ABP58 Full Light D IVMTQ
SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQALQTPITFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
107 ABP59 Full Light D IVMTQ
SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQALQTPITFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
109 ABP60 Full Light D IVMTQ
SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQALQTPITFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
111 ABP61 Full Light D IVMTQ
SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQALQTPITFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
8 ABP62 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
18 ABP63 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
25 ABP64 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLKYGVP SRF S GS GS GTDFTL TIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
154
CA 03042727 2019-05-02
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33 ABP65 Full Light D IQMTQ
SP S SL S A S VGDRVTITCRA SK SID SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
33 ABP66 Full Light D IQMTQ
SP S SL S A S VGDRVTITCRA SK SID SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
39 ABP67 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAD SLQSGVP SRF S GS GS GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
43 ABP68 Full Light D IQMTQ
SP S SL S A S VGDRVTITCRA S Q SINSYLNWYQQKP GKA
PKLLIYAASSLDSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
52 ABP69 Full Light D IQMTQ
SP S SL SASVGDRVTITCGASQSISTYLNWYQQKPGKA
PKLLIYSASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYC
QQEYNTPPSFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASV
VCLLNNFYPREAKVQWKVDNALQ S GN S QES V 1EQD SKD S TY
SLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
8 ABP70 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
18 ABP71 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
25 ABP72 Full Light D IQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLKYGVP SRF S GS GS GTDFTL TIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
155
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
33 ABP73 Full Light D IQMTQ SP S SL S A S VGDRVTITCRA SK SID
SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
33 ABP74 Full Light D IQMTQ SP S SL S A S VGDRVTITCRA SK SID
SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
39 ABP75 Full Light D IQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAD SLQSGVP SRF S GS GS GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
43 ABP76 Full Light D IQMTQ SP S SL S A S VGDRVTITCRA S Q SINSYLNWYQQKP
GKA
PKLLIYAASSLDSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
52 ABP77 Full Light D IQMTQ SP S SL SASVGDRVTITCGASQSISTYLNWYQQKPGKA
PKLLIYSASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYC
QQEYNTPPSFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASV
VCLLNNFYPREAKVQWKVDNALQ S GN S QES V 1EQD SKD S TY
SLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
107 ABP78 Full Light D IVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
107 ABP79 Full Light D IVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
109 ABP80 Full Light D IQL TQ SP S S VS A S VGDRVTIT CRA S Q GI S
SWLAWYQQKPGK
APKLLIYGASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATY
YCQQASLFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTA
156
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SVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD S
TY SL S STLTL SKADYEKHKVYACEVTHQGL S SPVTKSFNRGE
C
111 ABP81 Full Light
EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
111 ABP82 Full Light
EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
111 ABP83 Full Light
EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113 ABP84 Full Light
EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113 ABP85 Full Light
EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
107 ABP86 Full Light D IVMTQ
SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
107 ABP87 Full Light D IVMTQ
SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
157
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109 ABP88 Full Light D IQL TQ SP S S VS A S VGDRVTIT CRA S Q GI S
SWLAWYQQKPGK
APKLLIYGASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATY
YCQQASLFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTA
SVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD S
TY SL S STLTL SKADYEKHKVYACEVTHQGL S SPVTKSFNRGE
C
111 ABP89 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
111 ABP90 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
111 ABP91 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113 ABP92 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
113 ABP93 Full Light EIVLTQSPATLSLSPGERATL SCRASQSVS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
114 ABP94 Full Heavy QVQLQESGPGLVKPSETLSLTCAVSGYSISSGLGWGWIRQPPG
KGLEWIGGIYESGSTYYNPSLKSRVTISVDTSKNQFSLKLS SVT
AADTAVYYCAHERVRGYGDYGGHHAFDIWGQGTMVTVS SA
STKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSG
ALT S GVHTFPAVLQ S SGLYSLS SVVTVP S S SLGTKTYTCNVDH
KPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDT
LMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKP
REEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIE
KTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPS
158
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DIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLQE
S GP GL VKP SETL SLTCAVSGYSIS SGLGWGWIRQPPGKGLEWI
GGIYESGSTYYNPSLKSRVTISVDTSKNQFSLKLS SVTAADTA
VYYCAHERVRGYGDYGGHHAFDIWGQGTMVTVSSASTKGP
S VFPL AP S SK S T S GGTAAL GCLVKDYFPEP VTVS WNS GALT S G
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNT
KVDKRVEPKSC
115 ABP95 Full Heavy
QVQLQESGPGLVKPSQTLSLTCTVSGGSIS SGGAVVVSWIRQHP
GKGLEWIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCADENVRGYGDYGGHHAFDIWGQGTMVTVS
SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNV
DHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPK
DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
PSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSR
WQEGNVFSCSVMHEALHNHYTQKSLSLSLGKGGGGSQVQLQ
ESGPGLVKPSQTLSLTCTVSGGSISSGGAVVVSWIRQHPGKGLE
WIGGIAYSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVTAAD
TAVYYCADENVRGYGDYGGHHAFDIWGQGTMVTVSSASTK
GP SVFPLAP S SKSTS GGTAAL GCLVKDYFPEPVTVSWNS GAL T
SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS
NTKVDKRVEPKSC
116 ABP96 Full Heavy
QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSA
STKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSG
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDH
KPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDT
LMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKP
REEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIE
KTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPS
DIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLQE
S GP GL VKP SETL SLTCAVSGYSIS SGAGWGWIRQPPGKGLEWI
GLIVH S G STYYNP SLKSRVTIS VDT SKNQF SLKL S SVTAADTA
VYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSASTKGPS
VFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSG
159
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VHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNT
KVDKRVEPKSC
117 ABP97 Full Heavy
QVQLQESGPGLVKPSETLSLTCAVSGYSISSGAGWGWIRQPPG
KGPEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLS SVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA
STKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSG
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDH
KPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDT
LMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKP
REEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIE
KTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPS
DIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLQE
S GP GL VKP SETL SLTCAVSGYSIS SGAGWGWIRQPPGKGPEWI
GLIVH S G STYYNP SLKSRVTIS VDT SKNQF SLKL S SVTAADTA
VYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SASTKGPS
VFPL AP S SKST S GGTAALGCLVKDYFPEPVTVSWNS GALT S G
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNT
KVDKRVEPKSC
118 ABP98 Full Heavy
QVQLQESGPGLVKPSETLSLTCAVSGYSIS SGAGWGWIRQPPG
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA
STKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSG
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDH
KPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDT
LMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKP
REEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIE
KTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPS
DIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLQE
S GP GL VKP SETL SLTCAVSGYSIS SGAGWGWIRQPPGKGLEWI
GLIVH S G STYYNP SLKSRVTIS VDT SKNQF SLKL S SVTAADTA
VYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SASTKGPS
VFPL AP S SKST S GGTAALGCLVKDYFPEPVTVSWNS GALT S G
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNT
KVDKRVEPKSC
119 ABP99 Full Heavy
QVQLQESGPGLVKPSETLSLTCAVSGYSIS SGAGWGWIRQPPG
KGLEWIGLIVHSGSTYYNPSLKSRVTISVDTSKNQFSLKLSSVT
AADTAVYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVS SA
STKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSG
160
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PCT/US2017/062443
ALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDH
KPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDT
LMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKP
REEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIE
KTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPS
DIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRW
QEGNVFSCSVMHEALHNHYTQKSL SLSL GKGGGGSQVQLQE
S GP GL VKP SETL SLTCAVSGYSIS SGAGWGWIRQPPGKGLEWI
GLIVH S G STYYNP SLKSRVTIS VDT SKNQF SLKL S SVTAADTA
VYYCVLESVRGYGDYGGHHAFDIWGQGTMVTVSSASTKGPS
VFPL AP S SKST S GGTAALGCLVKDYFPEPVTVSWNS GALT S G
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNT
KVDKRVEPKSC
120 ABP100 Full Heavy
QVQLQESGPGLVKPSETLSLTCAVSGYSIS SEYMWG
WIRQPPGKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQF
SLKLSSVTAADTAVYYCAREADRGYGDYGGHHAFDIWGQG
TMVTVS SASTKGPSVFPLAPCSRSTSESTAAL GCLVKDYFPEP
VTVSWNSGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTK
TYTCNVDHKPSNTKVDKRVESKYGPPCPPCPAPEFL GGPSVF
LFPPKPKDTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVE
VHNAKTKPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVS
NKGLPS SIEKTISKAKGQPREPQVYTLPP SQEEMTKNQVSLTC
LVKGFYPSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRL
TVDKSRWQEGNVFSCSVMHEALHNHYTQKSL SLSL GKGGGG
SQVQLQES GP GLVKP SETL SL TCAVS GY SI S SEYMVVGWIRQPP
GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
SASTKGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWN
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNV
NHKPSNTKVDKRVEPKSC
121 ABP 101 Full Heavy QVQLQE S
GP GLVKP SETL SLT CAVS GY SI S SEYMWGWIRQPP
GKGLEWIGLIYHSGKTYYNPSLKSRVTISVDTSKNQFSLKLS S
VTAADTAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVS
SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNV
DHKPSNTKVDKRVESKYGPPCPPCPAPEFL GGPSVFLFPPKPK
DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
PSDIAVEWESNGQPENNYKTTPPVLD SD GSFFLY SRL TVDK SR
161
CA 03042727 2019-05-02
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PCT/US2017/062443
WQEGNVFSCSVMHEALHNHYTQKSLSLSLGKGGGGSQVQLQ
ES GP GLVKP S ETL SLTCAVS GY SI S SEYMWGWIRQPPGKGLE
WIGLIYH S GKTYYNP SLK SRVTI S VD T SKNQF SLKL S SVTAAD
TAVYYCAREADRGYGDYGGHHAFDIWGQGTMVTVSSASTK
GPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALT
SGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPS
NTKVDKRVEPKSC
122 ABP102 Full Heavy
QVQLVQSGAEVKRPGSSVKVSCKASGGTFSSYYISWVRQVPG
QGLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
SLRSEDTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVS
SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNV
DHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPK
DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
PSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSR
WQEGNVFSCSVMHEALHNHYTQKSLSLSLGKGGGGSQVQLV
QSGAEVKRPGSSVKVSCKASGGTFSSYYISWVRQVPGQGLE
WMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELSSLRSE
DTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVSSAST
KGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGA
LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
PSNTKVDKRVEPKSC
123 ABP103 Full Heavy
QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYYISWVRQAPG
QRLEWMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELS
SLRSEDTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVS
SASTKGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWN
SGALTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNV
DHKPSNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPK
DTLMISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKT
KPREEQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS
SIEKTISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFY
PSDIAVEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSR
WQEGNVFSCSVMHEALHNHYTQKSLSLSLGKGGGGSQVQLV
QSGAEVKKPGSSVKVSCKASGGTFSSYYISWVRQAPGQRLE
WMGGIIPVPGTANYAQKFQGRVTITADESTSTAYMELSSLRSE
DTAVYYCARAGGGYARGDHYYGMDVVVGQGTTVTVSSAST
KGPSVFPLAPSSKSTSGGTAALGCLVKDYFPEPVTVSWNSGA
162
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LTSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHK
PSNTKVDKRVEPKSC
124 ABP104 Full Heavy QVQLVQSGAEVKKPGSSVKVSCKASGGTFSRGAISWVRQAP
GQGLEWMGGIIPIEGTAYYAQKFQGRVTITADESTGTAYMEL
S SLR SED TAVYYCARQ SDYGLPRGMD VVVGQ GTTVTVS SAS T
KGPSVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGAL
TSGVHTFPAVLQSSGLYSLSSVVTVPSSSLGTKTYTCNVDHKP
SNTKVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLM
ISRTPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPRE
EQFNSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKT
ISKAKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIA
VEWESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEG
NVFSCSVMHEALHNHYTQKSLSL SLGKGGGGSQVQLVQ S GA
EVKKPGS SVKVS CKASGGTFSRGAISWVRQAPGQGLEWMGG
IIPIEGTAYYAQKFQGRVTITADESTGTAYMELS SLRSEDTAV
YYCARQSDYGLPRGMDVVVGQGTTVTVSSASTKGPSVFPLAP
SSKSTSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPA
VLQSSGLYSLSSVVTVPSSSLGTQTYICNVNHKPSNTKVDKRV
EPKSC
125 ABP105 Full Heavy QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
QGLEWMGGIIPISGFTNYAQKFQGRVTITADESTSTAYMEL SS
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
SVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSG
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTKTYTCNVDHKPSNT
KVDKRVESKYGPPCPPCPAPEFLGGPSVFLFPPKPKDTLMISR
TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPSSIEKTISK
AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYPSDIAVE
WESNGQPENNYKTTPPVLDSDGSFFLYSRLTVDKSRWQEGN
VFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLVQSGAE
VKKPGS SVKVSCKASGGTFS SYAISWVRQAPGQGLEWMGGII
PI S GFTNYAQKFQ GRVTITADE STS TAYMEL S SLRSEDTAVYY
CAREGGHYYSGWPYWGQGTLVTVS SASTKGP SVFPL AP S SKS
TSGGTAALGCLVKDYFPEPVTVSWNSGALTSGVHTFPAVLQS
SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNTKVDKRVEPKS
C
126 ABP106 Full Heavy QVQLVQSGAEVKKPGSSVKVSCKASGGTFSSYAISWVRQAPG
QRLEWMGGIIPI S GFTNYAQKFQGRVTITADES TS TAYMEL S S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVSSASTKGP
SVFPLAPCSRSTSESTAALGCLVKDYFPEPVTVSWNSGALTSG
163
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VHTFPAVLQS SGLYSLS SVVTVPS S SLGTKTYTCNVDHKPSNT
KVDKRVE SKYGPPCPP CPAPEFLGGP SVFLFPPKPKDTLMI SR
TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKTISK
AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYP SDIAVE
WE SNGQPENNYKTTPPVLD SD GSFFLYSRLTVDKSRWQEGN
VFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLVQSGAE
VKKPGS SVKVSCKASGGTFS SYAISWVRQAPGQRLEWMGGII
PI S GFTNYAQKFQ GRVTITADE STS TAYMEL S SLRSEDTAVYY
CAREGGHYYSGWPYWGQGTLVTVS SASTKGP SVFPL AP S SKS
TS GGTAAL GCLVKDYFPEPVTVSWNS GALT S GVHTFPAVLQ S
SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNTKVDKRVEPKS
C
127 ABP107 Full Heavy
QVQLVQSGAEVKKPGS SVKVSCKASGGTFS SYAISWVRQAPG
QGLEWMGGIIPISGFTNYAQKFQGKVTITADESTSTAYMEL S S
LRSEDTAVYYCAREGGHYYSGWPYWGQGTLVTVS SA STKGP
SVFPL APC SRSTSE STAALGCLVKDYFPEPVTVSWNS GAL TS G
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTKTYTCNVDHKPSNT
KVDKRVE SKYGPPCPP CPAPEFLGGP SVFLFPPKPKDTLMI SR
TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKTISK
AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYP SDIAVE
WE SNGQPENNYKTTPPVLD SD GSFFLYSRLTVDKSRWQEGN
VFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLVQSGAE
VKKPGS SVKVSCKASGGTFS SYAISWVRQAPGQGLEWMGGII
PI S GFTNYAQKFQ GKVTITADEST STAYMEL S SLRSEDTAVYY
CAREGGHYYSGWPYWGQGTLVTVS SASTKGP SVFPL AP S SKS
TS GGTAAL GCLVKDYFPEPVTVSWNS GALT S GVHTFPAVLQ S
SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNTKVDKRVEPKS
C
128 ABP108 Full Heavy QVQLVQS
GAEVKKP GS SVKVSCKASGGTFVRYAISWVRQAP
GQGLEWMGGIIPIFGEAQYAQRFQGRVTITADESTSTAYMELS
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVS SASTKGP
SVFPL APC SRSTSE STAALGCLVKDYFPEPVTVSWNS GAL TS G
