Note: Descriptions are shown in the official language in which they were submitted.
WO 93~07892 2 1 % ~ 3 ~ 7 PC~/IJS92/088~3
Compo~ition and Treatm~nt ~ith Biologically
Act~ve Peptid~s and l~ ibiotic
This application is a ce~ntinuation-in-part of Applica'ciDn
5erial No. 402,642, filed Septem~er S, 1989, which is a
continuation-in-part of Application 5erial No. 339"292, filed
Apri 1 17, 1989 .
This invention relates to biologically activ~ pept~d~s and
prc~teins, and more partic:ularly to compositlona; and use~
involving }:~iologically active pept~des or proteins and a~
antibicstic; in particular an An~ib~o~lc whic~l i2; ~ot an ior~
channel-forming peptide or protein, or derlvativ~2s or analogues
thereof .
I t has been d~ sclo~ed t~dat agent~ suc~ a~ polymyxin
norl~peptide, which di~rupt the outer membrane o~ Grnm-~egative
bacteria, increase the poterlcy of certain antil:~iotics,- J3ucl~ ~
~u~idic al~id, novobiocirl, and erythromycin, again~t th~ organi~ms
Vil~anen, et al, "5u cept{billty of Bram~N~gatîv~ Bacteria to
Polymyxin B Nonapeptide, " =~
Vol. 2~, No. 6, Pgs. 701-70~ (June 1984~. Vil~neal, et al, in
~5uscepti3bility of Gram~ Negative Bacteria to the Synerglst~c
Bactericidal Action of Serum and PolylTyxin B Nonapeptlde, n _ n.
J. Microbiol.Vol . ~2 pgs. 66-69 ( $986), disclo~e a ~ynergi~tic
efect of polymyxin B n~napeptide and serum in ~actericidal '
action against E. coli strain~, strains of 5almonella ~yphimurium,
Xlebisiella species, Enterobacter cloacae, P eudomona~
WO 93/07892 PCT/US92/08823
2120337
influenzae. Vaara, et al., have disclosed a ~ynergistic effect
of polymyxin B nonapeptide and novobiocin, fusidic acid,
ery~hromycin, clindamycin, nafcillin, and cloxacillin against
smooth encapsulated E.coli and smooth Salmonella tY~himurium.
Antimicrobial Aaents and Chemotherapy, Vol. 24, No. 1, pg~.
107-113. (July 1983).
In accordance with an aspect of the pre~ent invention, there
is provided a compos~tion which includes at least one
biologically active peptide or protein, said peptid~ or protein
being an ion-channel form~ng peptide or protein; and an
antibiotic.
In accordance with anoth~r a~pect of the pre~ent invention,
there is provided a process wherein there is administered to a
host at least one biologically active peptide or protein, said
peptide or protein being an ion channel-forming peptide or
protein, and ~n antibiotic.
The antibiotic in which i~ combined is different from such
peptide or protein; i.e., ~uch antibiotic iQ other than an ion
channel-forming peptide or protein.
Although the invention i~ not to be l~mited to~nny
theoretical reasoning, it is believed that the peptides or
proteins employed in the present invention interact with the
membranes of bacterial cells and such interaction enhances the
ability o the above-mentioned antibiotic~ to cro~s the me~brane.
It i8 to be u~derstood~ however~ that ~e scope of the invention
i~ not limited to such antibiotics.
Example~ of antibiotics which may be employed include, but
are not limited to, a tetracycline(s); a p~eudomonic acld;
viomycin; a cephalo~porin; benzoyl peroxlde; ~thambutol;
isoniazid; ethionamide; a 3ulfonamide; trlmethoprim;
nitroimidazole~; an antimalariral agent; cyclo~erine; a penem
antibiotic; an aminoglyco~ide; a hydrophobic antibiotic; a
pen~cillin; a monobactam; a 50-S ribo~ome inhibitor; and an
antibiotic having a large lipid-like lactone ring.
W093/07892 2 ~ 2 o 3 3 7 PCT/~S92/088~3
Tetracyclines which may be employed include, but are not
limited to, tetracycline, doxycycline, oxytetracycline, as well
as those isolated from new species of MicromonosPora and
Actimonadura, such as, for example, Sch-36969, Sch-332~6, and
Sch-34164 (Schering Plough).
In one embodiment, thee pseudomonic acid iE mupirocin
(pseudomonic acid A~.
Cephalosporins which may be employed include, but are not
limited to cefotaxime; cephalothin; cephalexin; cefazolin;
cephradine; cephapir~n; cefotetan; cefamandole; ceftazidime;
cefoxitin; cefuroxime; cefoperazone; ceftriaxone; ceftizoxime;
ceforanide; cefonicid; cefaclor; 3' quaternary ammonium
cephalosporins; broad-spectrum cephalo~porins such as MI-4646,
MI-4648, and MI-4659 (Mochida); YM-13115 (Yamanouchi); DQ-2522
and DQ-25~6 (Daiichi); B0-1232 and 80 ~236 (Banyu~; CL-118523
(American Cyanamid), and moxalactam. In o~e embodiment, the
cephalosporin is cefotaxime.
Sulfonamides which may be employed include, but are not
limit~d to, sulfanilamide; sulfadiazine; sulfabenzamide;
sulfamethoxazole; sulfisoxazole; sulfamethoxazole sulfathalidine;
sulfacetamide; sulfacytine; sulfadoxime; sulfamerazine;
sulfamethazine; sulsulfamethizole; sulfapyridine; sulfasalazi~e;
sulfathiazole; and sulfapyrazone.
Anti-malarial agents which may be employed include, but are
not limited to, pyrimethamine; primaguine; chloroguine; quinine,
and quinidine.
Penem antibiotics include, but are not limite~ to, imipenem,
car~apenems, and 2-(n-azolyl) alkyl-substituted penem~.
Aminoglycoside antibiotics include, but are not limited to,
tobramycin, kanamycin, amikacin, the gentamicins (e.g.,
gentamicin C1, gentamicin C2, gentamicin C1a), netilmicin,
kanamycin, neomycin, streptomycin, and deri~atives and analo~ues
thereof. The preferred aminoglycosides are tobramycin and the
W093/07892 PCT/US~/08823
2l2a3~7
gentamicins. The aminoglycosides, and the bacitracins
hereinafter described, tend to be hydrophilic and water-soluble.
Penicillins which may be employed in accordance with the
present invention include, but are not limited to benzyl
penicillin, ampicillin, methi~illin ~dimethoxyphenyl penicillin),
ticaricillin, penicillin V (phenoxymethyl penicillin), oxacillin,
cloxacillin, dicloxacillin, flucloxacilli~, amoxicillin, and
amidinocillin. Preferred penicillins which may be employed are
benzyl penicillin and ampicillin. A preferred monobactam which
may be employed is aztreonam.
As representative examples of hydrophobic antibiotics which
may be used in the present invention, there may be mentioned
macrolides such as erythromycin, roxythromycin, clarithromycin,
etc.; 9-N-alkyl derivatives of erythromycin; mid~camycin acetate;
azithromycin; flurithromycin; rifabutin rokitamycin; a
6-0-methyl ery~hromycin A known as TE-031 (Taisho); rifapêntine;
benzypipera~inyl rifamycins such as CGP-7040, C~P-5909,
CGP-279353 5Ciba-Geigy); an erythromy~in A derivative with a
cyclic carbamate fu~ed to the Cl1/C12 po~ition of a macrolide
ring known as A-62~14 (Abbott); AC-7230 ~Toyo Jozo);
benzoxazinorifamycin; difficidin; dirithromycin; a
3-N-piperdinomethylzaino methyl rifamy in ~V known as FCE-222~50
(Farmitalia); M-119-a (Kirin Brewery); a
6-0-met~yl-1-4"-0-carbamoyl erythromycin known as A-6307~
(Abbott); 3-formylrifamycin S~J-hydrazones with diazabicycloalkyl
side chains such as CGP-27S~7 and CGP-2986 ~Ciba-Geigy~; and
16-membered macrolides having a 3~0-alpha-~-cladi~o~yl moiety,
~uch as 3-0-alpha-L-cladinosyldeepoxy rosaramicin; tylosins and
acyl demycinosyl tylosins.
In addition to the macrolides hereinabove described,
rifamycin, rifampin, carbenicillin, and nafcillin may be employed
as well.
Other antibiotics which may be used (whether or not
hydrophobic) are antibiotics which are S0-S ribosome inhibitors
WO 93/07892 2 ~ 2 0 3 3 7 PCT/US92/08823
such as lincomycin; clindamycin; and chloramphenirol; etc.;
antibiotics which have a large lipid like lactone ring, such as
mystatin; pimaricin, etc.
The preferred hydrophobic antibiotics are the macrolides and
in particular erythromycin and derivatives and analogues thereof.
Peptide antibiotics which are not ion channel-forming
peptides or proteins which may be employed include, but are not
limited to, a bacitracin; gramacidin S; polymyxin; vancomycin;
teichoplanin; and capreomycin; and derivatives and analogues
thereof.
The biologically active amphiphilic peptideq employed in the
present invention are generally water soluble to a concentration
of at least 20 mg/ml at neutral pH in water. In addition, the
structure of such peptide provides for flexibility of the peptide
molecule. When the peptide i~ placed in wa~er, it does not
assume an amphiphilic structure. When the p~ptide encoun~ers an
oily surface or membrane, the peptide chain fold~ upon itself
into a rod-like or alpha-helical structure.
In general, such peptides ha~e at lea~t 11 amino acids, and
preferably at least 20 amino acid~. In most ca~es, such peptides
do not have in excess of 50 amino acids.
In general, the biolo~ically active peptides or proteins~
employed in the present in~ention are ion cha~nel-forming
peptides or proteins. An ion channel-orming peptide or protein
or ionophore is a peptide or protein which increa~es the
permeability for ions across a natural or synthetic lipid
membrane. B. Christensen et al. PNAS ~ol. 85 P. 5072-76 (July,
1988~ describes methodology which indicates whether or not a
pep~ide or; protein has ion channel~forming prop~rties and is
therefore an ionophore. As used herein an ion channel-forming
peptide or protein is a peptide or protein which has ion
channel-forming properties as determined by the method of
Christensen et al.
WO 93/0?892 PCT/US92/08823
212~3~7
An amphiphilic peptide or protein is a peptide which
includes both hydrophobic and hydrophilic peptide regions.
Although the biologically active amphiphilic (amphipathic)
ion channel-forming peptides or proteins are capable of forming
ion channels, the ability of such peptides or proteins and the
above-mentioned a~,tibiotics to potentiate each other is not
necessarily dependent upon the antibiotic crossing a membrane
through such channels. Thus, although the ability to form ion
channels may be a characteristic of a type of peptide or protein
used in the invention, the invention is not limited to the
formation and/or use of such channels as part of the mechani~m
for the peptide or protein potentiating the antibiotic or vice
versa. Similarly, although Applicant believes that such peptides
or proteins interact with the membrane of bacterial cells and
such interaction is the mechanism by which the antib.~.otic
potentiates the peptide or protein and vice versa, the pre~sent
invention is not limited to such a mechanism.
The term "potentiate", as used herein, means either that the
biologically active amphiphilic peptide or protcin is e~fective
in increasing the biological activity of the above-mentioned
antibiotics against a target cell so thereby the antibiotic may
be employed in an amount lower than that which would be requi~ed
for preven~ing, destroying or inhibiting growth of a tar~et cell,
and/or that the peptide or protein may be employed in~an amount
lower than that which would be required for preventing,
destroying, or inhibiting growth of a target c~llo
The administration of the biologically active amphiphilic
peptides or proteins and antibiotic to a target cell may be
direct administration to the cell or ~ystemic or topical
administration to a host which includes the target cell, in order
to prevent, destroy, or inhibit the growth of a target cell.
Target cells whose growth may be prevented, inhibited, or
destroyed by the administration of the biolog~cally active
W093/07892 2 ~ ~ 0 3 3 7 PCT/US92/OW23
amphiphilic peptide or protein and antibiotic include
Gram-positive and Gram-negative bacteria.
For example, erythromycin, when employed without the
above-mentioned peptides, is effective only against Gram-positive
organisms. Applicants have found unexpec~edly that erythromycin,
when employed in combination with the above-mentioned peptides or
proteins, is potentiated such that it becomes biolcgically
effective against Gram-negative bacteria. Moreover, the
erythromycin may be employed against Gram-positive bacteria in
amounts lower than tho~e normally used. Furthermore, such a
res~lt can be achieved by using peptide or protein amounts lower
tha~ those normally used.
The peptides or proteins employed in the present invention
are capable of interacting selectively with membranes of
bacteria.
In general, the peptide or protein is employed to provide
peptide dosages of from 1 mg to 500 mg per kilogram of host
weight, when administered systemically. When administered
topically, the peptide or protein is used in a concentration of
from 0.1% to 10%.
The antibiotic, such as those hereinabove described, or
derivatives or analogues thexeof, when used topically, is
generally employed in a concentration of about 0.1% to about 10%.
When used systemically, the anti~iotic or derivative or analogue
thereof is generally employed in an amount of from O.lmg to about
45mg per kg of host weight per day.
The use of a combination of peptide or protein and an
antibiotic such as those hereina~ove described, or derivatives or
analogues thereof in a~cordance with the present invention is
effective as an antibiotic, and may be employed to inhibit,
prevent or destroy the growth or proliferation of microbes, such
as bacteria.
The compositions have a broad range of poterlt antibiotic
activity against a plurality of microorganisms, including
WO ~3/07892 PCT/US92/08823
212~33~ ~
Gram-positive and Gram-negative bacteria. Such compositions may
be employed for treating or controlling microbial infection
caused by organisms which are sensitive to such composition. The
treatment may comprise administering to a host organism or
tissues acceptable to or affiliated with a microbial infection an
anti-microbial amount of such peptide or protein and an
antibiotic.
The compositions may also be used as preservatives or
sterilants for materials susceptible to microbial contamination.
In accordance with a preferred embodiment, the peptide used
in conjunction with an antibiotic such as those hereinabo~e
described, or derivatives or analo~es t~ercof i8 a basic
(positively charged) polypeptide having at least sixteen amino
acids wherein the polypeptide includes at least eight hydrophobic
amino acids a~d at least eight hydrophilic amino acids. Still
more particularly, the hydrophobic amino acids are in groups o
two adjacent amino acids, and each group of two hydrophobic amino
acids i8 spaced from another group of two hydrophobic amino acids
by at least one amino acid other than a hydrophobic amino acid
(preferably at least two amino acids) and generally by no greater
than four amino acids, and the amino acids ~etwe~n pairs of
hydrophobic amino acids may or may not be hydrophilic.
The hydrophilic amino acids are generally also in groups of
two adja~ent amino acids in which at lea~t one of ~he~two amino
acids is a basic hydrophilic amino acid, with ~uch groups of two
hydrophilic amino acids being ~paced from each other ~y at least
one amino acid other than a hydrophilic amino acid (preferably at
least two amino acids) and ~enerally no greater than four amino
acids, and the amino acids between pairs of hydrophilic amino
~cids may or may not be hydropho~ic.
In accordance with a particularly preferred embodiment, the
polypeptide comprises a chain of at least four group3 of amino
acids, with each group consisting of four amino acids. Two of
the four amino acids in each group are hydrophobic amino acids,
WO ~3/07892 2 1 2 0 3 3 7 PCT/US92/08823
and two of the four amino acids in each group are hydrophilic,
with at least one of the hydrophilic amino acids in each group
being a basic hydrophilic amino acid and the other beinq a basic
or neutral hydrophilic amino acid.
The hydrophobic amino acids may be selected from the class
consisting of Ala, Cys, Phe, Gly, Ile, Leu, Met, Val, Trp, Tyr,
norleucine (Nle), norvaline (Nva), and cyclohexylalanine (Cha).
The neutral hydrophilic amino acids may be selected from the
class consisting of Asn, Gln, Ser, snd Thr. The ba~ic
hydrophilic amino acids may be selected from the class con~isting
of Lys, Arg, and His, Orn, homoarginine (Har), 2,
4-diaminobutyric acid (Dbu), and p-aminophenylalanine.
Each of the groups of four amino acids may be of the
seguence ABCD, BCDA, CDA~, or DABC, wherein A and 8 are each
hydrophobic amino acids and may be the same or different, one of
C or D is a basic hydrophilic amino acid, and the other of C or D
is a basic or neutral hydrophilic amino acid and may be the same
or different. In a preferred embodiment, the polypeptide chain
may comprise 5 or 6 groups of one or more of the~e ~equences. In
each group, each of A, B, C and D may be the Jame in ~ome or all
of the groups or may be different in some cr all of the groups.
The polypeptide chain preferably has at least 16 amino
acids, and ~o greater than 50 amino acids. It is to be
understood, however, that the polypeptide does not have to
consist entirely of the groups described abo~e. The polypeptide
may have amino acids extendirlg rom either or both end~ of the
noted groups fo~ming the polypeptide chain and/or there may be
amino acids between one or more of ~he at le~st four groups and
still remain within the scope of the invention.
The groups of amino acids may be repe~t1 ng groups of amino
acids, or the amino acids in the various groups may vary provided
that in each group of the at least four groups of amino acids
there are two hydrophobic and two hydrophilic amino acids as
hereinabove noted.
W093/07892 PCT/U~92/~8X23
212~3~7 ~
-10- , ,,
Thus, in a preferred embodiment, the biologically active
polypeptide comprises a chain including at least four groups of ,~
amino acids, each containing four amino acid~. Two of the four
amino acids in each group are hydrophobic, at lea~t one amino
acid is basic hydrophilic, and the remaining one i8 basic or
neutral hydrophilic, with the polypeptide chain preferably having
at least 20 amino acids but no greater than 50 amino acids.
In one em~odiment, each of the at least four groups of àmino
acids which are in the peptide chain is of the seguence A-B-C-D,
B-C-D-A, C-D-A-B or D-A-8-C wherein A and ~ are hydrophobic ~:
amino acids, one of C or D is basic hydrophilic amino acid, and
the other of C or D is basic or neutral hydrophilic amino acid. :~
The resulting polypeptide chain, therefore, may have one of the
following sequences:
(X1)a(A~B~C~D)n(Yl)b
(X2 ) a(B~C~D~A)n(Y2 )~
(X3)a~C-D-A-B)n(Y3)b
(x4)a(D-A-~-c)n(y4)b
wherein X1 is D; C-D- or B-C-D-, Y1 is -A or :~
-A-B or -A-B-C -:
X2 is A-, D-A- or C-D-A-
Y2 is -B, -B-C or B-C-D
X3is B-, A-B-, D-A-B-
Y3 is -C, C-D, -C-D-A .
