Note: Descriptions are shown in the official language in which they were submitted.
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BOLAAMPHIPHILIC COMPOUNDS, COMPOSITIONS AND USES THEREOF
FIELD
[0001] Provided herein are bolaamphiphilic compounds, complexes thereof
with specific
small interfering RNAs (siRNAs), and pharmaceutical compositions thereof Also
provided are
methods of delivering siRNAs into the human and animal cell using the
compounds, complexes
and pharmaceutical compositions provided herein. Also provided are methods of
delivering
siRNAs into the human and animal organs, such as the brain, using the
compounds, complexes
and pharmaceutical compositions provided herein.
BACKGROUND
[0002] In the past decade, efforts to develop RNA-based therapeutic
technologies have
significantly intensified1-5. Triggering RNA interference (RNAi), in
particular, has become one
of the most widely used techniques for biomedical applications1-11. RNAi
employs a mechanism
of post-transcriptional sequence specific gene silencing by processing double-
stranded RNAs
into small-interfering RNAs (siRNAs) used as part of the RNA-induced silencing
complex
(RISC) to selectively cleave target mRNA12. After the discovery that synthetic
siRNAs can be
exogenously introduced into cells to activate RNAin' 14, this approach has
become a powerful
method for selective suppression of specific genes of interest in different
species, showing
potential for use in cancer therapeutics3' 5,6 However, the biomedical utility
of the synthetic
siRNAs is limited by several RNA structure-related factors such as the
negative charge (uptake
by cells that also have negatively charged surface) and instability in the
blood circulation (non-
modified siRNAs have a very short half-life in blood stream, mostly because of
degradation by
nucleases)4. These impediments can be overcome by using polymeric or lipid-
based carriers to
shield the negative charge and provide protection against nuclease activity15-
17.
[0003] Complexation of the anionic carboxyfluorescein (CF) with single
headed
amphiphiles of opposite charge in cationic vesicles, formed by mixing single-
tailed cationic and
anionic surfactants has been reported (Danoff et al. 2007).
[0004] Furthermore, WO 02/055011 and WO 03/047499, both of the same
applicant,
disclose amphiphilic derivatives composed of at least one fatty acid chain
derived from natural
vegetable oils such as vernonia oil, lesquerella oil and castor oil, in which
functional groups such
as epoxy, hydroxy and double bonds were modified into polar and ionic
headgroups.
[0005] Additionally, WO 10/128504 discloses a series of amphiphiles and
bolamphiphiles (amphiphiles with two head groups) useful for targeted drug
delivery of insulin,
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insulin analogs, TNF, GDNF, DNA, RNA (including siRNA), enkephalin class of
analgesics, and
others.
[0006] These synthetic bolaamphiphiles (bolas) have recently been shown to
form
nanovesicles that interact with and encapsulate a variety of small and large
molecules including
peptides18' 19, proteins20 and plasmid DNAs19' 21 delivering them across
biological membranes22.
These bolaamphiphiles are a unique class of compounds that have two
hydrophilic headgroups
placed at each ends of a hydrophobic domain. Bolaamphiphiles can form vesicles
that consist of
monolayer membrane that surrounds an aqueous core. Vesicles made from natural
bolaamphiphiles, such as those extracted from archaebacteria (archaesomes),
are very stable and,
therefore, might be employed for targeted drug delivery. However,
bolaamphiphiles from
archaebacteria are heterogeneous and cannot be easily extracted or chemically
synthesized.
Furthermore, bolas have a hydrophobic alkyl chain connected to positively
charged head groups,
that can potentially interact with negatively charged nucleic acids and
promote their delivery into
cells. However, the nature of these interactions as well as the possibility to
use bolas for
optimized delivery of therapeutic siRNAs remains a challenge.
[0007] Thus, there remains a need to make new specific bolaamphiphiles
which can be
useful for optimized delivery of siRNAs into cells and have desired
therapeutic utility. The
compounds, compositions, and methods described herein are directed toward this
end.
SUMMARY OF THE INVENTION
[0008] In certain aspects, provided herein are pharmaceutical compositions
comprising of
complexes between bolaamphiphiles and pharmacologically or biologically active
compounds.
[0009] In certain aspects, the bolaamphiphile vesicle complexes comprise
one or more
bolaamphiphilic compounds and the biologically active compound is siRNA.