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTKTYTCNVDHKPSNT
KVDKRVE SKYGPPCPP CPAPEFLGGP SVFLFPPKPKDTLMI SR
TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKTISK
AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYP SDIAVE
WE SNGQPENNYKTTPPVLD SD GSFFLYSRLTVDKSRWQEGN
164
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VFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLVQSGAE
VKKPGS SVKVSCKASGGTFVRYAISWVRQAPGQGLEWMGGI
IPIFGEAQYAQRFQGRVTITADESTSTAYMELS SLRSEDTAVY
YCAREGYYYGALPYWGQGTLVTVS SASTK GP SVFPL AP S SKS
TS GGTAAL GCLVKDYFPEPVTVSWNS GALT S GVHTFPAVLQ S
SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNTKVDKRVEPKS
C
129 ABP109 Full Heavy QVQLVQS
GAEVKKP GS SVKVSCKASGGTFVRYAISWVRQAP
GQGLEWMGGIIPIF GEAQYAQKFRGRATITADE ST STAYMEL S
SLRSEDTAVYYCAREGYYYGALPYWGQGTLVTVS SASTKGP
SVFPL APC SRSTSE STAALGCLVKDYFPEPVTVSWNS GAL TS G
VHTFPAVLQS SGLYSLS SVVTVPS S SLGTKTYTCNVDHKPSNT
KVDKRVE SKYGPPCPP CPAPEFLGGP SVFLFPPKPKDTLMI SR
TPEVTCVVVDVSQEDPEVQFNWYVDGVEVHNAKTKPREEQF
NSTYRVVSVLTVLHQDWLNGKEYKCKVSNKGLPS SIEKTISK
AKGQPREPQVYTLPPSQEEMTKNQVSLTCLVKGFYP SDIAVE
WE SNGQPENNYKTTPPVLD SD GSFFLYSRLTVDKSRWQEGN
VFSCSVMHEALHNHYTQKSL SLSLGKGGGGSQVQLVQSGAE
VKKPGS SVKVSCKASGGTFVRYAISWVRQAPGQGLEWMGGI
IPIFGEAQYAQKFRGRATITADESTSTAYMELS SLRSEDTAVY
YCAREGYYYGALPYWGQGTLVTVS SASTK GP SVFPL AP S SKS
TS GGTAAL GCLVKDYFPEPVTVSWNS GALT S GVHTFPAVLQ S
SGLYSLS SVVTVPS S SLGTQTYICNVNHKPSNTKVDKRVEPKS
C
8 ABP94 Full Light DIQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYATPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
18 ABP95 Full Light DIQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLQ S GVP SRF S GS G S GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
25 ABP96 Full Light DIQMTQ
SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAS SLKYGVP SRF S GS GS GTDFTL TIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
165
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
33 ABP97 Full Light D IQMTQ SP S SL S A S VGDRVTITCRA SK SID
SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
33 ABP98 Full Light D IQMTQ SP S SL S A S VGDRVTITCRA SK SID
SYLNWYQQKPGKA
PKLLIYAASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
39 ABP99 Full Light D IQMTQ SP S SL SASVGDRVTITCRASQSIS SYLNWYQQKPGKA
PKLLIYAAD SLQSGVP SRF S GS GS GTDFTLTIS SLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
43 ABP100 Full Light DIQMTQ SP S SL SA SVGDRVTITCRASQ S INSYLNWYQQKPGKA
PKLLIYAASSLDSGVPSRFSGSGSGTDFTLTISSLQPEDFATYY
CQQEYNTPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
52 ABP101 Full Light D IQMTQ SP S SL SAS VGDRVTITC GASQ SISTYLNWYQQKP
GKA
PKLLIYSASSLESGVPSRFSGSGSGTDFTLTISSLQPEDFATYYC
QQEYNTPPSFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTASV
VCLLNNFYPREAKVQWKVDNALQ S GN S QES V 1EQD SKD S TY
SLSSTLTLSKADYEKHKVYACEVTHQGLSSPVTKSFNRGEC
107 ABP102 Full Light D IVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
107 ABP103 Full Light D IVMTQ SPL SLPVTP GEPASIS CRS SQSLLH SNGYNYLDWYLQ
KPGQSPQLLIYLGSNRASGVPDRFSGSGSGTDFTLKISRVEAE
DVGVYYCMQKIGTPLTFGGGTKVEIKRTVAAPSVFIFPPSDEQ
LKSGTASVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTE
QD SKD S TY SL S STLTLSKADYEKHKVYACEVTHQGL S SPVTK
SFNRGEC
109 ABP104 Full Light DIQLTQSPSSVSASVGDRVTITCRASQGISSWLAWYQQKPGK
APKLLIYGASSLQSGVPSRFSGSGSGTDFTLTISSLQPEDFATY
YCQQASLFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTA
166
CA 03042727 2019-05-02
WO 2018/094300
PCT/US2017/062443
SVVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD S
TY SL S STLTL SKADYEKHKVYACEVTHQGL S SPVTKSFNRGE
C
111 ABP105 Full Light
EIVLTQSPATLSL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
111 ABP106 Full Light
EIVLTQSPATLSL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
111 ABP107 Full Light
EIVLTQSPATLSL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQRLVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
113 ABP108 Full Light
EIVLTQSPATLSL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
113 ABP109 Full Light
EIVLTQSPATLSL SP GERATL S CRA S Q S VS SYLAWYQQKPGQA
PRLLIYDASNRATGIPARF S GS GS GTDFTLTI S SLEPEDFAVYY
CQQHSVFPPTFGGGTKVEIKRTVAAPSVFIFPPSDEQLKSGTAS
VVCLLNNFYPREAKVQWKVDNALQSGNSQESVTEQD SKD ST
YSL S STLTLSKADYEKHKVYACEVTHQGLS SPVTKSFNRGEC
167