X~is C-, B-C-, A-B-C-
Y4 is -D, -D-A, -~-A-~
a i5 0 or 1; b is 0 or 1
and n is at least 4.
It is to be understood that the peptide chain may include
amino acids between the hereinabo~e noted groups of four amino
acids provided that the spacing between such groups and the
charge on the amino acids does not change the characteristics of
WO93J07892 2 1 2 Q 3 3 7 PCT/US~2/08823
the peptide chain which provide amphiphilicity and a positive
charge and do not adversely affect the folding characteristics of
the chain to that which is significantly different from one in
which the hereinabove noted group of four amino acids are not
spaced from each other~
As representative examples of peptides in accordance with
the present invention, there may be mentioned.
I Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-
Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys (SEQ ID
NO~
II Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-
Lys-Ala-Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-Phe-
Ser-Lys. (SEQ ID NO:2)
III Phe-Ser-Lys-Al~-Phe-Ser-
Lys-Ala-Phe-Ser-Lys-Ala-
Phe-Ser-Lys-Ala- (SEQ ID NO:3).
IV Ser-Lys-Ala-Phe-Ser-Lys-Ala- -
Phe-Ser-Lys-Ala-Phe-Ser-Lys-Ala-
~he-Ser-Lys-Ala-Phe- (SEQ ID NO:4~ -
V Lys-Ala-Phe-Ser-Ly~-Ala-Phe-Ser-~ys-Ala-Phe-Ser-
Lys-Ala-Phe-Ser (SEQ ID NO:5)
The peptide may have amino acids extending from either en~d
of the chain. For example, the chains may have a Ser-Lys
seguence before the "Ala" end, and/or an Ala-Phe seguence after
the "Lys" end. Other amino acid seguences may al~o be attached
to the "Ala" and/or the "Lys" end.
Simil~rly, in any polypeptide chain having at least four ~`
groups of amino acids of the se~ence as described above, the
chain may have, for example, a C-D seguence before the first
A-B-C-D group. Also other amino acid sequences may be attached
to the "A" and/or the "Dl' end of one of these polypeptide chains
Also there may be amino acids in the chain which ~pace one or
more groups of the hereinabove noted four amino acids from each
other.
WO 93/07892 P~T/US92/08823
2120~7 ::
-12-
The peptides may ~e produced by known techni~ues and
obtained in substantially pure form. For example, the peptides
may be synthesized on an automatic chemical peptide synthesizer.
Journal of the American Chemical Society, Vol. 85 Pages
2149-54(1963). It is also possi~le to produce ~uch peptides by
recombinant ~NA technology using genetic engineeri~g techni~ues.
Methods in Enz~molo~Y, Vol. 68: "Recombinant DNA" (Wu, et al.)
Academic Press, New York (1979); and Methods in Enz~oloaY~ Vol.
18~: "Gene Expression Technology" (~.V. Goeddel, et al.) Academic
Press, New York (l990).
In accordance with another preferred embodiment, the peptide
employed in conjunction with an antibiotic such as those
hereinabove described, or derivatives or analogues thereof may be
a magainin peptide.
A magainin peptide is either a magaini~ such as Magai~in I,
II or III or an analogue or derivative thereof. The magainin
peptides may inc'ude the following basic peptide structure X12
Rll R11 R12 ~13 Rll R14 R12 Rll R14
R12 ~ Rll R14a- (R15 ~n-R14a-R14 ~~
wherein R11 is a hydrophobic amino acid, R12 is a basic
hydrcphilic amino acid; Rl3 is a hydrophobi~, neutral
hydrophilic, or basic hydrophilic amino acid; Rl4 and R14a are~ -
hydropho~ic or basic hydrophilic amino acids, Rl5 ~ 5 ~l~tamic
acid or aspartic acid, or a hydrophohic or basic hydrophilic
amino acid, and n is 0 or 1. In a preferred ~mbodiment, ~13 is a
hydrophobic or net~tral hydrophilic amino acid, Rl4a is a
hydropho~ic amino acid, and R15 is glutamic acid or aspartic
acid.
Thus, for example, a magaini~ peptide may inrlude the
following structure:
Y12 X12'
where X12 is the here-~nabove described basic peptide
structure and Y12 is
(i) R12; .
WO 93/07892 PCT/US92/08823
2120~37
-13-
(ii) R14 - R12;
(iii) Rll-R14 -R12; or
(iv) ~14 Rll R14a R12
Rll, R12, R14, and R14a are as previously defined.
A magainin peptide may al80 have the following structure:
-X12-Z12,
wherein X12 is as previously defined and Z12 i5:
(i) R16 where R16 is a basic hydrophilic amino a~id or
asparagine or glutamine; or ~:
(ii) R16-R17 where R17 is a neutral hydrophilic amino
acid, a hydrophobic amino acid, or a basic hydrophilic amino
acid. Preferably, R17 is a neutral hydrophilic amino acid.
A magainin peptide may al50 have the following structure
(Y12)a X12 (Z12)b
where Xl7, Y1 , and Z12 are as previously defined, and a is
or 1 and b is 0 or 1. -:~
The magainin peptides may also include the f~llowing basic -~
peptide structure X13:
14 ~11 R14a R12 Rll~R~ 12~R13~~ R14~
12 R11 Rll ~12-'
Rll' R12' R13, R14, and R14a are amino acids as
here~na~ove describ~d.
The magainin peptide may also include the fo~lowing ~:
structure X13-Z~3; wherein X13 i8 the hereinaboYe described basic
peptide structure and Z13 is
(Rll)n-(Rll)n~ l)n-(Rl4a)n (Rls)n ( 14a)n
(R14) ~~R16)n-(R17~n-~ whe~ein R11~ R14~ R14a' R15' 16~
R17 are amino acids as hereinabo~e described, and n is O or 1,
and each n may be the same or different.
The magainin peptides generally include at least fourteen
amino acids and may include up to forty amino acids. A magainin
peptide preferably has 22 or 23 amino acids. A~cordingly, the
hereinabove described basic peptide structures of a magainin
W093/07892 PCT/US92/08823
2120337
peptide may incl~de additional amino acids at the amino end or at
the carboxyl end, or at ~oth ends. ..
As representative examples of such magainin peptides, there
may be mentioned peptides having the following primary sequences
as given in the accompanying sequence listing, as well as
appropriate analogues and derivatives thereof:
(a) (NH2) (SEQ ID NO:6) (OH) or (NH2)
(Magainin I)
(b) (NH2) (SEQ ID NO:7) (O~) or (NH2) :~
(Magainin II)
(c) (NH2) ~SEQ ID NO:8) (OH) or (NH2)
(Magainin III)
The following are examples of peptide derivatives or analo~s .
t~f the basi~ structure:
(d) ~NH~) (SEQ ID NO:9) (OH) or (NH2, .-
~e) (NH23 (SEQ ID NO:lO) (9H) or (NH23
(f) (NH2) ~SEQ ID NO:ll) (OH) or (NH2) .-
Magainin peptides are described in Proc. Natl. Acad Sc~. ~
Vol. 84, pp. 5449-53 (Aug. 1987). The term "magainin peptides" ~i
as used herein refers to the basic magainin stru~ture as well as
derivatives and analogs thereof, including but not limited to the
representative derivatives or analogs.
In ac~ordance with a further embodiment, the peptide
employed in conjunctio~ with an antibioti~ such as bac~tracin,
tobramycin or gentamicin or derivatives or analogues thereof may
be a pGr~a peptide or an XPF peptide.
A PGLa peptide is either PGLa or an analogue or derivative
thereof. The PGLa peptides preferably include the following
basic peptide ~tru~ture X14:
~ 17 ~12 ~ 14 ~
Rll R14 ~12 Rll Rll R12 11
Rll Rll ~12
where R~ 12~ Rl4, and R17 are as previously defined.
wo g3/~,892 2 1 2 o 3 3 7 Pcr/US92/08823
-15-
The PGLa peptides generally include at least seventeen amino
acids and may include as many as forty amino acids. Accordingly,
the hereinabove described basic peptide stru~ture for a P~La
peptide may include additional amino acids at the amino end or at
the carboxyl end or at both the amino and carboxyl end.
Thus, for example, a PGLa peptide may have the following
struct~re:
Y14 X14
where X14 is as previously defined and
Y14 iS
(i) Rll;
( ii ) R14 Rll
where Rll is as pre~iously defined.
For example, a PGLa like peptide may also have the following
structure:
14 14'
where X14 is as previously defined; and Z14 is:
~i) Rll; or
~ ii ) 11 11 " '
where Rll is as previously defined.
A PGLa peptide may als~ have the following structure: !'`
(Y14)a X14 (Z14)~
where X14; Y14 and Z14 are as previously deined,~ a is O or
1 and b is O or 1.
An XPF peptide is either XPF or an analogue or derivative
thereof. The XPF peptides preferably include the following ~asic
peptide structur* X16:
Rll R17 R12~ 14-R18-R17-
Rl1 R14-R12-Rl1-;Rl1-R12-
Rll-Rll- Rll-R12-~15-Rll--~
Rl1, R12, R14, R15 and ~17 are as previously defined
~nd R18 is glutamine or asparagine.
W093/07892 PCT/US92/08X23
2120337
The XPF peptides generally include at least nineteen amino
acids and may include up to forty amino acids. Accordingly, the
hereinabove described basic peptide structure of XPF may include ~:
additional amino acids at the amino end, or at the carboxyl end
or at both the amino and carboxyl ends. :
Thus, for example, an XPF peptide may include the following -~-
structure:
Y16 X16 '
where X16 is as previously defined and Y16 is
(i) Rll or
(ii) 14
where Rll and R14 are is as previously defined. ~
An XPF peptide may include the following structure: ~:
-X16-Z16,
where X16 is as previously defined and Z16 is
~i) R11; or
(ii) R11 R18; or
(iii) 11 Rlg-Proline; or
( iv ) Rll-R18-Prline R12
An XPF peptide may also have the following struc~ure: :
(Y16~a 16( 1~)b -~.
where X16, Y16 and ~16 are as previously defined: a is 0 or
1 and b is 0 or 1.~
Preferred are XPF or PGLa peptides, which are characteri~ed
by the following primary amino acid sequence as given in the
accompanying sequenc~ listing:
PGLa : (SEQ ID N0:12) (NH2)
XPF : (SEQ ID N0:13)
A review of XPF and PGLa can be found in Hoffman et al, EMB0
J. 2:711-714; 19~3; Andreu et al, J B ochem. 149:531-535, 1985;
Gibson et al J. Biol. Chem. 261:5341 5349, 1986; and Giovannini
et al, Biochem J. 243:113-120, 1987. ~
W093/07X92 2 1 2 ~ 3 3 7 PCT/US92/08823
In accordance with yet another embodiment, the peptide
employed in conjunction with an antibiotic such as those
hereinabove described, or derivatives or analogues thereof may be
a CPF peptide or appropriate analogue or derviative thereof.
A basic CPF peptide structure as well as analogues and
derivatives thereof are herein sometimes referred to collectively
as CPF peptides.
The CPF peptide is preferably one which includes the
following peptide structure X30: `
21 ~21 R22~R22~~21~R21~R23~R2~
-R21-R21-~'23-~?'21-~21-R24-R25-R21-
wherein R21 is a hydrophobic amino acid;
R22 is a hydrophobic amino acid or a basic hydrophilic amino
acid;
R23 is a basic hydrophilic amino acid; and
R24 is a hydrophobic or neutral hydrophilic amino acId; and ~:
R25 is a basic or neutral hydrophilic amino a~id.
The hereinabove basic structure ic hereinafter symbolically
indicated as X30.
The hydrophobic amino acids may be Ala, Cys, Phe, Gly, Ile,
Leu, Met, Val, Trp, and Tyr.
The neutral hydrophilic amino acids may be Asn, Gln, Ser,
and Thr.
The basic hydrophilic amino acids may be Ly~, Arg, and ~is,
Orn, homoarginine (Har), 2, 4-diaminobutyri~ acid lDbu), and
p-aminophenylalanine.
The CPF peptide may include only the hereinabove noted amino
acids or may include additional amino acids at the amino end or
car~oxyl end or both the amino and car~oxyl ~nd. In g~neral, the
peptide does not include more than 40 amino acid3.
The CPF peptides including the above ba~ic peptide structure
may have from 1 to 4 additional amino acids at the amino end.
Accordingly, such preferred peptides may be represented by the
structural formula:
~V~ 93/07X92 PCT/US92/08823
2120337
-18-
Y30-X30-
wherein X30 is the hereinabove described basic peptide -~
structure and Y30 is
(i) R25-' or
(ii) R22 R25; or
(iii) R2l-R22-R25 or
(iv) 22 R2l R22~R25; preferably
GlyCine -3~21-R22 R25
wherein R2~, R22, and R25 are as previously defined.
The carboxyl end of the basic peptide structure may also --
have additional amino acids which may r~nge from l to 13
additional amino acids. ~-~
In a preferred em~odiment, the basic structure may have from
1 to 7 additional amino acids at the carboxyl end, which may be
represented as follows~
-X30-Z30 wherein
X30 is the hereinabove defined basic peptide structure and
Z30 is ~:
(i) R21 ' ~:
( ii ) R21 R21
Iiii) R2l R21 ~24;
(iv) R2l R2l R2~ R~4;
(~) R2l i~21 ~24 ~24 R26;
(vi) R2l R2l R24~R24~~26-Gln; or
~vii ) R21 R21 i~24-~24-R26-G~ Gln~
wherein R21 and R24 are as previously defined, and R26 is
proline or a hydrophobic amino acid.
Preferred peptides may be represented by the following
structural ormula:
(Y30)a 30 (Z30)b
wherein X30, Y30 and Z30 are as previously defined and a is
0 or l a~d b is 0 or l.
WO ~3/0789Z 2 1 2 o 3 3 7 PCT/USg2/-88
- 19- `~ .
~ .
Representative examples of CPF peptides which are useful in
the present invention have been described in the literature and
comprise the following sequences as given in the accompanying
sequence listing:
(SEQ ID N0:14)
(SEQ ID N0:15)
(SEQ ID N0:16) `~
(SEQ ID N0:17)
(SEQ ID N0:18)
(SEQ ID N0:19) `
(SEQ ID N0:20)
(SEQ ID N0:21)
(SEQ ID N0:22)
(SEQ ID N0:23)
(SEQ ID N~:24)
(SEQ ID N0:25)
(SEQ ID NO:26)
A review of the CPF peptides can b- found in Richter, K.,
Egger, R., and Kreil (1986) J. Biol. Chem. 261, 3676-3680;
Wakabayashi, T. Kato, H., and Tachibaba, S. (198~) Nucleic Acids '`5'~'
Research 13, 1817-1828; Gibson, B.W., Poulter, L., Williams,
D.H., and Maggio, J.E. (1986) J. Biol. Chem. 261, 5341-5349.
CPF peptides which may be employed in the present invention
are represented by the following (single letter amino acid ~ode):
G12S3LG4ALKA5LKIG678LGG9(10)QQ
Where: -~
1 = F, L
- 2 = G, A
3 = F, L -
4 = K, L
= A, G, T
6 = A, T
7 = H, N
8 = A, M, F, L
W093/07892 PCT/~S92/08823
212~337 ~
-20-
9 = A, S, T
= P, L
The numbered amino acids may be employed as described in any ~;~
combination to provide either a basic CPF peptide ~tructure or an
analogue or derivative~ The term CPF peptide includes the ba~ic
peptide structure as well as analogues or derivatives thereof. ~
In accordance with yet another embodiment, the peptide may ~ :
include one of the following basic structures X31 through X37
wherein: :
X31 is lR3l-R32-~32-R33-R3l-~32~R32l~n; -
X32 is [R32 R32 ~33 R31~R32~R32~R313~n;
X33 is lR32 R33 R31 ~32 ~32 ~31 ~32] n;
X34 is lR33-~31-R32-R32-R31-R32-R32~-n;
X35 is -1R31-R32-R32-R31 R32 R32 33 n
X36 is IR3~-~32-R3l-R32-R32-R33-R3l~- ; and
37 lR32 R31 R32-R32-~33-~31-R321-n;
wherein R31 is a basic hydrophilic amino ac~d, R32 i~ a
hydropho~ic amino acid, R33 is a neutral hydrophilic or
hydrophobic amino acid, and n is from 2 to S.
The basic hydrophilic amino acids may be selected from the
class consisting of Lys, Arg, ~is, Orn, homoarginine (Har),
2,4-diamino-butyric acid (Dbu), and p-aminophenylalanine.
The hydrophobi~ amino acids may be selected ~rom the class
consisting of Ala, Cys, Phe, Gly, Ile, Leu, Net, Val, Trp and
Tyr,norleucine ~Nle), norvaline (Nval), and cyclohexylalanine
(Cha).
The neutral hydrophilic smino acids may be selected from the
class consisting of Asn, Gln, Ser and Thr.
In accordance with one embodiment, when ~he peptide includes
the structure X31, the peptide may include the following
structure:
Y31-X31, wherein X31 is as hereina~ove described, and Y
is:
(i) ~32; `
WO 93/07892 2 1 2 ~ ~ 3 7 PCT/US92/08823 ~
-21- ~:
(ii) R32-R3Z;
(iii) R31-R32 R32;
(iV) R33-R31-R32 R32;
(v) R32-R33-R3l R32 32
( ) R32 ~32 R33 R3l-R32-R32~ wherein R31, R32, and R33 are
as hereinabove described. -:
In accordance with another em~odiment, when the peptide ~::
includes the structure X31, the peptide may include the following
structure:
X31-Z3l, wherein X3l is as hereinabove described, and Z31
is:
(i ~ R31;
(ii) R31 R32;
(iii) R31-~32 R32;
(iv) R31-R32 R32 33
(v) R31 R32-R32-R33-R31; or
~ vi) R3l-R3~-R32-R33 ~31 3Z
In accordance with yet another embodiment, the peptide may ~:
include the following structure: -
(Y3l)a-x3l-(z3l)b~ wherein Y31 and Z31 are as pre~iously
defined, a is 0 or 1, and b is 0 or l.