[0010] In certain aspects, the bolaamphiphile vesicle complexes comprise
one or more
bolaamphiphilic compounds and the biologically active compound is a siRNA that
is a mixture of
two or more siRNA, wherein at least one siRNA is directed to a first target,
and at least one
siRNA is directed to a second target.
[0011] In further aspects, provided herein are novel siRNA and
bolamphiphilic vesicle
complex comprising siRNA and one or more bolaamphiphilic compounds.
[0012] In further aspects, provided herein are novel formulations of siRNA
with
bolaamphiphilic compounds or with bolaamhphilic vesicles.
[0013] In another aspect, provided here are methods of delivering siRNA
into animal or
human cells.
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[0014] In an additional aspect, this present disclosure is directed to
delivery of
siRNA-bolaamphiphile vesicle complexes or siRNA-bolaamphiphilic vesicle
complexes into
animals or human wherein the bolaamphiphile vesicle complex comprises one or
more
bolaamphiphilic compounds and siRNA.
[0015] In another aspect, provided herein are methods of delivering siRNA
into animal or
human cell comprising the step of administering to the animal or human a
pharmaceutical
composition comprising of a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA. In
one
embodiment, the cell is brain cell, liver cell, gall bladder, or a lung cell.
In other embodiments,
the cells are are cells of a lymph node, a CD4+ lymphocyte, or a cell of the
mononuclear
phagocyte system, including, without limitation, a monocyte, macrophage, a
resident brain
microglial cell and a dendritic cell. In a still further emobidment, the cell
is a cancer cell.
[0016] In another aspect, provided here are methods of delivering siRNA
into animal or
human organs comprising the step of administering to the animal or human a
pharmaceutical
composition comprising of a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA. In
one
embodiment, the organ is brain, liver, gall bladder, a lymph node or a lung.
In certain aspects of
this emobidment, the siRNA is delivered to a tumor.
[0017] In a further embodiment the active agent is an RNA-DNA heteroduplex
with
properties of siRNA molecules. In certain aspects of this embodiment, the
bolaamphiphile
vesicle complexes comprise one or more bolaamphiphilic compounds and the
biologically active
compound is a siRNA that is a mixture of two or more siRNA or a mixture
comprising at least
one siRNA and one RNA-DNA duplex, wherein at least one siRNA or RNA-DNA duplex
is
directed to a first target, and at least one siRNA or RNA-DNA duplex is
directed to a second
target.
[0018] In certain embodiments, the target is a promoter. In other
emodiments, the first
and second targets are sequences of separate and distinct genes.
[0019] In one embodiment, the bolaamphiphilic compound consists of two
hydrophilic
headgroups linked through a long hydrophobic chain. In another embodiment, the
hydrophilic
headgroup is an amino containing group. In a specific embodiment, the
hydrophilic headgroup is
a tertiary or quaternary amino containing group.
[0020] In one particular embodiment, the bolaamphiphilic compound is a
compound
according to formula I:
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HG2 -L1-HG1
or a pharmaceutically acceptable salt, solvate, hydrate, prodrug,
stereoisomer, tautomer, isotopic
variant, or N¨oxide thereof, or a combination thereof;
wherein:
each HG1 and HG2 is independently a hydrophilic head group; and
L1 is alkylene, alkenyl, heteroalkylene, or heteroalkenyl linker;
unsubstituted or
substituted with Ci-C20 alkyl, hydroxyl, or oxo.
[0021] In one embodiment, the pharmaceutically acceptable salt is a
quaternary
ammonium salt.
[0022] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
the bolaamphiphilic compound is a compound according to formula II, III, IV,
V, or VI:
¨)nio Z2
HG2 __________________________ r _______________________ HG1
0 0
0 0
HG2 __________________________________________ f
22 ______________________________________________________ HG1
III
HG2_oRla Rlb
f'Ll0
R2a( Z1 Z2 R2b
IV
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HG2 ¨ 0 R1 a
0
.... .-Nn10
R2a ( in8 ( ')n9 Zi 'Z2 ¨HG1
V ,or
HG 2 ¨ 0 R1 a
0
R2a ( )1-1--8 - Th( )n9 Zi
VI
or a pharmaceutically acceptable salt, solvate, hydrate, prodrug,
stereoisomer, tautomer, isotopic
variant, or N¨oxide thereof, or a combination thereof;
wherein:
each HG1 and HG2 is independently a hydrophilic head group;
each Z1 and Z2 is independently -C(R3)2-, -N(R3)- or ¨0-;
¨ ib,
each Ria, I(R3, and R4 is independently H or Ci-C8 alkyl;
each R2a and R2b is independently H, Ci-Cs alkyl, OH, alkoxy, or 0-HG1 or 0-
HG2;
each n8, n9, n11, and n12 is independently an integer from 1-20;
n10 is an integer from 2-20; and
each dotted bond is independently a single or a double bond.