When the peptide includes the structure X32, the peptideCmay
include the following structure: ;
Y32 ~ X32, wherein X32 is as hereinabo~e described, and Y32
i s :
(i) ~31; :~
(ii~ R32-R31;
(iii) R32-~32 ~31
(iv) R3l~3~-R32 ~31'
R33 R31 R32 R32 31;
R32-R33-~31 R32 R32 R31-
In another embodiment, when the peptide includes the
structure X32, the peptide may include the following structure: :
WO 93/07892 PCT/US92/08823
2l 20337
-22~
X32 ~ Z32~ wherein X32 is as hereinabove described, and Z32
i s :
( ) R32;
(ii) R32-R32;
(iii) R32-R32 R33;
(iv) R32-R32-R33 ~31;
(v) R3~-R32-~33-R31-R32; or
(vi) R32-R32-~33 R31 ~32 R32-
In accordance with yet another embodiment, the peptide may :-
include the following structure:
(Y32)a X32 (Z3Z)b' wherein Y3~ and Z32 are as previously
defined, a is 0 or 1, and b is 0 or 1.
In accordance with another embodiment, when the peptide
includes the structure X33, the peptide may include the following
structure:
Y33 - X33 wherein ~33 is as hereinabove described, and Y33
is:
(i) R32;
(ii3 R3i R32;
(iii) R32-~31 R32;
(iv) R32-~32-}?31 R32;
(v) ~31 ~32 ~32 R31 R32; or
t ) 33 ~31 R32 R32 R31 R32~ wherein R31, R32, and R33 are
as hereinabove d~scribed.
In accordance with another embodiment, when ~he peptide
includes the stru~ture X33, the peptide may include the following
structu~:
X33 - Z33 wherein X33 is as hereinabove described, ~nd Z33
is: .
(i) ~32;
( i) R32 R33;
(iii) R32-R33 ~31;
(iv) ~32-R33-R31 R32;
(v) R32-~33-~31-R32-R32; or
~ ~ ~ r ~ ~ ~ PCT/US92/08823
WO 93/07892
-~3-
(vi) R32-R33-R3l-R32 R32 31
In accordance with yet another embodiment, the peptide may
include the following structure: `
(Y33)a X33 (Z33)b~ wherein Y33 and Z33 are as previously
defined, a is 0 or l, and b is 0 or 1.
In accordance with yet another embodiment, when the peptide
includes the structure X34, the peptide may include the following
structure: ,
Y34 - X34, wherein X34 is as hereinabove described, and Y34
i s :
(i) R32;
(ii) R32 ~32;
(iii) R3l-~32 R32;
(iv) R3 Y2-R3l-R32 R32;
( ) R32 R32 R31 ~32 ~32; or -~
( ) R31 R32 ~32 ~31 R32-~32~ wherein R31, R32 and R33 are ~;
as hereinabove described.
In accordance with another embodiment, when the peptide -~
includes the structure X34, the peptide~may include the followin~
structure:
- X34-Z34, wherein X34 is as hereinabove described, and Z3
is:
(i) R33;
(ii) ~33-~31;
(iii) R33-R3l ~32;
(iv) R33-R3l-R32 R32;
( ) R33 ~31 R32 ~32-R31; or
(Vi ) R33-~.31; R32-R32R31-R32 '' '
In accordance with yet another embodiment, the peptide may
include the followiny structure:
(Y34)a X34 tZ34)b~ wherein X34 and Z34 are as previously
defined, a is 0 or l, and b is 0 or l.
W093/07892 PCT/US92/08823
2~2~337
-24-
In accordance with a further embodiment, when the peptide
includes the structure X35, the peptide may include the following
structure:
Y35-X3~, wherein X35 is as hereinabove described, and Y
is:
(i) R33;
(ii) R32-~33;
(iii) R32~R32~R33;
(i~) R3l-R32 R32 R33;
(v) R32-R3l-R3~ R32 33;
( ) R32 R32 ~31 R32 R32-R33~ wherein R31, ~32~ and R33 are
as hereinabove described.
In accordance with another embodiment, when the peptide
includes the structure X35, the peptide may include the following
structure:
X35 - Z35 wherein X35 is as hereinabove described, and Z
is:
(i ) R31;
(ii) R31'R32;
(iii) R31-~32 R32;
(iV) R31-~32-R32 R31;
(v~ 31 R32 R32~R31~R32; or
(vi) R3l-~32-~32-R31 R32 32
In accordance with yet another embodîment, the peptide may
include the followin~ structure:
~ Y35)a- X35 ~Z35)~, wherein X35 and Z35 are as previously
defined, a is 0 or 1, ~nd b is 0 or l.
In accord~nce with a further embodiment, when the peptide
includes the structure X36, the peptide may include the following
structure:
Y3~ ~ X36 wherein X36 is as hereinabove described, and Y36
is:
(i) ~31;
(ii) R33-R31;
W093~07892 - 2 1 2 0 3 3 7 PCT/US92/08823
-25
(iii) R32~R33 R31;
(iv) R32~~32~R33 R31;
(v) R31 R32-R32-R33-R31; or
( ) R32 R31 R32 ~32-R33-R31~ wherein R31, R32, and R33 are
as hereinabove described.
In accordance with another ~m~odiment, when the pe~tide :~
includes the structure X36, the peptide may include the following
structure:
X36-Z36, wherein X36 is as hereina~ove described, and Z
i s :
(i) R32; ~'
(ii) R32-~32;
(iii ) R32-~32 R31;
(iv) R32-R32 R31 32
(v) R~2-R32-R3l-R32 R32;
(vi) R32-R32-R31 ~32 32 33
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y ) - X (Z 6)~ wherein Y36 and Z36 are as previ~uslY
defined, a is O or 1, and b is 0 or 1.
I~ accordance with one embodiment, when the peptide includes -~
the structure X37, the peptide may includes the structure . ~:
Y37-X37, wherein X37 is as hereinabove de3cribed, and Y37 is:
~i) R3~,
(ii) R31-R32;
(iii) R33-~31 R32;
(iv) R32-~33-R31 R32;
(v) R -~ -~33-~31-R32; or
( ) 31 ~32 R32 R33 R31 R32, wherein R31, R32, and R33 are
as hereinabove described.
In accordance with a further embodiment, when the peptide
includes the structure X37, the peptide may include the follo,wing
structure:
W093/07892 PCT/US92/08823
21203~7 `
-26-
X37 - Z37 wherein X37 is as hereinabove described, and Z37 -
i s :
( ) 32;
(ii) R32-R31;
(iii~ R32-R31 R32;
(iv) R32-R31-R32 R32;
(v) R32 ~31 R32 32 33;
(vi) R32-R3l-R32 ~32 33 31
In accordance with yet another embodiment, the peptide may
include the following structure:
(Y37)a X37 (Z37)b' wherein Y37 and Z37 are a~ previously
defined, a is O or 1, and b is O or 1.
In a preferred embodiment, n is 3, and most preferably the
peptide is of one of the following structures as given in the
accompanying sequen~e listing:
(Lys Ile Ala Gly Lys Ile Ala)3 ~SEQ ID NO:27).
(Lys Ile Ala Lys Ile Ala Gly)3 (SEQ ID NO:28).
~Lys Ile Ala Gly Lys Ile Gly)3 (SEQ ID NO:29).
(~ys Leu Ala Gly Lys Leu Ala)3 (SEQ ID NO:30).
(Lys Phe Ala Gly Lys Phe Ala)3 (SEQ ID NO:31).
(Lys Ala Leu Ser Lys Ala ~eu~3 (SEQ ID NO:32). :~
(Lys Leu Leu Lyæ Ala Leu Gly)3 (SEQ ID NO:33). -~
(Lys Ala Ile Gly Ly~ Ala Ile)3 (5EQ ID NO:34~.
~Gly Ile Ala Lys Ile Ala Lys)3 (SEQ ID NO:35). -
(Lys Ile Ala ~ys Ile Phe Gly)3 (SEQ ID NO:36).
(Gly Ile Ala Ar~ Ile Ala Lys)3 ~SEQ ID NO:373.
(Lys Phe Ala Arg Ile Ala Gly)3 (SEQ ID NO:38).
(Gly Phe Ala Lys Ile Ala Lys)3 (SEQ ID NO:39).
(Lys Ile Ala Gly Orn Ile Ala)3 (SEQ ID NO:40).
(Lys Ile Ala Arg Ile Ala Gly)3 (SEQ ID NO:41).
(Orn Ile Ala Gly Lys Ile Ala)3 (SEQ ID NO:42).
(G~y Ile Ala Arg Ile Phe Lys)3 (SEQ ID NO:43).
(Lys Nle Ala Gly Lys Nle Ala)3 (SEQ ID NO:44).
(Lys Nle Ala Gly Lys Ile Ala)3 (SEQ ID NO:4~
W093/07892 2 1 ~ ~ 3 3 7 PCT/US92/08823
-27-
tLYs Ile Ala Gly Lys Nle Ala)3 (SEQ ID NO:46).
(Lys Nva Ala Gly Lys Nva Ala)3 (SEQ ID NO:47).
(Lys Nva Ala Gly Lys Ile Ala)3 (SEQ ID NO:48).
(Lys Leu Leu Ser Lys Leu Gly)3 (SEQ ID NO:49~.
(Lys Leu Leu Ser Lys Phe Gly)3 (S~Q ID NO:50).
(Lys Ile Ala Gly Lys Nva Ala)3 (SEQ ID NO:51).
(His Ile Ala Gly His Ile Ala)3 (SEQ ID NO:52).
(Ala Gly Lys Ile Ala Lys Ile)3 (SEQ ID NO:53).
(Ile Ala Lys Ile Ala Gly Lys)3 ~SEQ ID NO:54). -~
(Lys Ile Ala Gly Arg Il* Ala)3 ~SEQ ID NO:55).
(Arg Ile Ala Gly Arg Ile Ala)3 (SEQ ID NO:56).~:
(Lys Val Ala Gly Lys Ile Ala)3 (SEQ ID NO:57). -~
~Lys Ile Ala Gly Lys Val Al~)3 (SEQ ID NO:58).
(Ala Lys Ile Ala Gly Lys Ile)3 (SEQ ID NO:59).
(Orn Ile Ala Gly Orn Ile Ala)3 (SEQ ID NO:60).
(Lys Phe Ala Gly Lys Ile Ala)3 (SEQ ID NO:61).
(Lys Ile Ala Gly Lys Phe Ala)3 (SEQ ID NO:62).
(Lys Ch~ Ala Gly Lys Ile Ala)3 (SEQ ID NO:63).
(Lys Nle Ala Lys Ile ,~la Gly)3 (SEQ ID NO:643. :
(Arg Ile Ala Gly Lys Ile Ala)3 (5EQ ID NO:65~. :
(Har Ile Ala Gly Har Ile Ala)3 (SEQ 'D NO:663.
(X~a Ile Ala Gly Lys Ile Ala)3 (SEQ ID NO:67~.
(Lys Ile Ala Gly Xaa Ile Ala)3 (SEQ ID NO:68~.
In (SEQ ID NO:67) and ~SEQ ID NO:68~, Xaa i~
p-aminophenylalanine.
In accordance with another embodiment, the biologically
active amphiphilic peptide in~ludes the following basic structure
X40
R31-R32-R32-R33-R34-R32-R32-~?31-R32_R32_~.3~ R34-~.32_~,32,
wherein R31 ~ R32, and R33 are a~ hereinabove described, a
R34 is a basic hydrophilic or hydrophobic amino acid.
In accordance with one em~odiment, the peptide may includ
the following structure:
W~g~/~7~9~3 7 PCT/US92/08823
-28-
Y40-X40, wherein X40 is as herei~abo~e described, and Y40
i s :
(i) R32;
(ii) R32-R32;
(iii) R34-~32-R32;
(iv) R33-R34-R32-~32;
(v) 32 33 R34 ~32 R32;
(v) R32 R32 R33 R34 R32-R32, or
R ( ) R31 R32 R32 ~33-R34 ~32~R32,wherein R31, ~32~ R33 and
34 are as hereinabove de~cribed.
In accordance with another embodiment, the peptide may
include the following structure:
X40-Z40, wherein X40 is as hereinabove described and Z40 is:
(i ) R31;
(ii) R31 ~32;
(iii ) R31-R32 R32;
(iv) R31-R3~ R32 33
(v) R31-~32 ~32 33 34
(vi) R31 ~32 ~32 ~33 R34 R32; or `~
( ) ~31 R32 ~32 ~33~~34~R32~~32, wherein R31, R32, ~33,
and R34 are as hereinabove described.
In accordance with yet another embodim~nt the peptide may~ -:
include the following structure:
(Y40)a-x4o-(z4o)b~ wherein Y40 and ~40 are as previously
defined, a is O or 1, and b is O or 1. In a preferred
embodiment, ~he peptide has the following ~tructural formula 8S ~ :-
given in the accompanying sequence listing:
~ SEQ ID NO:69)
In another preferred em~odiment, the ~eptide has the
following structural formula as given in the accompanying
sPquence li sting:
(SEQ ID NO:70)
,
W093/07892 2 1 2 o ~ 3 7 PcT/us92/o8823
-29- ~:
In accordance with a further embodiment, the peptide has one ~.
of the one of the following structural formulae as given in the
accompanying sequence listing:
(SEQ I~ N0:71) ~
(SEQ ID N0:72) .
(SEQ ID N0:73) ~:
(SEQ ID N0:74)
(SEQ I~ N0:7S) ~
~SEQ ID N0:76) -~:
~SEQ ID N0:77)
~SEQ ID N0:78) .
(SEQ ID N0:79)
(SEQ ID N0:80)
(SEQ ID NO:B1)
~SEQ ,ID N0:82)
~SEQ ID NO:B3) --
~SEQ ID NO:B4)
~SEQ ID N0:85
In acc~rdance with another embodiment, the peptide may ~`
include the following stru~tural ~ormula: .~:
- ~Lys Ile Ala Lys Lys Ile Ala)-n, wherein n is from 2 to 5. .'
Preferably, n is 3, and the peptide has the following structu~al ~:`
formula:
~Lys Ile Ala Lys Lys Ile Ala)
(SEQ ID NO:B6
In accordanee with another embodiment, the peptide may be
selected from the group consisting of the followinq structural
formulae as given in the accompanying sequence listing~
~SEQ ID N0:87) ~ E~
(SEQ ID N0:88) ~;
~SEQ ID N0:89)
(SEQ IP N0:90)
In accordance with another em~odiment, the peptide may
includes the ollowing basic structure XS0: `
W093/07892 PCT/US92/0~823
212Q337
-30-
41 R42 R42 R41 42 42 41 41 42 41 R41
R41 is a hydrophobic amino acid, and R42 is a basi.c
hydrophilic or neutral hydrophilic ami~o acid.
In one embodiment, the peptide includes the basic structure
Y50-X50 wherein X50 is as hereinabove described and Y50 is: -
~ i) R41;
(ii) R42-R41; or
(iii) R42-R42-R41, wherein R41 and R42 are as hereinabo~e
described.
In one embodiment, R41 is leucine. In another embodiment,
R42 is lysine. Representative examples of peptides in acordance
with this aspect of the present invention include those having
the following structures:
(SEQ ID N0: 91)
(SEQ ID N0: g2)
(SE~ ID N0: 93)
~SEQ ID N0: 94)
In accordance with another em~odiment, the includes the
~ollowing basic structure X~2:
42 41 R42 R42 R41 R41 ~42-R42-R41-Rg2-R42, wherein R~l is
a hydropho~ic amino acid, and R2 is a basic hydrophilic or
neutral hydrophilic amino acid.
In o~e embodiment R41 is leucine. In another embodiment,
R42 is lysine-
In one embodiment, the peptide includes the basic structure
Y52-X52, wherein X52 is as hereinabove described, and YS2 is: ~-
(i) R42; ~;,
(ii) R41 42;
(iii) R41 ~41 R42;
(iv~ R42-~41-~41 R42;
(v3 R42 ~42 ~41 R41 R42-
In one embodiment, the peptide may have the following
structure:
WO 93/07892 2 l 2 0 3 3 7 PCT/US92/08823
-31-
Lys Lys Leu Leu Lys Lys Leu Lys Lys Leu
Leu Lys Lys Leu Arg Arg
(SEQ ID NO.:9~)
In another embodiment, the peptide includes the basic
structure X~2 ~ Z~2' wherein X~2 is as hereinabove described, and :
Z~2 is:
(i) R41; ..
(ii) R41 R41;
liii) 41 R41 R42; :
(iv) R4l-R41-R42 ~42'
(v) R41 R41 R42-~4~ 41;
In one embodiment, the peptide may have the following
structure: ~.
Lys Leu Lys Lys Leu Leu Lys Lys Leu Lys Lys Leu Leu Lys Lys Leu
~ 10 15 :.
(SEQ ID NO: 96)
In another embodiment, the peptide may include the
structure: ~ `
52 a ~2 ~ ~23b~ wherein X52, Ys2 and Z52 are as
hereinabove described, and a i5 0 or 1, and b is 0 or 1.
In one embodiment, the hereinabove described peptides may be ~:
acetylated with a CH3~O-group at the N-terminal.
In accordance with another embodiment, each of the amino
acid residues may be a D-amino acid residue or glycine. ;~.
Although the scope of this particular embodimen~ is not to
be limited to any theoretical reasoning, it is b~lieved that the
peptides and proteins of the present invention, when consisting
entirely of D-amino acid or glycine residue~, have in~reased
resistance to proteolytic enzymes while retaining their
biological activity. Such peptides may thus be administered
WO 93/07892 P~T/USg2/08~3
2~203~7
-32-
orally. Also, in accordance with another embodiment, all of the
amino acid residues may be D-amino acid or glycine residues, or
L-amino acid or glycine residues.
In still another embodiment, the peptide employed in
conjunction with an anti~iotic such as those hereinabo~e
described, or derivatives or analogues thereof i8 a cecropin.
The term cecropins includes the basic structure as well as
analogues and derivatives thereof. The cecropins and analogues
and deri~atives thereof are described in Ann. Rev. Microbiol
1987, Vol. 41 pages 103-26, in particular p. 108 ~nd Christensen
at al PNAS Vol. 8S p. 5072-76, which are hereby incorporated by
reference.
The term cecropin includes the basic structure as well as
analogues and derivati~es.
In yet another embodiment, the peptide employed in
conjunction with an anti~iotic such as those hereinabove
described, or derivatives or analogues thereof is a sarcotoxin.
The sarcotoxins and analogues and derivatives thereof are ~`
described in Molecular Entomoloav, pages 369-78, in particular p.
375 Alan R. Liss Inc. (1987), which is hereby incorporated by
r~ference.
The term sarcotoxin includes the basic materials as well as
analogues and derivati~es.
Ion channel-forming proteins or peptides which may be
employed include defensins, also known a~ human neutrophil
antimicrobial peptides (~NP), major basic protein (MBP) of
eosinophils, bactericidal permeability-increasing protein (BPI),
and a pore-forming cytotoxin called variously perforin,
cytolysin, or pore-forming protein. Defensins are described in
Selsted, et al., J. Clin. Invest., Vol. 76, pgs. 1436-1439
(198~). MBP proteins are described in Wa~moen, et al., J. Biol .
Chem., Vol. 263, pgs. 12559-12563 (1988). BPI proteins are
described in Ooi , et al., J. Biol. Chem., Vol. 262, pgs.
14891-14894 (1981). Perforin is described in HenXart, et al. J.
W0 93/0~892 2 1 2 0 3 3 7 PCT/US92/08823
-33-
EXP. Med., 160:75 (1984), and in Podack, et al. J. EXP. Med.,
160:695 (1984). The above articles are hereby incorporated by
reference.
The term ion channel-forming proteins includes the basic
structures of the ion channel-forming proteins as well as
analogues or derivatives.
Pseudomonic acids are produced by Pseudomonas fluoressens.
A preferred pseudomonic acid which may be employed i5 mupirocin,
or pseudomonic acid A, which has the following sturcture:
~'j ~h C; ~c_o~
JYO
The cephalosporins include a 7-aminocephalosporanic acid
nucleus. A preferred cephalosporin is cefotaxime, which has the :
following structure:
~O-~c~ _ O _ ~/_ C,~
C00f~a
Etham~utol, isoniazid, and ethionamide are especially useful
in treating mycobacterial infections, and in particular
infections of M. tuberculosis. Ethambutol is a synthetic,
WOg3/0~892 . PCT/US92/08823
2120 ~7
- 3~ -
water soluble, heat stable compound and is the D-isomer of
the structure below: .~
CH20H C2H5 . :,
H--C--NH--( CH2)2--Nt~ CH
C2H5 CH20H
Ethambutol may be dispensed as the dihydrocholoride
salt. Ehtionamide, a close chemical relative of isoniazid, H
has the following structure: . ;
H5C ~ ~
.'.
S" `NH2 -~ '
Sulfonamides have the following nucleus:
S02NHR
NH2
to which various R radical in the amido`group ~ S02NHR) have
been attached or in which various substitutions of the amino
group ~NH23 are made. Representative exa~ples of the
sulfonamides and their structure are given hereinbelow:
52 NH~
~J ;
SUBSTITUTE SHEET
W093/078s2 2 1 2 0 3 3 7 PCT/US92/~8823
- 3s - ~-
sulfadiazine :
S 2 N H\~
CH3
sulfisoxazole
S2 NH~
[~3 ~CH3
sulfamethoxazole
SO2NH~
COOH
NHCO~
sulfathalidine
Penem antibiotics which may be employed include~
but are not limited to, imipenem, carbapenems such as RS-533
(Sankyo), 2-(N-azolyl~ ~lkyl-substitu~ted penems, sucb as CGP-
29~718 (Ciba-Geigy), and broad-spectrum penems such as Sch-
34343 (Schering Plough). Imipenem, also known as a
thienamycin antibiotic, has the following structure:
NH
COOH
SVB~T~ i ~ J ~ FT
W093/07892 PCT/U~92/08823
212~337 36 -
Preferred aminoglycoside antibiotics which may be
employed are tobramhcin and the gentamicins. Tobramycin has
the following structure:
H~C--NH2 N~ NHz
~0~ ;.
H H~
Tbe gentamicins (Gentamicin C~, Gentamicin C2, and
Gentamicin C1~), as well as netilmicin, have the following
basic structure: R1 :
C~NHR2 NH2 NHR3 . ~:
~ ''
N~2HO ~ OH ~
~ H3 :
HO NH-CH3
For Gentamicin Cl, R1 and R2 are each CH3, the Cb-C5 bond is a
single bond, and R3 is H. For Gentamcin C2, R1 is CH3, R2 and
R3 are each H, and the C~-C5 bond is a single bo~d. For
Gentamicin C1a, R1, R2 and R3 ea h are H, and the C~-C5 bon~ ic
a single bond. For netilmicin, R1 and R2 each are H, R3 is
C2~5, and the Cb-C5 bond is a double bond.
Other aminoglycosides which may al~o be employed
within the scope of the present invention include, but ~re
not limited to, kanamycin and amikacin, as well as
metilmicin, neomycin, and streptomycin, which is alo
effective in treatihg tuberculosis. Kanamycin ~nd amikacin
both have the following basic structure:
H2C--N11,! N~NII-R
HO~ ~(
., ~LI CH20
H1:1 0U HO ~
HD NH2
S ~J 3S ~1 , . . _ ~ ~
W093~07892 2 1 2 ~ 3 3 ~ PCT/US92/08823
- 37 -
For kanamycin, R is H, and for amikacin, R i5:
o OH
Il /
C - CH - CH2- CH2 - NH2
Erythromycin, which is isolated from Streptomyces
erythreus, is a member of a group of compounds known as
macrolides. The basic structure of a group of compounds ~;
known as macrolides. The basic structure is a large lactone
ring to which unusual sugars are attached. The term
"macrolide" refers to a large ring formed from a chain of 14
to 20 carbon atoms by lactone condensation of a carboxyl and
hydroxyl group. Other macrolides include oleandomycin,
spiramycin, kitasamycin, and carbonmycin.
Erythromycin has the the following structure:
~ 3
0/~ ~
CH3 CIIJCII3~l 1 1
\/ o/\~\
/ ¦ N(Cl13)2
1~1 (I I oll
H3 ~ ~ - Cll 3
Cll 2 0 0~ O y Cll 3
Cli3
Cl130 ~
~ IP..C~T'T~ ~ ~T`~
W093/07892 PCT/US92/0~23
212~337
_ 38
It is also to be understood that other macrolide
antibiotics, such as roxythromycin, clarithromycin, and
others hereinabove described, may be employed as well.
For purposes of illustration of exanples of other
macrolide structures, the following examples, in addition to :
the above structure of erythromycin, are given below.
Rokitamycin is of the following structure:
CH3 C}i~CH2
~CH~
HO~ O ~ H ¦ CH3
-^OH ~ 1 OCCH~CHJ
Cll~ ~ H\ N~C~/¦ ~ OdCN.CH,CN.
O D ~ H -
CH3
CGP-7040, a benzapiperazinyl rifamycin, has the
following structure:
CH 3 ~H 3 Cll 3
CH3CDO ~ ~
I CH3 ~ -
J OH OH ~ ~ CH 3
~CH3 1 ¦ I
¦ ~ ~ N ~ N ~ H
CH3 ~;
SUBSTITUTE SHEE~T ~
., .". .,. ~ ." .~:
W093/07892 2 1 2 0 3 3 7 PCT/US92/0~23
- 39 -
It is to be understood, however, that the scope of
the present invention is not to be limited to the specific
macrolides or macrolide structures hereinabove described.
Preferred penicillins which may be employed in
accordance with the present invention are benzyl penicillin
(penicillin G) and ampicillin (alpha-amino-benzyl
penicillin). Benzyl penicillin is of the following
structure:
O C11J
~Cll 2 - C- NllF~ C~l 3
COOli
Ampicillin has the following structure:
O Cll~
C}l-C-N1l ~ ~ ~ Cll~
COO}l
A preferred monobactam which may be employed in
accordance with the present invention is aztreonam, which has
the following structure:
O
` 11 (;113
~1 1 G--~--Nl~
~N N ~N
z I O ~S~
O - C ( C ~ 3 ) 2
C~QII
IR.C~T~T~
W093/07892 . PCTrUS92/0~23
2 12 ~ 3 3 l - 39/1-
The bacitracins which may be employed in accordance
with the present invention tend to be hydrophilic and water ~:
solubleO The preferred bacitracin is bacitracin A, which has
the following structure:
,~S ~ G,H --CIICII z Cll 3
~¦ Nl12 Cll~
N
.1 ' ':
C= O .
L-l{is- D-Asp-L-Asn L-Leu
D-phe~ D-Glu
L-Ile -D-Orn- L- a -Lys-L-lle
Peptide antibiotics, which are not ion channel-
forming peptides or proteins, which may be employed include,
but are not limited to, gramacidin-S, polymyxin, vancomycin, :
teichoplanin, and capreomycin.
~J~ ~T'.~JT~ ~H~FT
W093/07892 2 1 2 0 3 ~ 7 PCT/US92/08823
-4~-
Vancomycin is a tricyclic glycopeptide antibiotic. Its
chemical formula is C66H15C12H9024, and it hafi a molecular weight
of 1,44~.
It is to be understood, however, that the ~cope of the
invention is not to be limited to the specific antibiotics and
antibiotic structures hereinabove described.
The present invention will be further described with respect
to the following examples, however, the ~cope of the invention is
not to be limited thereby.
ExamDle 1 `:
S. aure~s organisms are grown to mid log phase, and then
diluted to 10 organisms/ml in 1/~ ~trength trypticase soy broth.
After the incubation of the organisms, the minimal inhibitory ~`-
concentraion in ug~ml for Z-52 MG-2(amide-terminated), PGLa, CPF -~
Z-50, A-97, Magainin II, and Z-74 peptides, agai`nst S.aureus was
measured when each peptide was added alone to the organisms.
"MG-2 (amide)" is amide-terminated Magainin II, and 1'Magainin~
is car~oxy-terminated Magainin II. A-C7 peptide is of the
following structureo
(SEQ ID N0:97) amide
Z-74 peptide is of the following str~lcture:
~ 5EQ ID N0:98)-amide
The PGLa peptide is of the followinq structure:
~ SEQ ID N0:12)-amide.
The minimal inhibitory concentration (MIC) for each peptide
was then measured when 20% of the minimal inhi~itory
concentration of bacitracin, to~ramycin or gentamicin,
respecitvely, is added to the organisms along with the peptide.
The gentamicin employed is a mixture of Gentamicin Cl, Gentamicin
WO 93/07892 PCT/US92/08X23
212Q337
-41-
Cla, and Gentamicin C2. For Examples 1-3, the MlC values for
bacitracin are 2 ~g/ml against S. aureus, 64 ~g/ml against E. :
coli, and >2~6 ~g/ml against 8~e9~e~a~YD~o~kqh~8~- The MIC
values for gentamicin are 256 ~g/ml again~t S. aureus, 2 ~g/ml
against E. coli, and >256 ~g/ml against P. aeru~inosa. The MIC
values for tobramycin are >256 ~g/ml against S. aureus, 2 ~g/ml
against E. coli, and 128 ~g/ml again~t P. aeru~inosa. CP~ Z-50
peptide is (5EQ ID N0:21) of the CPF peptides hereinabove
described. Z-52 peptide is of the structure (SEQ ID NO:~9)-NH2.
The minimal inhibitory concentrations are given below in Table I.
TABLE I
MIC With
Peptide _dded Antibiotic
Pe~tide Alone _Bacitracin~ Gentamicin Tobram~cin
Z-S2 32 8 16 16 -
MG-2 amide256 32 ~ 32
PGLa 32 16 2 32 ~-
A-97 32 16 4 32
Maqainin 2256 128 4 32
Z-50 16 16 2 8 -~
Z-74 1~ 16 4 8 -
EXAMPLE 2
E.coli organisms were incu~ated according to the prcedure
described in Example 1. After the incubation, the minimal
inhibitory concentrations against E.coli of each of the peptides
described in Example 1 alone, as well as of each peptide when
employed in ~ombination with 20% of the MIC for bactracin, were
then measured. The results are given in Table 2 below.
wo g3/07892 2 1 2 ~ 3 3 7 Pcr/US92~08823
Ta~le 2
PePtide MIC
PePtide Pe~t_de with
Alone _ Bacitracin
Z-52 ~ ~
MG-2 amide 16 4
PGLa 16 8
Z-50 16 16
A-97 8 4
Magainin II32 8
(amide)
Z-7~ 16 8
ExamPle 3
In this example, P.aeru~inosa organisms were incubated
according to the procedure described in Example 1. After the
incubation, the minimal inhibitory ~oncentration~ of the peptides -~.
hereinabQve described alone, as well a~ the peptides in
combination with 20~ of the MIC of bacitracln, were then
measured. The results are given below in Table
Table 3
PePtide MIC .
Peptide Pe~tide with
Alone Bacitracin Added
Z-52 ~6
MG-2 amide128 8
PGLa 64 8
Z-~0 32 4
A-97 32 4
Magainin ~I256 1
Z-74 16 16
EXAMPLE 4 :~
7 PCT/US92/08823
-43-
In this example, E. coli or P. aeruqinosa organisms were
incubated according to the procedure described in Example 1.
After, the incubation, the minimal inhibitory concentrations of
the peptides hereinabove described alone, as well a5 the peptides
in combination with 20% of the minimal inhibitory concentrations
(MIC) of benzyl penicillin or ampicillin, were then measured.
The MIC of benzyl peni illin against E. coli i~ 64 ~g/ml, and of
ampicillin against E. coli is 4 ~g/ml. The results are given
below in Table 4.
TABLE 4
E. coli-
MIC ..
Peptide Peptide and Peptide and :~
PePtide alone Ben~yl ~enicill_nAm~icillin ~.
Z-52 16 16 -
Mg 2-amide 64 32 8
PGT.a 32 16
Magainin
II 64 32 16
Z-50 32 8
A-97 32 16 . 16
Z-74 4 8
P. aeru~inosa-
Pe~tide
PGLa 128 32 32
Mg~2
amide 256 128 128
W O 93/07892 . 2 1 2 0 3 3 7 PC~r/US92/08823
-44-
E~U~MPLE S
E. coli and P. aeruq nosa organisms were incubated according
to the procedure described in Example 1. After the incubation,
the minimal inhibitory concentrations of the peptides hereina~ove
described alone, as well as the peptides in combination with 20%
of the MIC of the monobactam antibiotic aztreonam, were then
measured. The MIC of aztreonam against E. col$ is a 2 ~g/ml, and ~
against P. aeru~inosa i~ 8 ~g/ml. The results are given below in ~.:
Table 5.
:.:
':
- .
WO 93/0~8g2 PCT/US92/08823
2l2n337 :~
-45-
TABLE S
E. coli-
MIC
Peptide Peptide with :~
PePtide alone Aztreonam
Z-52 16 2
Mg 2-amide 64 32
PCLa 32 4
Z-50 32 8
A-97 32 8
Magainin
II 64 16
Z-?4 4 8
P. aeru~inosa-
Pe~tide
Z-~2 32 32
Mg-2
amide 128 64
PGLa 128 64
Z-~0 64 3X
A-97 32 32
Magainin
II 256 128
Z-74 32 8
The above r~sùlts indicate that when one of the biologically
active ion-channel forming peptides hereinabove described is
added in combination with bacitracin, tobramycin, gen~amicin,,
benzyl penicill~n, ampicillin, or aztreonam, in an amount of 20%
of the MIC of these antibiotics, against S.aureus, E.coli, or
WO 93/07X92 2 1 2 ~ 3 3 7 PCT/US92/08823
4~
P.aeruqinosa, there is, in most cases, a resulting synergy
between the peptide and the antibiotic. In most cases, less
peptide and less antibiotic may be used against these organisms
when peptide and antibiotic are employed in combination, than if
peptide or antibiotic alone were employed.
Exam~le 6
In this example, the effect of a combination of erythromycin
and biologically active amphiphilic ion channel-forming peptide
will be measured against K.Pneumoniae, P.aeruqinosa, E.coli, and
5.aureus.
For each of the organisms listed in Table 6 below, 105
organisms were mid-log inoculated into 200 ~1 of one-half
strength trypticase soy broth. The organisms were incubated for
15 hrs. at 37C. After the incubation of the organisms, the
minimal inhi~itory concentration in ~g/ml for Magainin II
(amide-terminated), shown as MGN2, CPF Z-50 (amide-terminated)
peptide, Z-52 (amide-terminated) peptide, against each species of
organism was measured wherein 10 ~g~ml of erythromycin was added
and wherein no erythromycin was added. The minimal inhibitory
concentration of erythromycin alone against each species of
organism was also measured. The minimal inhibitory
concentrations in ~g/ml are given below in Table 6. For purpQses
of explanation, the "~" and "-" signs below the "Erythromycin (10
~g/ml)" column indicate the presence or a~sence, respectively,
of erythromycin ~dministered in combination with one of the `-
biologically active peptides. K.~neumQniae, P.aeruqinosa, and
E.coli are Gram-negative bacteria.
W093/07892 PCT/US92fO8B23
~33~ `
- 47
Table 6
Minimal Inhibitory
Concentration (~g/ml)
Erthro- MGN2 CPF Z-52 Erytho
mycin
Organism (10~/ml) amide z-50 amide mycin
amide
K.Pneumoniae - 5 ~0.5 1.5 >100
+ <0.5 <o.~~0.5
P.ae~uquinosa - 50 100 ~500 5-15 ~100
+ <5.0 <0.5~1.5
E.coli - 5 <5.0 l.S >100
+ 0.5 <0.5<0.5
The above results show that when one of the ~-
biologically active ion channel-forming peptides shown in
Table 6 is added to 10 ~g/ml of erythromycin, such a
combination of peptide and erythromycin is effective against
the Gram-negative organisms shown, whereas grea~er than 100
~g/ml of erythromycin alone is required for effective
biological a~tivity against these Gram-negative organisms.
The addi*ion of erythromycin to the biologically active
peptides also enables one to use less of the biologically
active peptide against Gram-negatiYe organis~. Thus, there
is provided a synergistic effect against Gram-negatiYe
organisms when erythromycin and a biologically active
amphiphilic ion channel-fo~ming peptide are administered to
inhibit growth of Gram-negative organisms.