[0023] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, or VI, each HG1 and HG2 is independently selected from:
0 0 0 R8
( ')FI,1 /
X ( ')FI,1 X
,
0
R8 X
)riil 40
X and
)n13 0 0
wherein:
X is ¨NR5aR5b, or ¨N+R5aR5bR5e; each R5a, and R5b is independently H or
substituted or
unsubstituted C1-C20 alkyl or R5a and R5b may join together to form an N
containing
substituted or unsubstituted heteroaryl, or substituted or unsubstituted
heterocyclyl;
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each R5e is independently substituted or unsubstituted Ci-C20 alkyl; each R8
is independently
H, substituted or unsubstituted Ci-C20 alkyl, alkoxy, or carboxy;
ml is 0 or 1; and
each n13, n14, and n15 is independently an integer from 1-20.
[0024] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
the bolaamphiphilic compound is selected from the bolaambphilic compounds
listed in Table 1,
and wherein the compound ID is GLH-7, GLH-9, GLH-10, GLH-11, GLH-14, GLH-15,
GLH-
17, GLH-18, GLH-22, GLH-23, GLH-24, GLH-25, GLH-27, GLH-28, GLH-30 to GLH-48,
GLH-55, GLH-56, or GLH-57.
[0025] Other objects and advantages will become apparent to those skilled
in the art from
a consideration of the ensuing detailed description.
FIGURES
[0026] Figure 1: Double stranded siRNA and fluorescently labeled siRNA used
for the in
vitro and the in vivo experiments
[0027] Figure 2: Transfection of FITC-siRNA into dendritic cells and
silencing of HSP60
gene in these cells by bolaamphiphilic vesicles with siRNA. Human primary
dendritic cells
(obtained by differentiating human monocytes by the cytokines IL-4 and GM-CSF)
were exposed
to GLH-19 vesicles with FITC-siRNA for 5 hours and cells were observed by a
fluorescence
microscope and examined by flow cytometry. The fluorescence micrographs show
that all the
cells became fluorescent after exposing them for 5 hours to bolaamphiphilic
vesicles that
contained FITC-siRNA whereas only few cells were fluorescent after
transfecting them with the
same concentration of FITC-siRNA by electroporation. These results were
confirmed by flow
cytometry studies shown in the lower part of the Figure. When the dendritic
cells were exposed
to GLH-19 vesicles with specific siRNA for HSP60, a very significant silencing
of the gene was
seen on a western blot compared to cells treated with empty vesicles (control
cells). (V-smart
vesicles - bolaamphiphilic vesicles).
[0028] Figure 3: Silencing of GFP gene in stably transfected macorophages
by GFP
siRNA. Macrophages cell line from a mouse that stably express eGFP were
exposed to GLH-19
vesicles containg eGFP- siRNA. Not all the cells expressed the GFP as can be
seen from
comparison of the phase contrast micrograph to the fluorescence micrograph of
untreated cells
(lower right micrograph and upper right micrographs, respectively). Yet, the
eGFP fluorescence
in the cells that expressed the GFP gene disappeared almost completely when
the cells were
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treated with GLH-19 vesicles containing eGFP-siRNA (lower left micrograph),
whereas in cells
that were treated with empty vesicles all the cells that expressed the eGFP
remained fluorescent
(upper left micrograph) (V-smart vesicles - bolaamphiphilic vesicles).
[0029] Figure 4: Silencing of eGFP in MDA-MB-231/GFP cell line treated with
GLH-19
vesicles containing eGFP-siRNA. The breast cancer cell line MDA-MB-231 that
stably express
eGFP were exposed for 5 hours to GLH-19 vesicles containing egFP-siRNA. 72
hours after the
exposure a very significant silencing of the eGFP gene was observed. No
silencing was seen in
cells exposed to empty vesicles (not shown).
[0030] Figure 5: Biodistribution of siRNA-AF555 in organs from mice after
i.v.
administration of bolavesicles containing siRNA-AF555. GLH-19 vesicles
containing AF555-
siRNA were injected via the tail vein into mice and 30 minutes later mice were
acrificed, organs
collected and imaging for AF-555 fluorescence was performed. Bakground
fluorescence was
adjusted to show brown color and AF-555 fluorescence was adjusted to give
green color. AF-555
fluorescence is seen only in gall bladder (left image), blood vessels (around
the lung, see middle
image) and thoughout the brain (right image). No fluorescence is seen in the
liver at the 30 min
time point nor in the lung.