It was also found that a concentration from a~out
0.6 ~g/ml to about 1.25 ~g~ml of erythromycin alone was
required for effectiYe biological activity agai.nst S._aureus,
a Gram-positive organism, and that greater than 250 ~g/ml of
amide-terminated organism, and that greater than 250 ~g/ml of
amide-te~minated Magainin II alone was required for effective
biological activity against S.aureus. A combination of lO
~g~ml of amide-terminated
SUBS~TU ~ r ~HFFT
WO 93/0789~ 2 1 2 0 3 3 7 PCT/US92/08823
-48-
M~gainin II and 0.03 ug/ml of erythromycin, however, al80 showed
effective biological activity against S.aureus. Thus, it has
also been shown that an enhanced effect against Gram-positive
organisms is also obtained when erythromycin and a biologically
active amphiphilic ion channel-forming peptide are administered
to inhi~it growth of Gram-positive organisms. The addition of
erythromycin to the bioloyically active peptide enables one to
use less of the biologically active peptide against a
Gram-positive organism.
ExamPle 7 -:~
The procedure for t~is assay, which is a checkerboard assay,
is performed as described in Antibiotics and Laboratorv Medicine,
2 nd ed., Victor Lorian, M.D., Editor, pgs. 540-546 (19~6).
The checkerboard assay is carried out in a microtiter plate
having wells arranged in rows and columns, 100 ~1 of plain broth
is added to every row of wells. 100 ~1 of peptide (SEQ ID
N0:100) at 512 ~g/ml is added to the top row of wells, and
serially dil~ted (1:2) through the columns. 50~1 o vancomycin
in 4X concentrations is added to each appropriate column of
wells to give a range of vancomycin concentrations from 16 to
1,024 ~g/ml. 50~1 of P. aeru~lnosa strain ATCC 27853 is thenr
added to each well. The plate is incubated at 35-37C for 18 to
24 hours, and the wells are then examined for the presence of
visible growth, i.e., turbidity.
Peptide (SEQ ID N0:100~-N~2 was tested alone or in
combination with vancomyein for activity against _ aeruainosa --
strain ATCC 27853.
In this assay, the MIC of (SEQ ID N0:100)-NH2 wa~ 32 ~g/ml,
and the MIC of vancomycin was greater than 1,024 ~g/ml; i.e.,
vancomycin at a concentration of 1,024 ~g/ml did not inhibit
growth of P. aeru~inosa.
WO g3J07892 PCT/US92/08823
2~2D337
-4g-
The following combinations of (SEQ tD N0:100)-N~2 and
vancomycin were found to be inhibitory.
~ ~q/ml ~SEQ ID NO:100)-NH2 plus 16 ~g/ml ~ancomycin
8 ~g/ml (SEQ ID NO:100)-NH2 plus 32 ~g/ml vancomycin
8 ~g/ml (SEQ ID N0:100)-NH2 plus 64 ~g/ml vancomycin
8 ~g/ml (SEQ ID N0:100)-NH2 plus 128 ~g/ml vancomycin
8 ~g/ml (SEQ ID NO:100)-NH2 plus 256 ~g/ml vancomycin
8 ~g/ml (SEQ ID NO:100)-NH2 plus S12 ~g/ml vancomycin
8 ~g/ml (SEQ ID N0:100)-NH2 plus 1,024 ~g/ml vancomycin
The above results thus indicate that an enhanced effect
against P. aeruainosa is obtained when vancomycin and a
biologically active ion channel-forming peptide are administered
to inhibit growth of P. aeruqinosa.
The peptide or protein and antibiotic such as those
hereinabove des~ribed, may be enployed for treating a wide
variety of hosts. In accordance with a preferred embodiment, a
host is an animal, and such animal may be a human or non-human
animal~ It is also possible to administer the peptide or protein
and antibiotic in separate forms. For example, the antibiotic
may be administered systemically and the peptide or protein may
be administered topically.
The peptide or pr~tein and/or antibiotic such as those
hereina~ove described, may be employed in a wide variety of
pharmaceutical compositions in combination with a non-toxic
pharmaceutical carrier or vehicle such a~ a filler, nun-toxic
buffer, or physiological saline solution. Such pharmaceutical
compositions may be used topically or systemically and may be in
any suitable form such as a liquid, solid, semi-solid, injectable
soiution, tablet, ointment, lotion, paste, capsule, or the like.
The peptide or protein and/or antibioti~ such as those
hereinabove described may also be used in combination with
adjuvants, protease lnhibitors, or compatible drugs where such a
combination is seen to be desirable or advantàgeous in
controlling infection caused by harmful microorganisms.
WO 93/07892 2 1 2 0 3 3 7 P~T/US92/08X23 ~
-50-
When the peptide or protein is administered topically, it is
administered in combination with a water-soluble vehicle, said
water-soluble vehicle being in the form of an ointment, cream,
lotion, paste, or the like. Examples of water-soluble vehicles
which may be employed include, but are not limited to, glycols,
such as polyethylene glycol, hydroxycellulose, and KY Jelly. The
water-soluble vehicle is preferably free of an oily substance.
The combination of peptide or protein and antibiotic of the
present invention may be administered to a host; in particular an
animal, in an effective antibiotic amount. When used to inhibit
growth of bacterial cells, the combination, whether administered
as a mixture or separately, is employed in an effective
antibacterial amount. When used to inhibit growth of fungi, such
components are administered in an effective antifungal amount.
As representative examples of administering the peptide or ~-
protein and antibiotic for topical or local administration, the ~-
peptide or protein could be administered in an amount of from
about O.l,o to about 10% weight to weight; and the antibiotic is
delivered in an amount of from about 0.1~ to about 10% weight to
weight. -~;
Numerous modifications and variations of the present
invention are possible in light of the above teachings and,
therefore, within the scope of the appended claims, the invention
may be practiced otherwise than as partiGularly d~cribed.
W093/07892 PCT/US92J08823
2 1203 37
SEQUENCE Ll STING
(1) GENERAL INFORMATION:
(i) APPLICANT: Berkowitz, Barry A.
Zasl~ff, Michael
(ii) TITLE OF INVENTION: Composition and Treatment wi~h
Biologically Active Peptides and Antibiotic.
(iii) NUMBER OF SEQUENCES: 100
(iv) CORRESPONDENCE ADDRESS:
(A) ADDRESSEE: Carella, Byrne, Bain, Gilfillan,
Cecchi ~ Stewart
(B) STREET: 6 Becker Farm Road
(C) CITY: Roseland
~D) STATE: New Jer~ey
(E) COUNTRY: USA
(F) ZIP: 07068
~v) COMPUTER READABLE FORM: ~
(A) M~DIUM TYPE: 3.5 inch diskette ~ -
(B) COMPUTER: IBM PS/2
(C) OPERATING SYSTEM: PC-DOS
~D) SOFTWARE: DW4.V2
(~i) CURRENT APPLICATION DATA
(A) APPLICATION`NUMBER:
(B) FILING DATE:
(C) CLASSIFICATION:
(vii) PRIOR APPLICATION DATA:
WO 93/07B92 2 1 2 0 3 3 7 PCT/US92/08823
-5~-
(A~ APPLICATION NUMBER:
U507402642
~B) FILING DATE: 05-SEP-1989
(A) APPLICATION NUMBER: ~:
US073392g2 "'
~8) FILING DATE: 17-APR-1989
(viii) ATTORNEY/AGENT INFORMATION:
(A) NAME: Ol~tein, Elliot M.
(B) REGISTRATION NUMBER: 24,025
(C) REFERENCE/DOCKET NUMB~: 421250
(ix) TELECOMMUNICATION INFORMATION:
(A) TELEPXONE: 201-994-1700
~B) TELEFAX: 201-994-1744
(2) I~FORMATION FOR S Q ID NO:1: `~
(i) SEQUENCE CHARACTERISTICS .~;
(A) LENGTH: 20 amino acids ~
(B) TYPE: amino acid ,l:
(C) STRANDEDNESS~
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(x) PUBLICATION INFO~MATION:
(H) DOCUMENT NUM8ER: W089/11290 -~
(I) FILING DATE: 19-MAY-1989 ~:
(J, PUBLICATION DRTE: 30-NOV-1989 ~:
WOg3t0~892 PCT/US92/~8823
2120337 "`
..
-53-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:1:
Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe
Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys
(2) INFORMATION FOR SEQ ID NO:2
(i) SEQUENCE C~ARACTERISTICS:
(A) LENGT~: 24 amino acids
(B) m E: amino acid
(C~ STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(x) PUBLICATION INFORMATION: ;
(H) DOCUMENT NUMBER: W089/11290
(I) FILI~G DATE: l9-MAY-19~9
(J) PUBLICATION DATE: 30-NOV-1989
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:2:
Ala Phe Ser Lys Ala Phe Ser Lys Ala Phe
Ser Lys Ala Phe Ser ~ys Ala Phe Ser Lys
1~ 20
Ala Phe Ser Lys
(2) IN~ORMATION FOR SEQ ID NO:3:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 16 amino acids
(B) m E: amino acid
(C) STRANDEDNESS:
(D~ TOPOLOGY: linear
WO 93~078922 1 2 0 3 ~ ~ PCI/US92/0~823
-54- -:
(ii) MOLECULE TYPE: peptide
(x) PUBLICATION INFORMATION:
(H) DOCUMENT NUM8ER: W089/11290
(I) FILING DATE: l9-MAY-l9~
(J) PUBLICATION DATE: 30-NOV-1989
(xi) SEQUENCE DESCRIPTION: SEQ ~D NO:3:
Phe Ser Lys Ala Phe Ser Lys Ala Phe Ser :~
Lys Ala Phe Ser Lys Ala
(2) INFORMATION FOR SEQ ID NO:4~
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH~ 20 amino acids
(B) TYP : amino acid ::
(C) STRANDEDNESS:
~D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(x) PUBLICATION TNFORMATION:
(H) DOCUMENT NUM8ER: W089/11290
(I) FILING DATE: 19~MAY-1989
(J) PUBLICATION DATE: ~O-NOV~1~89
~xi) SEQUENCE DESCRIPTION: SEQ ID NO:4: ~-
Ser Lys Ala Phe Ser Lys Ala Phe Ser Lys
5 10
Ala Phe Ser Lys Ala Phe 5er Lys Ala Phe
(2~ INFORMATION FOR S~:~ ID NO:5:
W093/~7892 PCT/US92/08~23
212~3~7
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 16 amino acids
( B ) TYPE: ami ~o acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(x) PUBLICATION INFORMATION:
(~) DOCUMENT NUMBER: W089/11290
(I) FILING DATE: l9-MAY-1989
(J) PUBLICATION DATE: 30-NOV-1989
(xi~ SEQUENCE DE~CRIPTION: SEQ ID NO:5:
~ Lys Ala Phe Ser Lys Al~ Phe Ser Lys Ala
Phe Ser Lys Ala Phe Ser
(2) INFORMATION FOR SEQ ID NO:6:
~i) SEQUENCE CHARACTERISTICS :.
(A) LENGTH: ~23 ~mino acids
(8) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
.
MOLECULE TYPE: peptide
(A) NAME/KEY: Magainin I peptide.
~x~ PU~LICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNA~: Proceedin~s ~f ebe N~t_ooal Academ~
of Sciences
(D) VOLUME: 84
':~
:
W093~07~92 2 1 2 0 3 3 7 PCT/US92/08823
. -56-
( F ) PAGES: 5449-5453
(G) DATE: AUG - 1987
~H) DOCUMENT NUMBER: US 4810777
(I) FILING DATE: 04-MAR-19B7
(J) PUBLICATION DATE: 07-MAR-1989
'~
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:6
Gly Ile Gly Lys Phe Leu His Ser Ala Gly
5 10 ~
Lys Phe Gly Lys Ala Phe Val Gly Glu Ile :
Met Lys Ser
(2) INFORMATION FOR SEQ ID NO:7: ;
(i) SEQUENCE CHARACTERISTICS
(A3 LENGTH: 23 amino acids
(B) TYPE: amino acid
~C) ST~ANDE~N%SS:
(D~ TOPOLOGY: linear -
, .
(ii) MOLECU~E TYPE: peptide
(ix) FEAT~RE:~
(A) NAME/KEY: Magainin II peptide.
(x~ PUBLICATION INFO~MATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedinqs of the National AcademY
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-5453
(G) DATE: AUG - 1987
~H) DOCUMENT NUMB~R: US 4810777
(I) FI~ING DATE: 04-MA~-1987
W093/07892 PCT/US92/08823
2120337
~J) PUBLICATION DhTE: 07-MAR-19B9
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:7
Gly Ile Gly Lys Phe Leu His Ser Ala Lys
S 10
Lys Phe Gly Lys Ala Phe Val Gly Glu Ile
15 20
Met Asn Ser
(2) INFORMATION FOR SEQ ID NO:8: -:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 22 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide ~-
(ix) FEATURE: -
(A) NAME/KEY: Magainin III peptide.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedinqs of the National Aca~emv
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-54S3
(G) DATE: AUG - 1987
~H) DOGUMENT NUMBER: US 4810777
(I) FILING DATE: 04-MAR^1987
(J) PUBLICATION DATE: 07-MAR-1989
W093/07892 2 1 2 ~ 3 3 7 PCT/US92/08823
-5fi-
(xi) SEQUENCE DESC~IPTION: SEQ ID NO:8
Gly Ile Gly Lys Phe Leu His Ser Ala Ly~
5 10
Lys Phe Gly Lys Ala Phe Val Gly Glu Ile
}~ 20
Met Asn :
(2) INFORMAT~ON FOR SEQ ID NO:9:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 22 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOL~GY: linear
(ii~ MOLECULE TYPE: peptide ~;
(ix) FEATURE:
(A) Nt~lE/KEY: magainin peptide.
( x ) PUBL I CAT I ON I NFORMAT I ON:
~A) AUTHOR: Zasloff, Michael
(C) JOURNAL: Proceedinqs of the National Academv
of Sciences ~;
( I) ) VOLUME: 84
(F) PAC;ES: 5449-5453
(G) DATE: AUG - 1987
(H) DOCUMENT NtJMBER: US 4810777 :-
( I ) FILING DATE: 04-MAR-1987
(J) PUBLICATION DATE: 07-MAR-1989
WO 93/07892 PCT/USg~/08823
21~Q33~
-59- .
(xi3 SEQUENCE DESCRIPTION: SEQ ID NO:9:
Ile Gly Lys Phe Leu His Ser Ala Lys Lys
5 10
Phe Gly Lys Ala Phe Val Gly Glu Ile Met
15 20
Asn Ser
(2) INFORMATION FOR SEQ ID NO:10:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid ::
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: magainin peptide.
(x) PU8LICATION INFORMATION: ~ -:
~A) AUTHOR: Zasloff, ~ichael
(C) JOURNAL: ~3~E~L~g__of the National Academy
of Scien es
(D) VOLUME: 84
(F3 PAC;ES: 5449-5453
(G) DAT~: AUG 1987
(H) DOCUMENT NUMBER: US 4810777
(I) FILING DATE: 04-MAR-19B7
~J) PUBLICATION VAT : 07-MAR-1989
WO 93/07892 2 1 2 0 3 3 ~ PCT/US92/08~23
-6~- .
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:10:
Gly Lys Phe Leu His Ser Ala Lys Lys Phe
Gly Lys Ala Phe Val Gly Glu Ile Met Asn
1~ 20
Ser
(2) INFORMATION FOR SEQ ID NO:ll:
(i~ SEQU~NCE CHARACTERISTICS
5A~ LENGTH: 20 amino acids
(B~ m~ amino acid ~`
(C) STRANDEDNESS:
(D) TOPOLOGY: linear ~ -
.'.
(ii) MOLECULE mE peptide
(ix) FEATURE:
~A) NAME/KEY: magainin peptide. ~:
(x) PUBLICATIQN INFORMATION:
~A) AUIHOR: Za510ff, Michael
(C) JOURNAL: Proceedinqs of the Nati nal AcademY
of Sciences
(D) VOLUME: 84
(F) PAGES: 5449-5453
~G) DATE: AUG - 1987 -:
~H) DOCUMENT NUMBER: US 4810777
~I) FILING DATE: 04-MAR-1987
(J) PUBLICATIO~ DATE: 07-MAR-1989
W093/0~892 PCT/US92/08823
2120337
-61-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:ll:
Lys Phe Leu His Ser Ala Lys Lys Phe Gly ~:
S 10 ,
Lys Ala Phe Val Gly Glu Ile Met Asn Ser - ..
15 ~ :
(2) INFORMATION FOR SEQ ID NO:12: :~
(i) SE~UENCE CHARACTERISTICS ~
(A) LENGTH: 21 amino acids ::
(B) m E: amino acid :~
(C) STRANDEDNESS:
(D) TOPOLOGY: linear ;
'~."
(ii) MQLECULE TYPE: peptide -:
(ix) FEATURE: ~
(A) NAME/KEY: PGLa peptide. .~.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Hoffman, et al.
(C) JOURNAL: EMBO J.
(D) VOLUME: 2
(F) PAGES: 711-714
~G) DATE: l9B3
(A) AUTHOR: Andreu, et al.
(C) JOURNAL: Journal of Bio hemis~rY
(D3 VOLUME: 149
(F) PAGES: 531-535 :~:
(G) ~ATE: l9a5~
(A) AUT~OR: Gibson, et al.
(C~ JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G1 DATE: 1986
WO 93/07892 2 1 2 0 3 3 7 PCT/US92/0~23 . :
-62-
(A) AUIHOR: Giovannini, et al.
(C) JOURNAL: Biochem J.
(D) VOLUME: 243
(F) PAGES: 113-120
(G) DATE: 1987 -:~
(xij SEQUENCE DESCRIPTION: SEQ ID NO:12:
Gly Met Ala Ser Lys Ala Gly Ala Ile Ala .
.~,.
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala
lS 20 ~-
Leu
(2) INFORMATION FOR SEQ ID NO:13:
(i) S~QUE~CE CHARACTERISTICS
(A) LENGTH: 25 amino acids
(B) TYP~: amino acid
(C) ST~ANDEDNESS:
(D) TOPOLO5Y: linear :
(ii) MOLECU~E TYPE: peptide
(ix) FEATURE.
(A) NAME/KEY: XPF peptide.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Hoffman, et al.1
(C) JOURNAL: MBO J.
~D) VOLUME: 2
(F) PAGES: 711-714
(~) DATF~: 1983
(A) AUTHOR: Andreu, et al.
~C) JOURNAL: Journal of Biochemistr~
tD) VOLUME: 149
WO 93/07892 P~T/US92/08823
2120337
-63-
(F) PAGES: 531-535
(G) DATE: 1985
(A) AUTHOR: Gibson, et al.
~C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(A) AUTHOR: Giovannini, et al.
(C) JOURNAL: Biochem J.
(D3 VOLUME: 243
(F) PAGES: 113-12
(G) DATE: 1987
(xi) CE~UENCE DESCRIPTION: SEQ ID NO:13:
Gly Trp Ala Ser Lys Ile Gly Gln Thr Leu
5 lO
Gly Lys Ile Ala Lys Val Gly Leu Lys Glu
Leu I le Gln Pro Lys
:
(2) INFORMATION FOR SEQ ID NO: 14:
( i ) SEQUENCE CHA~ TERISTICS
(A) LENGTH: 27 amino acids
~ B ) TYPE: amino acid
( C ) STRANDEDNESS:
(D) TOPOLOt;Y: linear
( ii ) MOLECULE TYPE: peptide
( i x ) FEATURE:
~A) NAME/KEY: C~F peptide. ~
( x ) PUBLICATION INFORMATION: -
WO 93/07892 . 2 ~ 2 0 3 3 7 PCT/US92/0%823
-64- ~:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: 1986
(A) AU~HOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Re~earch
(D) VOLUME: 13
~F) PAGES: 1817-1828
(G) DATE: 198$
(A1 AUTHOR: Gibson, ~.W.
Poulter, L.
Williams, D.~.
Maggio, J.E. ``
~C) JOURNAL: J Biol. Chem.
(D~ VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
~H) DOCUMENT NUMBER: WOa0/04407
(I) FILING DATE: 16-OCT-1989
~J) PUBLICATION DATE: 03-MAY-1990
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:14:
Gly Phe Gly Ser Phe Leu Gly 1eu Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
Leu Gly Gly Ala Pro Gln Gln
~5
W093/~78g~ PCT/US92/08823
-65-
(23 INFORMATION FOR SEQ ID NO:lS:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
tc) STRANDEDNESS: ~
(D) TOPOLOGY: linear :
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Rirht~r, K
Egger, R. :
Kreil ::.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 251 ~:
(F) PAGES: 3~76-3680 ~:
(G) DATE: 198S
(A) AUTHOR: Wakabayashi, I.
Kato, H.
Tachibaba, S.
(C) JOURNAL: ~Es~h~ a~s_Research
(D~ VOLUME: 13
(~) PAGES: 1817-1828
(~) DATE: lg85
(A) AUTHO~: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) ~OLUM~: 261 .
(F) PAGES: 5341-5349 - ~
' '.
2 1203~7
W093/07~92 PCT/US9~/08823
-66
(G) DATE: 1986
(H) DOCUMENT NUMBER: W090/04407
(I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DA~E: 03-MAY-1990
(xi) 5EQUENCE DESCRIPTION: SEQ ID NO:lS:
Gly Leu Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Gly Leu Lys Ile Gly Ala His Leu
lS 20
Le~ Gly Gly Ala Pro Gln Gln :
2) INFORMATION FOR SEQ ID NO:16:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 27 amino acids
~B) TYPE: amino acid
(C) STRANDEDNESS:
~D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(x~ PUBLICATION IN~ORMATION:
(A~ AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOU~NAL: J. Biol. Chem
(D) VOLUME: ~61
(F~ PAGES: 3676-368
(G) DATE: 1986
(A) AUTHOR: Wakabayashi~ T.
WO 93/07892 . PCI`/US92/08823
212OJ37
Kato, H.
Ta ::hibaba, S .
(C) JOURNAL: Nucleic Acids Resear~h
(D ) VOLVME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985 ::
( A ) AUTHO~: Gibson, B . W .
Poulter, L.
Williams, D.H. -
Maggi o, J . E .
(C) JOURNAL: J. Biol. Chem.
( D ) VOL~ME: 2 61
(F) PAGES: 5341-~34
( G ) DATE: 1986
(H) DOCUMENT NUMBER: W090/04407
(I) FILING DATE: 16-OCT-1~8g
( J ) PIJBLICATION DATE: 03-MAY- 1990
(xi ) SEQUENCE DESCRIPTION: SEQ ID NO: 16: ::
Gly Leu Ala Ser Leu Leu Gly Lys Ala Le~
S 10
Lys Ala Gly Leu Lys Ile Gly Thr E~is Phe
2û
Leu Gly Gly Ala Pro Gln Gln
(2 ) INFORMATION F5:)R SEQ ID NO: 17:
i ) SEQUENCE C~ARACTERI STICS
(A) LENGTH: 27 amino acids
(B~ TYPE: amino acid
( C ) STRANDEDNESS:
( D ) TOPOLOC;Y: 1 ine ar
( ii ) MOLECULE TYPE: peptide
2120337
WO 93/07892 ~ PCT/US92/08823
-68-
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. 8iol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
~G) DATE: 1986 :
(A) AUTHOR: Wakabayashi, T. :
Kato, H.
Tachibaba, S.
(C) ~OURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F) PAGES: lB17-1828
~) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E. ~ ~`
(C~ JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NUMBER: WO90/04407
(I) FILING DATE: 16-OCT-1989
~J) PUBLICATION DATE: 03-MAY l99O
WO 93/07892 PCT/USg2/08823
2121)337
-6g- :
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:17:
Gly Leu Ala Ser Leu Leu Gly Lys Ala Leu .
5 10
Lys Ala Thr Leu Lys Ile Gly Thr His Phe
~.
Leu Gly Gly Ala Pro Gln Gln -
(2) INFORMATION FOR SEQ ID NO:18~
(i) SEQUE~CE CHARACTERISTICS
(A) LEN~TH: 27 amino acids
~B) TYPE: amino acid
(C) STRANDEDNESS: .
(D) TOPOLOGY: linear ::
(ii) MOLECULE TYPE: pe~tide ~
(ix) FEATURE:
(A) NAME/KEY: C~F pepti~e.
(x) PUBLICATION INFORMATIO~:
~A) AUIHOR: Richter, K. , :-
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem
(D) VOLUME: 261 -
(E) PAGES: 3676-3680
(G) DATE: 1986
(A) AUTHOR: Wakabayashi, T.
:
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids_Research
(D) VOLUME: 13
(F) PAGES: 1817-1828 -
wo 93/0,8g2 2 1 2 0 3 3 7 PCT/US92/~8823
-70~.
(G) DATE: 1985 .~
(A) AUTHOR: Gibson, B.W. :
Poulter, ~.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986
(H) DOCUMENT NVM~ER: WO90/04407
~I) FILING DATE: 15-OCT-1~89
(J) PUBLICATION DATE: 03-MAY-1990
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:18:
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Met
~0
Leu Gly Gly Thr Pro Gln Gln
~:.
.
~2) INFORMATION FOR SEQ ID NO:1:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino arid
~C) STRANDEDNESS:
~D) TOPOLOGY: linear
~ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(A) NAME~KEY: CPF peptide.
(x) PUBLICATION INFORMATION:
W093/07~2 PCT/US92/~823
2120337
(A) AUIHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. ~iol. Chem. -~-
(D) VOLUME: 261
(F) PAGES: 3676-3680
(Gj DATE: 1986
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: I~ucleic Acids_Researoh
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: 1985
(A) AUIHOR: Gibson, B.W.
Poulter, L.
Williams, D.B.
Maggio, J.E.
(C) JOURNAL- J. Biol. Chem.
~D) VOLVME: 261 :~
(F) PAGES: 5341-5349
(G~ DATE: la86
~H) DOCUMENT NUMBER: WO90/04407
~I) FILING DATE: 16-OCT-1989
(J) PUBLICATION DATE: 03-MAY-1990
~xi~ SEQUENCE DESC~IPTI~N: SEQ ID NO:19:
Gly Phe Gly Ser Phe Leu 51y Lys Ala Leu :~
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
Leu Gly Gly Ala Pro Gln Gln
::.
W093/07892 2 1 2 ~ 3 3 7 pCT~US92/08823
(2) INFORMATION FOR SEQ ID NO:20: -
(i) SEQUENCE CHARACTERISTIC5
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE: ~
(A) NAME/KEY: CPF peptide.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G3 DATE: l9B6
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Taçhibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
(F~ PAGES: 1817-1828
(G) DATE: 1985
(A) AUTHOR: Gibson, B.W.
Poulter, L.
William , D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
WO 93/07892 PCI/USg2/08823
2 12~337
-73-
( G ) DATE: 1986
(H) DOCUMENT ~UM8ER: W090/04407
( I ~ FILING DATE: 16-OCT- 1989
(J) PUBLICATION DATE: 03-MAY-1990
(xi) SEQUENCE DE5CRIPTION: SEQ ID NO:20
Gly Phe Gly Ser Phe Leu Gly Lys Al a Leu
S 10 . ,,~
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
1~ 20 ~
I.eu Gly Gly Ser Pro Gln Gln .~.
~2) INFORMATION FOR SEQ ID NO:21: ~.
~i ) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids ~:
(8) TYPE: amino a~id
( C ) STRANDEDNESS:
(D ) TOPOLOGY: linear
~ii ) MOLECULE TYPE: peptide
( i x ) FE:ATURE:
~ A ) NAME/KEY: CPF pepti de .
( x ) PUBL I CATI QN INFORMAT I ON:
( A ) AUTHOR: Ri chter, K .
Egger, R.
Krei 1
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: l9B6
(A) AUTHOR: Waka~ayashî, T.
212~337
WO 93/07892 PCT/USg2/0~823
-74-
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D~ VOLUME: 13
(F) PAGES: 1817-1828
(G~ DATE: 198~
(A) AUTHOR: Gibson, B.W.
Poulter, L. :~
Williams, ~
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261
(F) PAGES: 5341-5349
(G) DATE: 1986 :
(H) DOCUMENT NUMBER: WO90/04407
(I) EILING DATE: 16-OCT-1989
~J) PUBLICATION DATE: 03-MAY-1990 -~
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:21:
Gly Phe Ala Ser Phe Leu Gly Ly~ Ala Leu
Lys Ala Ala Leu Lys lle ~ly Ala Asn Leu
1~ 20
Leu Gly Gly Thr Pro ~ln Gln
2~
(2) INEORMATION FOR SEQ ID NO:22:
~i) SEQUENCE CRARACTERISTICS :`
(A) LENGTH: 27 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
~D) TOPQLOGY: linear
(ii) MOLECVLE TYPE: peptide
WO g3/078g2 . PCI`~US92/08823
212~337
-75-
.
( ix ) FEATURE:
(A) NAME/KE;Y: CPF peptide.
. .
( x ) PUBLIC~TION INFORMATION:
( A ) AUTHOR: Ri chter, K .
Egger, R.
Kre i l
~C) JOURNAL: J. Biol. Chem ~.
(D) VOLUME: 261
(F) PAGES: 3676-3680 ::.
( G ) DATE: 1986 ~;
~A) AUT~OR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOL~JME: 13
~ F ) PAGES: 1817-1828
( C; ) DATE: l9S5 ~ -
( A ) AUT~OR: Gi}:~son, B . w .
Poult~r, L.
Williams, D.H.
Maggi o, J . E .
(C) JOIJRNAL J. Biol. Chem.
~ D ~ VOLUME 2 61
(F) PAGE5: 5341-5349
~ G ) DATE: 1986
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:22:
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
Leu Gly ~;ly Ala Pro Gln Gln
- :
wo 93/078g2 2 1 2 0 3 3 7 PCT/US92/08X23
-7~- :
(2) INFORMATION FOR SEQ ID NO:23~
~i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B) TYPE: amino a~id `
(C) STRANDEDNESS: -
(D) TOPOL WY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATU~E:
(A) NAME/KEY: CPF peptide.
(x) PUBLICATION INFO~MATION:
(A) AUTHOR: Richter, ~. ~
Egger, R.-~.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLUME: 261 ~::
(~) PAGES: 3676-3680
(G) DATE: 1~86
(A) AUIHOR: Wakabayashi, T. ~-
Kato, H.
Tachibaba, S.
(C) JOURNAL:
~D~ VOLUME: 13
(F~ PA~ES: 1817-18~8
(G) DATE: 1985
~A~ AUT~OR: Gibson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J. Biol. Chem. ~:
~D) VOLUME: 261
(F) PAGES: 5341 5349
-:.
.-:
W~ ~3,~,0 ~ 7 PCT/US92/~8823
-77-
(G) DATE: 1986 .
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:23:
Gly Phe Ala Ser Phe Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Met
lS 20
Leu Gly Gly Ala Pro Gln Gln
- 25
(2) IN~ORMAT~ON FOR SEQ ID NO:24:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 27 amino acids
(B~ TYPE: amino acid
(C) STRAN~EDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE ~YPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(x) PUBLICATION I~FORMATION:
(A) AUT~OR: Richter~ K.
Egger, R.
Kreil
(C) JOURNAL: J. ~iol~ Chem.
(~) VOLUME: 261
(F) PAGES: 3676-3680
(G) DATE: l9B6
(A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, S.
(C) JOURNAL: Nucleic_Acids Research
W093/07892 2 1 2 0 3 3 7 Pcr/US92/0~823
-78- ::
(D) VOLUME: 13
~F) PAGES: 1817-1828 :~.
(G) DATE: 1985
~A) AUTXOR: Gibson, 8.W.
Poulter, L.
Williams, D.H.
Maggio, J.E. :
(C) JOURNAL: J. Biol. Chem.
tD) VOLUME: 261
~F) PAGES: 5341-5349
(G) DATE: 1986
~xi) SEQUENCE DESCRIPTION: SEQ ID NO:24: ::~
Gly Phe Gly Ser Phe Leu Gly Lys Ala Leu
5 10 :
Ly5 Ala Ala Leu Lys Ile Gly Ala Asn Ala
lS 20
Leu Gly Gly Ser Leu Gln Gln
:
(2~ INFORMATION FOR SEQ ID NQ:2S:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 27 amino acids
(B) TYPE: ami~o acid
(C) STRANDEDNES5:
(D) TOPOLOGY: linear
(ii) MOLECULE IYPE: peptide
(ix) FEATURE:
(A) NAME/KEY: CPF peptide.
(x) PUBLICATION INFORMATION:
(A) AUTHOR: Richter, K.
W093/07892 PCT/US92~08823
2 1 ~3~7
-79-
Egger, R.
Kreil
(C) JOURNAL: J. Biol. Chem.
(D) VOLVME: 261
~F) PAGES: 3676-3680
(G) DATE: 1986
~A) AUTHOR: Wakabayashi, T.
Kato, H.
Tachibaba, 5.
(C) JOURNA~: Nucleic Acids R~cear~h
(D) VOLUME: 13
(F) PAGES: 1817-1828
(G) DATE: l9a5
(A) AUTHOR: G~bson, B.W.
Poulter, L.
Williams, D.H.
Maggio, J.E.
(C) JOURNAL: J Biol. Chem.
(D) VOLUME: 261
~F) PAGE5: 5341-534
(G) DATE: 1986
. .
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:25:
Gly Phe Gly Ser Phe Leu Gly Lys Ala Leu
5 1~
Lys Ala ~ly Leu Lys Ile Gly Thr Asn Phe
~5 ~0
Leu Gly Gly Ala Pro Gln Gln
(2~ INFO~MATION FOR S~Q ID NO:26:
(i) SEQUENCE CHARACTERISTICS
(A) ~ENGTH: 27 amino aeids
W093/û7892 2l2n337 PCI`/US~2/08823
-80-
( B ) TYPE: amino acid
( c ) STRANl:~EDNESS:
(D ) TOPOLOGY: linear
( ii ) MOLECULE TYPE: peptide
( i x ) FEATURE:
(A) NAME/KEY CPF peptide.
( x ) PUBLI CATI ON INFORMP.TION:
(A) AUTHOR: Richter, K
Egger, R.
Krei 1 ~:
(C~ JOURNAL: J. Biol. Chem.
(D~ VOLUME: 261
(F) PAGES: 3676-3680
( G ) DATE: 1986
( A ) AUTHOR: Wakabayashi, T .
Kato, H.
Tachi~aba, S.
(C) JOURNAL: Nucleic Acids Research
(D) VOLUME: 13
( F ) PAGES: 1817-1828
(G) DATE: l9B5
( A ~ At~OR: Gibson, B . W .
Poulter, L.
Williams, D.H.
Maggio, J. E.
( C ) JOURNAL: J . _ Biol . Chem .
( D ) VOLUME: 2 61
(F) PAGES: 5341-53
(G) DATE: 1986
WO 93/07892 PCT/uS92/0X823
2l2n.~ 7
-81-
(xi) SEQUENCE DESCRIPTION: SEQ I~ NO:Z6:
Gly Leu Ala Ser Leu Leu Gly Lys Ala Leu
Lys Ala Ala Leu Lys Ile Gly Ala Asn Ala
lS 20
Leu Gly Gly Ser Pro Gln Gln
(2~ INFO~MATION FOR SEQ ID NO:2~:
(i) SEQUENCE CHAR~CTERISTICS
~A) LENGTH: 2l amino acids
(B) TYP~: amino acid
(C~ STRANDEDNESS:
~) TOPOLOGY: linear
~ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:27:
Lys Ile Ala Gly Lys Ile Ala Lys Ile A1B
Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile
lS 20
Ala
(2) I~IFORMATION FOR SEQ ID NO:28:
(i) SEQUENCE CHA~ACTERISTICS:
~A) LEN~TH: 2l amino acids
(B~ m E: amino acid
(C) STRANDEDNESS:
[D) TOPOLOGY: linear
(ii) MOLECVLE TYPE: peptide
WO 93/07892 2 1 2 0 3 ~ 7 PCT/US92/08823
-82-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:28:
Lys Ile Ala Lys Ile Ala Gly Lys Ile Ala .