DEFINITIONS
Chemical Definitions
[0031] Definitions of specific functional groups and chemical terms are
described in
more detail below. The chemical elements are identified in accordance with the
Periodic Table of
the Elements, CAS version, Handbook of Chemistry and Physics, 75th ¨d.,
E inside cover, and
specific functional groups are generally defined as described therein.
Additionally, general
principles of organic chemistry, as well as specific functional moieties and
reactivity, are
described in Thomas Sorrell, Organic Chemistry, University Science Books,
Sausalito, 1999;
Smith and March, March's Advanced Organic Chemistry, 5th Edition, John Wiley &
Sons, Inc.,
New York, 2001; Larock, Comprehensive Organic Transformations, VCH Publishers,
Inc., New
York, 1989; and Carruthers, Some Modern Methods of Organic Synthesis, 3rd
Edition, Cambridge
University Press, Cambridge, 1987.
[0032] Compounds described herein can comprise one or more asymmetric
centers, and
thus can exist in various isomeric forms, e.g., enantiomers and/or
diastereomers. For example,
the compounds described herein can be in the form of an individual enantiomer,
diastereomer or
geometric isomer, or can be in the form of a mixture of stereoisomers,
including racemic
mixtures and mixtures enriched in one or more stereoisomer. Isomers can be
isolated from
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mixtures by methods known to those skilled in the art, including chiral high
pressure liquid
chromatography (HPLC) and the formation and crystallization of chiral salts;
or preferred
isomers can be prepared by asymmetric syntheses. See, for example, Jacques et
al.,
Enantiomers, Racemates and Resolutions (Wiley Interscience, New York, 1981);
Wilen et al.,
Tetrahedron 33:2725 (1977); Eliel, Stereochemistry of Carbon Compounds
(McGraw¨Hill, NY,
1962); and Wilen, Tables of Resolving Agents and Optical Resolutions p. 268
(E.L. Eliel, Ed.,
Univ. of Notre Dame Press, Notre Dame, IN 1972). The invention additionally
encompasses
compounds described herein as individual isomers substantially free of other
isomers, and
alternatively, as mixtures of various isomers.
[0033] When a range of values is listed, it is intended to encompass each
value and sub¨
range within the range. For example "Ci_6 alkyl" is intended to encompass, Ci,
C2, C3, C4, C5,
C6, C1-6, C1-5, C1-4, C1-3, C1-2, C2-6, C2-5, C2-4, C2-3, C3-6, C3-5, C3-4, C4-
6, C4-5, and C5-6 alkyl.
[0034] The following terms are intended to have the meanings presented
therewith below
and are useful in understanding the description and intended scope of the
present invention.
When describing the invention, which may include compounds, pharmaceutical
compositions
containing such compounds and methods of using such compounds and
compositions, the
following terms, if present, have the following meanings unless otherwise
indicated. It should
also be understood that when described herein any of the moieties defined
forth below may be
substituted with a variety of substituents, and that the respective
definitions are intended to
include such substituted moieties within their scope as set out below. Unless
otherwise stated, the
term "substituted" is to be defined as set out below. It should be further
understood that the
terms "groups" and "radicals" can be considered interchangeable when used
herein. The articles
"a" and "an" may be used herein to refer to one or to more than one (i.e. at
least one) of the
grammatical objects of the article. By way of example "an analogue" means one
analogue or
more than one analogue.