S 10
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly
(2) INFORMATION FOR SEQ ID NO:29:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
~C~ STRANDEDNESS: :
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide -
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:29:
Lys Ile Ala Gly Lys Ile Gly Lys Ile Ala
Gly Lys Ile Gly Lys Ile Ala Gly Lys Ile
1~ 20
Gly
(2) INFORMATION ~OR SEQ ID NO:30:
(i) SEQUENCE C~ARACTERISTICS
(A) LENGTH: 21 amino acids
~B) TYPE- amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
WO 93/~7892 - PCT/US92/08823
21203~7 ` ~`
-83- :
.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:30:
Lys Leu Ala Gly Lys Leu Ala Lys Leu Ala
Gly Lys Leu Ala Lys Leu Ala Gly Lys Leu
Ala
(2) INEORMATION ~OR SEQ ID NO:31:
(i) SEQUENCE ~HARACTERISTICS :~:
(A) LENGT~: 21 ami~o acids
(B) TypF: amino acid
(C~ STRANDED~ESS:
~D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
~xi) SE~UENCE D~SCRIPTION: SEQ ID NO:31: 5''
Lys Phe Ala Gly Lys Phe Ala Lys Phe ~la
5 10
Gly Lys Phe Ala Lys Phe Ala Gly Lys Phe
Ala
(2~ INFORMATION FOR SEQ ID NO:32:
(i3 SEQUENCE CHARACTERISTIC5
(A) LENGTH: 21 amino acids
(B) TYPE: amino a~id
~C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
WO 93/O~X92 2 ~ 2 ~ 3 3 7 PcT/vs92/o88~3
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:32: -
Lys Ala Leu Ser Lys Ala Leu Lys Ala Leu
5 10 :-~
Ser Lys Ala Leu Lys Ala Leu Ser Lys Ala ~-~
lS 2D ::
Leu :
(2) INFORMATION FOR SEQ ID NO:33:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acid~
~B) m E: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ~D NO:33:
Lys Leu Leu Lys Ala Leu Gly Lys Leu Leu
Ly5 Al a Leu Gly Lys Leu Leu Lys Ala Leu
~;ly
(2) INFORMATION EOR SEQ ID NO:34:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino a~ids
(B) m E: amino acid
(C) STRANDEDNESS;
~D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
.~. .
, . .
W093/~7892 PCT/US92/08823
212033~
-85-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:34:
Lys Ala Ile Gly Lys A~a Ile Lys Ala Ile
5 10
Gly Lys Ala Ile Lys Ala Ile Gly Lys Ala -
Ile
(2) INFORMATION FOR SEQ ID NO:35:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino a~id
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
~ .
(xi~ SEQUENCE DESCRIPTION: SEQ ID NO:35: ~-
Gly Ile Ala Lys Ile Ala ~ys Gly Ile Ala
5 10 ~
Lys Ile Ala Lys Gly Ile Ala Lys Ile Ala :
Lys
(2) INFORMATION FOR SEQ ID NO:36:
(i) SEQUENCE CHARACTERISTICS
(A~ LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
.,,~ .
,~ .
WO 93/07892 2 1 2 0 3 3 7 PCT~US92/0~8~3 ~
-8~- :
~xi) SEQUENCE DESCRIPTION: SEQ ID NO:36:
Lys Ile Ala Lys Ile Phe Gly Lys Ile Ala
Lys Ile Phe Gly Lys Ile Ala Lys Ile Phe
Gly
~2) INFORMATION FOR SEQ ID NO:37:
(i) SEQUENCE CHARACT~RISTICS
(A) LENGTH: 21 amino acid~
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) NOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ I~ NO:37:
_Gly ~le Ala Arg Ile Ala Lys Gly Ile Ala
Arg Ile Ala Lys Gly Ile Ala Arg Ile Ala
Lys
(2) INFORMATION FOR SEQ ID NO:38:
(i~ SEQUENCE CRARACTERISTICS
~A) LENGTH: 21 amin~ acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MO~ECULE ~YPE: peptide
W093/07892 PCT/US92/08823
2120337
-B7-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:3B:
Lys Phe Ala Arg Ile Ala Gly Lys Phe Ala
Arg Ile Ala Gly Lys Phe Ala Arg Ile Ala
Gly
~2) IMFOXMATION FOR SEQ ID NO:39:
(i) SEQUENCE CHARACTERISTICS
- (A) LENGTH: 21 amino acids
(B) TYPE: amino acid
~C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQ~ENC~ DESCRIPTION: SEQ ID NO:39:
_Gly Phe Ala Lys Ile Ala Lys 51y Phe Ala
Lys I le Ala Lys Gly Phe Ala Lys Ile Ala
15 20 -
Lys `~
(2) INFORMATION FOR SE~ ID NO:40:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TY}?E: amino acid
~ C ~ STRANDEDNESS:
( 1) ) TO~OLQGY: 1 i ne ar
.
(ii) MOLECULE TYPE: peptide
212~3~7
WO 93/07892 PCT/US92/08823
-8B-
.
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:40:
. Lys Ile Ala Gly Orn Ile Ala Lys Ile Ala
Gly Orn Ile Ala Lys Ile Ala Gly Orn Ile
Ala
(2) INFORMATION F0R SEQ ID N0:41:
(i) SEQUENCE C~AR~CTERISTICS
~ ~A) LENGTH: 21 amino acids ::
~B) TYPE: amino acid
( C ) STRANDEDNESS:
D ~ TOPOLOGY: linear
:.
(ii~ MOLECULE TYPE;: peptide ::
(xi) SEQIJENCE DESCRIPTION: SEQ ID NO:41:
Lys Ile Ala Arg Ile Ala Gly Lys lle Ala
Arg Ile Ala Gly Lys Ile ~la Arg Ile Ala
~0
Gly
~2) IN~0RMATI0N ~OR SEQ ID NO:42:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 21 amino acids
(B) TYPE: amino acid
C ) STRANDEDNE S S:
~ D ) TOPOI.OGY: l i ne a r
(ii) MOLECULE TYPE: peptide
WO 93/07892 PCT/US92/08~23
212D337
:
-89-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:42:
Orn Ile Ala Gly Lys Ile Ala Orn Ile Ala
5 10 .:
Gly Lys Ile Ala Orn Ile Ala Gly Lys Ile
15 20 :
Ala ~:~
J (2) INFORMATION FOR S~Q ~D NO:43:
(i) SEQUENCE CHARACTERISTICS ::
- (A) LENGTH: 21 amino acids
(B) m E: amino acid
(C) 5TRANDE~NESS: ~
~D) TOPOLOGY: linear :
.:
Iii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:43:
Gly Ile Ala Arg Ile Phe Lys Gly Ile Ala
Arg Ile Phe Lys Gly Ile Ala Ar~ Ile Phe .~:
Lys
:~
(2) INFO~;ATION FOR SEQ ID NO:44:
(i) SEQUENCE C~ARACTERISTICS
(A) LENGTH: 21 amino acids
(B) m E: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
21203~7
WO 93!07892 PCT/US92/08823
-90-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:44:
Lys Nle Ala Gly Lys Nle Ala Lys Nle Ala
5 10 ;
Gly Lys Nle Ala Lys Nle Ala Gly Lys Nle
Ala
(2) INFORMATION FOR SEQ ID NO:45:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDN~SS:
(D) TOPOLOGY: linear
(ii) MOLECUL~ TYPE: peptide
(xi) SEQUENCE DESCRI~TIO~: SEQ ID NO:54:
Lys Nle Ala Gly Lys Ile Ala Lys Mle Ala
Gly Lys Ile Ala ~ys Nle Ala Gly Ly~ Ile
1~ 20
Ala
~2) INFORMATION ~OR SEQ ID NO:46:
(i) SEQUENCE C~ARACTERISTICS
~A) LENGTH: 21 amino acids
(~1 TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOG~: linear
(ii) MOLECULE mE peptide
WO 93r07892 PCI~US92/08~23 :
212U3~7 :
-gl- ~.
(xi ) SEQUENCE DESCRIPTION: SEQ II) NO: 46:
Lys Ile Ala Gly Ly~i Nle Ala Lys Ile Ala
S 10
Gly Lys Nle Ala Lys Ile Ala Cly Lys Nle
-
Ala
( 2 ) INFORMATION FOR SEQ ID NO: 47:
( i ) SEQUENCE CHARACTERISTICS :~
(A) LENGTEI: 21 amino acid~ ~'
(B) TYPF~: amino acid
( C ) STRANDEDNE S S:
(D) TOPOLOGY: linear
( ii ) MOLECULE TYPE . peptide :
(xi ) SEQUENCE DESCRIPTION: SEO ID NO: 47:
- Lys Nva Ala Gly Lys Nva Ala Lys Nva Ala
5 10 .
Gly Lys Nva Ala Lys Nva Ala Gly Lys Nva
Ala
2 ) INFORMATION FC)R SEQ ID NO: ~a
(i ~ SEQUENCE C~ARACTERISTICS
(A) LENGTHo 21 smino acids
( ~ ~ TYPE o amino acid
~ C ) STRANDEDNESS:
(D~ TOPOLOGY: linear
( i i ) MOLECULE TYPE: pept i de
WO 93/07892 2 ~ 2 D 3 3 7 Pcr/US92/0~823
-92-
(xi) SEQUENCE DESC~IPTION: SEQ ID NO-48:
Lys Nva Ala Gly Lys Ile Ala Lys Nva Ala
lQ
Gly Lys Ile Ala Lys Nva Ala Gly Ly5 Nva
Ala
( 2 ) INFORMATION FOR SEQ ID NO: 49:
( ~ ) SEQUENCE CHARACTERISTICS
- (A) I.ENGl~l: 21 amino acid~
(B) TYPE: amino acid
( C ) STRAN~EDNES S:
(D ) TOPOLOGY: linear
( i i ) MOLECULE ~YPE: peptide
(xi) SE5~UENCE D5C~IPTION: SEQ ID NO:49:
_ Lys I.eu Leu Ser Lys Leu Gly Lys Leu Leu
Ser Lys Leu Gly l.ys Leu Leu Ser Lys Leu
Gly
(2) INFORMATION FOR SEQ Il:) NO:50:
~i ) SEQIJENGE CH~RAC:TERISTIC5
(A) LENGTH: 21 amino acids
( B ) ~:PE: aminc~ acid
C ) STRANDE:DNESS:
~D) TOPOLOGY: lYne~r
( ii ) MOLECULE TYPE: peptide
WO93/078g2 PCT/uS92/08$23 ::~
2:12a337 ~
-93- -~
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:50: ~
Lys Leu Leu Ser Lys Phe Gly Lys Leu Leu ~:
Ser Lys Phe Gly Lys Leu Leu Ser Lys Phe
Gly
(2) INFORMATION FOR SEQ ID NO:51:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 21 amino a~ids
~B) TYPE: amino acid ::
(C~ STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide :
(xi) SEQUENCE DESC~IPTION: SEQ ID NO:51:
~ Lys Ile Ala Gly Lys Nva Al~ Lys Ile Ala
Gly Lys Nva Ala Lys Ile Ala Gly ~ys Nva
Ala
(2) INFO~MATION FOR S~Q ID NO:52:
EQUENCE CHARACTERISTICS
(A) ~ENGT~: 21 ami~o acids
(B) m E: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MO~ECULE TYPE: peptide
wo 93~0~8g2 2 1 2 0 3 3 7 PCT/~S92/08823
-94-
.
(xi) S~QUENCE DESCRIPTION: SEQ ID NO:52:
His Ile Ala Gly His Ile Ala His Ile Ala
Gly His Ile Ala His Ile Ala Gly His Ile
Ala
(2) INFORMATION FOR SEQ ID NO:53:
(i) SEQUENCE CHARACTERI5TICS
(A) LENGTH: 21 amin~ acids
(B) TYPE: amino acid
tC) STRANDEDNE55
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:53:
~Ala Gly Lys Ile Ala Lys Ile Ala Gly Lys
Ile Ala Lys Ile Ala Gly ~ys Ile Ala Lys `~
Ile
(2) INFORMATION FOR SEQ ID NO:54:
(i) SEQUENCE CHARACTERISTICS
(A) LENGT~: 21 amino acids
(B~ TYPE: amino acid
SC) STRANDEDNESS
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
WO 93/07892 PCT~US92108823
2121)337
-95- :
(xi) SEQUENCE DESCRXPTION: SEQ ID NO:54:
Ile Ala Lys Ile Ala Gly Lys Ile Ala Lys
Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly
1~ 20
Lys
.
(2) INFORMATION FOR SEQ ID NO:55:
(i) SEQUENCE CHARACT~RISTICS
(A) LENGTH: 21 amino acids `
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE- peptide
(xi) SEQUENCE DESC~IPTION: SEQ ID NO:55: -.
Lys Ile Ala Gly Arg Ile Ala Lys Ile Ala
f 5 10
~ly Arg Ile Ala Lys Ile Ala Gly Arg Ile
15 20
Ala
(2) INFORMATION FOR SEQ ID NO:56:
(i) SEQUENCE ~HARACTERISTICS
(A) LENGTH: 21 amino acids
~B) TYPE: amino acid
~C) STRANDEDNESS: -:
(D) TOPOLOGY: linear
(ii) MOLECULE mE peptide
WO 93/07892 2 12 D 3 3 ~ PCr/lJS92/08X23
_9~_
(xi ) SEQUENCE DESCRIPTION: SEQ ID NO: S6:
Arg Ile Ala Gly Arg Ile Ala Arg Ile Ala
- 5 10
C;ly Arg Ile Ala Arg Ile Ala Gly Arg Ile
i~0 .
Ala
(2) INFORMATION FOR SEQ ID NO:57:
( i ) SEQUENCE CHARACTERI STIC:S
(A) LENGS~I: 21 amis~o acids
(~) mE: arnino acid
( C ) STRA~DEDNESS:
( D ) TOPOLOGY: 1 ine ar
( ii ) MOLE5ULE TYPE: peptide
( xi ) SEQUENCE DESCRIPTION: SEQ ID NO: 57: ~:~
Lys Val Ala Gly Lys Ile Ala Lys Val Ala ;~
Gly Lys Ile Ala Lys Val Ala Gly Lys Ile :~
1~ 20 :~:
Ala
~`
(2) INFORMATION FOR SEQ ID NO:58:
( i ) SEQUENt:E CHARACTERISTICS
(A) LENGTH: 21 aminc: acids
(B~ mE: amino acid
C ~ 5TRANDE:DNESS: -
(D) TOPOLOGY: 1 inear
(ii) MOLECULE TS!PE: peptide
WO 93/07892 PCT/US92/08823 ~
2120337 ~
-97-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:58:
Lys Ile Ala Gly Lys Val Ala Lys Ile Ala
S 10 .,:
Gly Lys Val Ala Lys Ile Ala Gly Lys Val
Ala
(2) INEORMATION FOR SEQ ID NO:59:
(i) SEQUENCE CHARACTERIST$CS -~
~ (A) LENGTH: 21 amino acid~ :
(B) TYPE: amino a~id ~:
~C) STRANDEDNESS:
(D) TOPOLOGY: linear
~ii) MO~ECULE m E: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:59:
~Ala Lys Ile Ala Gly Lys Ile Ala Lys Ile :-
",."
Ala Gly Lys Ile Ala Lys Ile Ala Gly Lys .
15 20 -~
Ile
(2) INFORMATION ~OR SEQ ID NO:60:
(i) SEQUENCE CHARACTERISTICS
(~ LENGTH: 21 amin~ acids
(B) TYPE: amino acid
~C) STRANDEDNESS: :
(D) TOPOLOGY: 1inear
MOLECULE TYPE: peptide
W093/~7892 2 1 2 ~ 3 3 7 PCT/US92/08823
-98-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:60:
Orn Ile Ala Gly Orn Ile Ala Orn Ile Ala
S 10
Gly Orn Ile Ala Orn Ile Ala Gly Orn Ile
Ala
(2) INFORMATION FOR SEQ ID NO:61:
(i) SEQUENCE CHARACTERISTICS
~A) LENGTH: 21 amin~ acids -
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:61:
Lys Phe Ala Gly Lys Ile Ala Lys Phe Ala
Gly Lys Ile Ala L~s Phe Ala ~ly Lys Ile ~
lS 20 t:~'
Ala
(2~ INFORMATION FOR SEQ ID NO:62:
(i) SEQUENCE ~HARACTERISTICS
~A) LENGTH: 21 amino acids
(B3 ~YPE: amino a~id
(C3 STRANDEDNESS:
~D) TOPOLOGY- line~r --
~ii) MOLECULE TYPE: peptide
WO 93/07892 PCT/US92/08~23
2120337
99
~xi) SEQUENCE DESCRIPTION: SEQ ID NO:62:
Lys Ile Ala Gly Lys Phe Ala Lys Ile Ala
5 10
Gly Lys Phe Ala Lys Ile Ala Gly Lys Phe .:
~:
Ala
-
(2) INFORMAT~ON FOR SEQ ID NO:63:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amin~ acids .:
(B) TYPE: amino acid :
(C) ST~ANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:63: ::
Lys Cha Ala ~y Lys Ile Ala Lys ~ha Ala
Gly Lys Ile Ala Lys Cha Ala Gly Lys Ile ~:
lS 2
Ala
(2) INFORMATTON FOR SEQ TD NO:64:
~i) SEQUENCE CHARACTER~STICS
(A) LENGTH: 21 amino acids
~B) TYP~: amino acid
(C) STRANDEDNESS:
~D) TOPOLOGY: linear
~ii) MOLECULE TYPE: peptide
WO 93/07892 2 1 2 0 3 3 7 PCT/US9~/08823
-100-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:64: ;
Lys Nle Ala Lys Ile Ala Gly Lys Nle Ala
Lys Ile Ala Gly Lys Nle Ala Lys Ile Ala ::~
Gly
(2) INFORMATION FOR SEQ ID NO:65:
(i) SEQUENCE CHARACTERISTICS:
~A) LENGTH: 21 AMINO ACI~S
(B) m E: amino acids j.