[0035] "Alkyl" refers to a radical of a straight¨chain or branched
saturated hydrocarbon
group having from 1 to 20 carbon atoms ("C1_20 alkyl"). In some embodiments,
an alkyl group
has 1 to 12 carbon atoms ("C1_12 alkyl"). In some embodiments, an alkyl group
has 1 to 10
carbon atoms ("C1_10 alkyl"). In some embodiments, an alkyl group has 1 to 9
carbon atoms
("C1_9 alkyl"). In some embodiments, an alkyl group has 1 to 8 carbon atoms
("C1_8 alkyl"). In
some embodiments, an alkyl group has 1 to 7 carbon atoms ("C1_7 alkyl"). In
some
embodiments, an alkyl group has 1 to 6 carbon atoms ("C1_6 alkyl", also
referred to herein as
"lower alkyl"). In some embodiments, an alkyl group has 1 to 5 carbon atoms
("C1_5 alkyl"). In
some embodiments, an alkyl group has 1 to 4 carbon atoms ("C1_4 alkyl"). In
some
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embodiments, an alkyl group has 1 to 3 carbon atoms ("C1_3 alkyl"). In some
embodiments, an
alkyl group has 1 to 2 carbon atoms ("C1_2 alkyl"). In some embodiments, an
alkyl group has 1
carbon atom ("C1 alkyl"). In some embodiments, an alkyl group has 2 to 6
carbon atoms ("C2_6
alkyl"). Examples of Ci_6 alkyl groups include methyl (C1), ethyl (C2),
n¨propyl (C3), isopropyl
(C3), n¨butyl (C4), tert¨butyl (C4), sec¨butyl (C4), iso¨butyl (C4), n¨pentyl
(C5), 3¨pentanyl (C5),
amyl (C5), neopentyl (C5), 3¨methyl-2¨butanyl (C5), tertiary amyl (C5), and
n¨hexyl (C6).
Additional examples of alkyl groups include n¨heptyl (C2), n¨octyl (C8) and
the like. Unless
otherwise specified, each instance of an alkyl group is independently
optionally substituted, i.e.,
unsubstituted (an "unsubstituted alkyl") or substituted (a "substituted
alkyl") with one or more
substituents; e.g., for instance from 1 to 5 substituents, 1 to 3
substituents, or 1 substituent. In
certain embodiments, the alkyl group is unsubstituted C1_10 alkyl (e.g.,
¨CH3). In certain
embodiments, the alkyl group is substituted C1_10 alkyl.
[0036] "Alkylene" refers to a substituted or unsubstituted alkyl group, as
defined above,
wherein two hydrogens are removed to provide a divalent radical. Exemplary
divalent alkylene
groups include, but are not limited to, methylene (-CH2-), ethylene (-CH2CH2-
), the propylene
isomers (e.g., -CH2CH2CH2- and -CH(CH3)CH2-) and the like.
[0037] "Alkenyl" refers to a radical of a straight¨chain or branched
hydrocarbon group
having from 2 to 20 carbon atoms, one or more carbon¨carbon double bonds, and
no triple bonds
("C2_20 alkenyl"). In some embodiments, an alkenyl group has 2 to 10 carbon
atoms ("C2_10
alkenyl"). In some embodiments, an alkenyl group has 2 to 9 carbon atoms
("C2_9 alkenyl"). In
some embodiments, an alkenyl group has 2 to 8 carbon atoms ("C2_8 alkenyl").
In some
embodiments, an alkenyl group has 2 to 7 carbon atoms ("C2_2 alkenyl"). In
some embodiments,
an alkenyl group has 2 to 6 carbon atoms ("C2_6 alkenyl"). In some
embodiments, an alkenyl
group has 2 to 5 carbon atoms ("C2_5 alkenyl"). In some embodiments, an
alkenyl group has 2 to
4 carbon atoms ("C2_4 alkenyl"). In some embodiments, an alkenyl group has 2
to 3 carbon
atoms ("C2_3 alkenyl"). In some embodiments, an alkenyl group has 2 carbon
atoms ("C2
alkenyl"). The one or more carbon¨carbon double bonds can be internal (such as
in 2¨butenyl)
or terminal (such as in 1¨buteny1). Examples of C2_4 alkenyl groups include
ethenyl (C2), 1¨
propenyl (C3), 2¨propenyl (C3), 1¨butenyl (C4), 2¨butenyl (C4), butadienyl
(C4), and the like.
Examples of C2_6 alkenyl groups include the aforementioned C2_4 alkenyl groups
as well as
pentenyl (C5), pentadienyl (C5), hexenyl (C6), and the like. Additional
examples of alkenyl
include heptenyl (C2), octenyl (C8), octatrienyl (C8), and the like. Unless
otherwise specified,
each instance of an alkenyl group is independently optionally substituted,
i.e., unsubstituted (an
"unsubstituted alkenyl") or substituted (a "substituted alkenyl") with one or
more substituents
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e.g., for instance from 1 to 5 substituents, 1 to 3 substituents, or 1
substituent. In certain
embodiments, the alkenyl group is unsubstituted C2_10 alkenyl. In certain
embodiments, the
alkenyl group is substituted C2_10 alkenyl.