(C) STR~NDEDNE55:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:65:
Arg Ile Ala ~ly Lys Ile Ala Arg Ile Ala
- 5 lO
Gly Lys Ile Ala Arg Ile Ala Gly Lys Ile :
1~ 20
Ala
(2) INFORMATION FOR SEQ ID NO:66:
(i) SEQUENCE CHARACTERISTlCS
(A) LEN~TH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide ;~
W~93/07892 PCT/Us92/08823
2~ 20337 ~:
- 101 - :~
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:66:
Har Ile Ala Gly Har Ile Ala Har Ile Ala :
Gly Har Ile Ala Har Ile Ala Gly Har Ile
Ala
(2)INFORMATION FOR SEQ ID NO:67:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 21 amino acids
(B) m E: amino a~id
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE: Xaa is p-aminophenylalanine
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:67:
Xaa Ile Ala Gly Lys Ile Ala Xaa Ile Ala
5 1~ :
Gly Lys Ile Ala Xaa Ile Ala Gly Lys Ile
1~ ~0
Ala
(2) INEORMATION FOR SEQ ID NO:6B:
(i) SEQUENCE CHARACTERISTICS
(A) ~ENGTH: Zl amino acids
(B) TYP : amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
WO 93/07892 2 ~ 2 ~ 3 3 7 PCT/US92/0~823
-102-
(ix~ FEATURE: Xaa is p-aminophenylalanine :~
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:68:
Lys Ile Ala Gly Xaa Ile Ala Lys Ile Ala
Gly Xaa Ile Ala Lys Ile Ala Gly Xaa Ile
Ala
(2) INFORMATION FOR SEQ ID NO:69:
SEQUENCE C~ARACTERISTICS
(A) ~NGTH: 21 amino acids .:
~B) TYPE: amino acid :~
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
~xi) SEQUENCE DESCRIPTION: SEQ ID NO:69:
Lys Leu Ala Ser Lys Ala Gly Lys Ile Ala Gly :-
Lys Ile Ala Lys Val Ala Leu Lys Alà Leu
15 20 ~:
(2) INFORMATION EOR SEQ ID NO:70:
(i) SEQUENCE C~ARACTERISTICS
(A~ LENGTH: 21 amino acids
(B) TYPE: amino acid
~C) STRANDEDNESS: :~
(D) TOP~LOGY: linear
(ii) MOLECULE TYPE: peptide
WO 93/07892 PCI'/US92/08823
2120337
-103-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:70:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala Gly
5 lO j;
Orn Ile Ala Lys Ile Ala Gly Lys Ile Ala
15 2~ ~-
(2) INFORMATION FOR SEQ ID NO:71:
(i) SEQUENCE CHARACTERISTICS
tA) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C~ STRANDEDNESS:
(D) TOPOLOGY: linear
~ii) MOLECU~E TYPE: peptide
~ xi) SEQUENCE DESCRIPTION: SEQ ID NO:71:
! Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Arg Ile Ala Lys Ile Ala Gly Lys Ile
~O
Ala
(~) INFO~MATION ~OR SEQ ID NO:72:
(1) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C~ STRANDEDNES5:
(D) TOPOLOGY: linear
(ii) MOLECU~E TYP~: peptide
WO 93/07892 2 1 7 ~ ~ 3 7 Pcr/US92/08823
-104-
'
( xi ) SEQUENC~; DESCRIPTION: SEQ ID NO: 72:
Lys Ile Ala Gly l.ys Ile Ala Lys Ile Ala
Gly Nle Ile Ala Lys Ile Ala Gly Lys lle
Ala
( 2 ) INFORMATION FOR SEQ ID NO: 73:
(i) SEQUEMCE CHARACTERISTICS ;~
(A) LE:NGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D ) TOPOLOGY: linear
(ii) MOLECU~E TYPE: peptide
~xi) SEQUENCE DESCRIPTI~N: SEQ ID NO:73:
Lys Ile Ala Gly Lys lle Ala Lys Ile Ala
Gly Nva Ile Ala Lys Ile Ala Gly Lys Ile ~
15 20 .
Ala
(2) INFORMATION FOR SEQ ID NO:74:
(i) SEQUENCE CHARACTER}STICS
~A) LE~GTH: 21 amino acids
(B) TY~E: amino acid
(C) STRAND D ~ESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
W093/07892 PCT/US92/08823
2120337
-105-
(xi) SE~UENCE DESCRIPTION: SEQ ID NO:74:
Lys Phe Ala Gly Lys Phe Ala Lys Phe Ala Gly
Orn Phe Ala Lys Phe Ala Gly Lys Phe Ala
15 20
(2) INFORMATION FO~ SEQ ID NO:75:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ }D NO.75:
Lys Tle Ala Gly Lys Phe Ala Lys Ile Ala
5 10
Gly Orn Phe Ala Ly~ Ile Ala Gly Lys Phe
15 20
Ala .
(2) INFORMATION FOR SEQ ID NO:76:
(i) SEQUENCE CHARACTERISTICS
~A) LENGT~: 21 amino acids
(B) TYPE: amino acid
~C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
WO 93/07892 2 1 2 0 3 3 7 PCT/US92~0B823
- l o~ - ,
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:76: :
Lys Ile Ala Gly Lys Nle Ala Lys Ile Ala
Gly Orn Nle Ala Lys Ile Ala Gly Lys Nle ~:
lS 20 -;
Ala .:
~2) INFORMATION FO~ SEQ ID NO:77:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acid~ :
~B) TYPE: amino acid
(C) STRAN~EDNESS: ;:
~D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide :~.
(xi) SEQUENCE D~SCRIP~ION: SEQ ID NO:77:
Lys Me~ Ala Ser Lys Ala Gly Lys Ile Ala :
5 1~ :~
Gly Lys Ile Ala Lys Vai Ala Leu Lys Al~
Leu
(2) INFORMATION FOR SEQ ID NO:78:
(i) SEQUENCE CHARACTERISTICS
(A) LENGTH: 21 amino acid3
(B3 TYPE: amino acid
(C) STRANDEDNESS:
(D) TO~OLOGY: linear
(ii) MOLECULE TYPE: peptide
W093/n7892 PCT/US92/08~23
2120337
-107-
(xi) S~QUENCE DESCRIPTION: SEQ ID NO:78
Lys Ile Ala Ser Lys Ala Gly Lys Ile Ala
5 10
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala Leu
1~ 20
(2) INFORMATION FOR SEO ID NO:79:
~i) SEQUENCE CHARACTERISTICS
(A) ~ENGTH: 21 amino acid~
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii).MOLECULE TYPE: peptide
~xi) SEQUENCE DESCRIPTION: SEQ ID NO:79:
Lys Ile Ala Ser Lys Ala Gly Lys Nle Ala
5 10
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala Leu
~2) INFORMATION FOR SEQ ID NO:BO:
(i) SEQUENCE CHA~ACTERI 5T I CS
tA) LEN~TH: 21 amino acids
~B) TYPE: amino acid
(C) STRANDEDNES s
~D) TOPOLOGY: linear
(ii) MOLECULE TYP~: peptide
W093/07892 2 1 2 ~ 3 3 7 PCT/US92/08823
-108- :~
(xi) SEQUENCE D~SCRIPTION: SEQ ID NO:80:
Lys Leu Ala Ser Lys Ala Gly Lys Nle Ala
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala
15 20 ~:.
Leu ~.
(2) INFORMATION FOR SEQ ID NO:Bl:
(i) SEQUENCE CHARACTERISTICS
~ (A) LEN~TH: 21 amino acids
(B) TYPE: amino acid
(C) STRAND~DNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:81: ~.
Lys Nle Ala Ser Lys Ala Gly Lys Nle Ala
5 10 ~:
~ly Lys Ile Ala ~ys Val Ala Leu Lys Ala Leu
1~ 20 `~
(2) INFORMATION FOR SEQ I~ NO:82:
(i) SEQUENCE CHARACTERISTICS:
(A~ LENGTH: 21 amino acids
(B) TYPE: aminc acid
(C) STRANDEDNESS:
(~) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: Xaa is p-aminophenylalanine.
~r'~ ,r~
W0 93/0789~ PCT/U592/U8823
-109-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:~2:
Lys Ile Ala Gly Ly~ Ile Ala Lys Ile Ala
Gly Xaa Ile Ala Lys Ile Ala Gly Lys Ile
Ala
(2) INFORMATION FOR SEQ ID NO:83:
(i) S~QUENCE CHARACTERI5TICS:
tA) LENGTH: 21 amino acids
~B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide ~
'`;
(xi) SEQUENCE DESCRIPTION: SEQ I~ NO:83:
~ Lys Ile Ala Gly Ala Ile Ala ~ys Ile Ala
S 10 -~
Gly Lys Ile Ala Lys Ile Ala Gly Ly~ Ile
Ala
(2) INFORMATION FOR SEQ ID NO:84:
(i) SEQUEN OE CHARACTERISTICS: :
(A) LENGTH: 21 amino acids
(B) TYPE: aminc) acid
( C ) ST~DEDNESS:
(D ~ TOPOLOGY: l inear
(ii) MOLECULE TYPE: peptide
WO 93/07892 2 1 ~ ~ 3 3 7 PCl`/US92/0~823
- 110-
(xi ) SEQUENCE DESCRIPTION: SEQ ID NO: B4:
Lys Ile Ala Gly Lys Ile Ala Lys Ile Ala
Gly Ala Ile Ala Lys Ile Ala Gly Lys Ile
-
Ala :
(2) INFORMATION FOR SEQ ID MO:85
( i ) SEQUENCE C~IARACTER I ST I CS: ~
(A) LENGTH: 21 amino acids ~;
(B) TYPE: amino acid
(C) STRANDEDNESS:
( D ) TOPOLOGY: l i ne ar
( ii ) MOLECULE TYPI:: peptide
~ xi ~ SEQUE;NC DE:SCRIPTION: SEO ID NO: 85:
~ Lys Ile Ala Gly Lys Ile Ala Lys Ile P,la
5 10 :
Gly Lys Ile Ala Lys Ile Ala Gly Ala Ile
Ala
~2) INFORMATION EOR SEQ ID NO:86:
i ) 5EOIJENCE CHARACTERI STIC5
(A) LENGTH: 21 amino acids
(B) mE: ~mino acid
(C) STRANDEI)NESS:
~D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
2 1 2 0 3 3 7 PCT/US92/~8823
..
(xi) S~QUENCE DESCRIPTION: SEQ ID NO:96:
- Lys Ile Ala Lys Lys Ile Ala Lys Ile Ala
5 10 :~:
- Lys Lys Ile Ala Lys Ile Ala Lys Lys Ile
15 20 ~
Ala .
(2) INFORMATION FOR SEQ ID NO:87:
(i) SEQUENCE CHARACTERISTICS
(A) ~ENGTH: 21 amino acids
(B) m E: amino acid
(C) STRANDEDNESS:
~D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:87:
Ala Ile Ala Gly Lys Il~ Ala ~yci Ile Ala
- S 10 ~,
Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile :
1~ 20 ~
Ala ~-:
~2) INFORMATION FOR SEQ ID NO:8~:
(i) S~QUENCE CHARACTERISTICS
~A) LENGTH: 21 amin~ acids
(B) mE amino acid
(C) STRA~DEDNESSi
(~) TOPO~OGY: ~inear :
(ii3 MOLECULE TYPE: peptide
WO 93/07892 2 1 2 0 3 3 7 PcT~us92/o8x23
-112-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO 88:
Lys Ile Ala Gly Lys Ile Ala Ala Ile Ala
Gly Lys Ile Ala Lys Ile Ala Gly Lys Ile
lS 20
Ala
(2) INFORMATION FOR SEQ ID NO:B9:
(i) SEQUEN OE CHARACTERISTICS
- (A) LENGTH: 21 amino acids
(B) TYPF.: amino acid
~C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(xi) SEQUENCE DE5CRIPTION: SEQ ID NO:89:
Lys Ile Ala Gly Lys Ile Ala ~ys Ile Ala
~ 5 10
Gly Lys Ile Ala Ala Ile Ala Gly Lys Ile
Ala
~2) INFORMAT1ON FOR SEQ ID NO:90:
(i~ SEQUENCE C~ARACTERISTICS
(A) LENGTH: 21 amino acids
(B) TYPE: amino acid
~C1 STRANDEDNESS:
~D) TOPOLOGY. linear
(ii) MOLECULE TYPE: peptide
2 ~ ~ ~ 3 ~ 7 PCT/~592/08~3
-113-
(xi) SEQUENCE DESCRIPTION: SEQ ID NO:9~:
Gly Met Ala Ser Lys Ala Gly Lys Ile Ala
Gly Lys Ile Ala Lys Val Ala Leu Lys Ala
Leu
(2) INFORMATION FOR SEQ ID NO:91:
~i) SE~UENCE CHARACTERISTICS
~ (A) LE~G~H: 11 amino acids
~B) TYPE: ami~o ~cid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
~:.
(ix~ FEATURE~
(D) OTH~R INFORMATION: May be a C-terminal amide,
and/or may be acetylated at
N-terminus. :
.:
(xi~ SEQUENCE DESCRIPTION:SEQ ID NO~
Leu Lys Lys Leu Lys Lys Leu Leu Lys Leu
5 10 -~.
Leu
(2) INFORMATION ~OR SEQ ID NO:92: ~-
(i) SEQUENCE C~ARACTERISTICS
(A) ~ENGT~: 12 amino acids
(B) TYPE: amino acid
(C) STRANDEDN~SS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
WO 93/07892 2 1 2 ~ 3 3 7 PCT/US92/08823
-114-
(ix) FEATURE:
(D~ OTHER INFORMATION: May be a C-terminal amide,
and/or may be acetylated at
N-terminus.
(xi) SEQUENCE DESCRIPTION:SEQ ID NO:92:
Leu Leu Lys Lys Leu Lys Lys Leu Leu Lys
5 10
Leu Leu a
(2) INFORMATION FOR SEQ ID NO:a3:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 13 amino acids :.
(B) TYPE: amino acid
(C) 5TRANDE9NESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: May be a C-terminal amide,
and/or may be acetylated at
N-terminus.
(xi) SEQUENCE DESCRIPTION:SEQ ID NO:93:
Lys Leu t~eU ~ys Lys Leu Lys Lys Leu Leu
Lys Leu Leu
(2) INFORMATION FOR SEQ ID NO:94:
(i) SE~UENCE CHARACTERISTICS:
(A) LENGTH: 14 amino acids
(B) TYPE: amino a~id
WO 93/07892 PCT/US92/08823
212D337
-llS-
(C) STRANDED~ESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
(D) OTHER INFORMATION: May be a C-terminal amide,
and/or may be acetylated at
N-terminus.
(xi) SEQUENCE DESCRIPTION:SEQ ID NO:94:
Lys Lys Leu Leu Lys Lys L~u Lys Lys Leu
- 5 10
Leu Lys Leu Leu
,,.
(2) INFORMATION FOR SEQ ID NO:95:
(i) SEQU~NCE CHARACTERISTICS:
(A) LENGTH: 16 amino acids
(B) TYPE: amino acid .
(C) STRANDEDNESS:
(D) TOPOLOGY: linear -
(ii) MOLECULE TYP : peptide
(ix) F~ATURE:
(D) OTHER INFORMAT$0N: May be a C-terminal
amide, and/or may be acetylated at
N-term~nu~.
(ix) SEQUENCE DESCRIPTION: SEQ ID MO:95~
Lys Lys Leu L~u Lys Lys Leu Lys LYB Leu Leu Lys Ly~ Leu Arg Arg
~ 10 lS
(2) INFORMATION FOR SEQ ID NO:96:
(i) SEQUENCE CHARACTERISTICs:
(A) LENCTH: 16 amino acids
W093/07892 2 1 2 0 3 3 ~ PCT/US~2/08823
-l16-
.:
(B) TYPE: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE TYPE: peptide
(ix) FEATURE:
~D) OT~ER INFORMATION: May be a C-t~rminal amide,
andfor may be acetylated at
N-terminu~.
~xi~ SEQUENCE DESCRIPTION:SEQ ID NO:96:
Lys Leu Lys Lys Leu Leu Lys Lys Leu Lys
Lys Leu Leu 1ys Leu Leu
(2) INFORMATION FOR SEQ ID NO:97:
(i) SEQUNCE CHARACTERISTIGS
(A) LENGTH: 22 amino acids
(B) TYP~: amino acid
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii~ MOLECULE TYPE peptide
(xi) SEQUENCE ~ESCRIPTION: SEQ ID NO:97:
Gly Ile Gly Lys Phe Leu His Ser Ala Gly
Lys Phe Gly Lys Ala Phe Val Lys Ile Met
Lys Ser
W093/07892 PCT~S9~/08823
2~21)337
-117- . :~
(2) INFORMATION FOR SEQ ID NO:98:
(i) SEQUENCE CHARACTERISTICS:
(A) LENGTH: 24 amino acids
(B) TYPE: amino acid -.
(C) STRANDEDNESS:
(D) TOPOLOGY: linear
(ii) MOLECULE m E: peptide
~xi) SEQUEN OE DESCRIPTION: SEQ ID NO:98:
Ala Gly Ile Gly Lys Phe Le~ His Ala Ala
Lys Lys Phe Ala Lys Ala Phe Val Ala Glu
15 20 ~:
Ile Met Asn Ser
(2) INFORMATlON FOR SEQ ID NO:99:
~- (i) SEQUE~C~ CHARACTERISTICS:
(A) LENGTH: 24 amino acids
(B~ TYPE: amino acid
~C) STRANDED~SS: ~:
(D) TOPOLOGY: linear
~ MOLECULE TYPE: peptide ~`
(xi~ SEQUENCE DESCRIPTION: SEQ ID NO:99:
Ala Leu Ser Lys Ala Leu Ser Lys Ala Leu
Ser Lys Ala Leu S~r Lys Ala Leu Ser Lys
~5 20
Ala Leu Ser Lys
(2) INFORMATION FOR SEQ ID NO:100:
(i) SEQUENCE CHARACTERIST~CS:
WO 93/07892 2 1 2 0 3 3 7 p~/U~i92/08~23
~ 118-
(A) LENGTH: 16 am~no acids
(B) TYPF~: amino acid
( C ) STRANDEDNESS:
(D) TOPOLOGY: linear
( ii ) MOLECULE ~FYPE: peptide
(xi ) SEQUENCE DESCRIPTION: SEQ ID NO: lOO
Gly Ile Lys Lys Phe Leu Lys Lys Ala Gly
Lys Phe Gly Lys Ala Phe
,.