[0038] "Alkenylene" refers a substituted or unsubstituted alkenyl group, as
defined
above, wherein two hydrogens are removed to provide a divalent radical.
Exemplary divalent
alkenylene groups include, but are not limited to, ethenylene (-CH=CH-),
propenylenes (e.g., -
CH=CHCH2- and -C(CH3)=CH- and -CH=C(CH3)-) and the like.
[0039] "Alkynyl" refers to a radical of a straight¨chain or branched
hydrocarbon group
having from 2 to 20 carbon atoms, one or more carbon¨carbon triple bonds, and
optionally one or
more double bonds ("C2_20 alkynyl"). In some embodiments, an alkynyl group has
2 to 10
carbon atoms ("C2_10 alkynyl"). In some embodiments, an alkynyl group has 2 to
9 carbon atoms
("C2_9 alkynyl"). In some embodiments, an alkynyl group has 2 to 8 carbon
atoms ("C2-8
alkynyl"). In some embodiments, an alkynyl group has 2 to 7 carbon atoms
("C2_7 alkynyl"). In
some embodiments, an alkynyl group has 2 to 6 carbon atoms ("C2_6 alkynyl").
In some
embodiments, an alkynyl group has 2 to 5 carbon atoms ("C2_5 alkynyl"). In
some embodiments,
an alkynyl group has 2 to 4 carbon atoms ("C2_4 alkynyl"). In some
embodiments, an alkynyl
group has 2 to 3 carbon atoms ("C2_3 alkynyl"). In some embodiments, an
alkynyl group has 2
carbon atoms ("C2 alkynyl"). The one or more carbon¨carbon triple bonds can be
internal (such
as in 2¨butynyl) or terminal (such as in 1¨butyny1). Examples of C2_4 alkynyl
groups include,
without limitation, ethynyl (C2), 1¨propynyl (C3), 2¨propynyl (C3), 1¨butynyl
(C4), 2¨butynyl
(C4), and the like. Examples of C2_6 alkenyl groups include the aforementioned
C2_4 alkynyl
groups as well as pentynyl (C5), hexynyl (C6), and the like. Additional
examples of alkynyl
include heptynyl (C7), octynyl (C8), and the like. Unless otherwise specified,
each instance of an
alkynyl group is independently optionally substituted, i.e., unsubstituted (an
"unsubstituted
alkynyl") or substituted (a "substituted alkynyl") with one or more
substituents; e.g., for instance
from 1 to 5 substituents, 1 to 3 substituents, or 1 substituent. In certain
embodiments, the alkynyl
group is unsubstituted C2_1() alkynyl. In certain embodiments, the alkynyl
group is substituted
C2-10 alkynyl.
[0040] "Alkynylene" refers a substituted or unsubstituted alkynyl group, as
defined
above, wherein two hydrogens are removed to provide a divalent radical.
Exemplary divalent
alkynylene groups include, but are not limited to, ethynylene, propynylene,
and the like.
[0041] "Aryl" refers to a radical of a monocyclic or polycyclic (e.g.,
bicyclic or tricyclic)
4n+2 aromatic ring system (e.g., having 6, 10, or 14 it electrons shared in a
cyclic array) having
6-14 ring carbon atoms and zero heteroatoms provided in the aromatic ring
system ("C6_14 aryl").
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In some embodiments, an aryl group has six ring carbon atoms ("C6 aryl"; e.g.,
phenyl). In some
embodiments, an aryl group has ten ring carbon atoms ("C10 aryl"; e.g.,
naphthyl such as 1¨
naphthyl and 2¨naphthyl). In some embodiments, an aryl group has fourteen ring
carbon atoms
("C14 aryl"; e.g., anthracyl). "Aryl" also includes ring systems wherein the
aryl ring, as defined
above, is fused with one or more carbocyclyl or heterocyclyl groups wherein
the radical or point
of attachment is on the aryl ring, and in such instances, the number of carbon
atoms continue to
designate the number of carbon atoms in the aryl ring system. Typical aryl
groups include, but
are not limited to, groups derived from aceanthrylene, acenaphthylene,
acephenanthrylene,
anthracene, azulene, benzene, chrysene, coronene, fluoranthene, fluorene,
hexacene, hexaphene,
hexalene, as-indacene, s-indacene, indane, indene, naphthalene, octacene,
octaphene, octalene,
ovalene, penta-2,4-diene, pentacene, pentalene, pentaphene, perylene,
phenalene, phenanthrene,
picene, pleiadene, pyrene, pyranthrene, rubicene, triphenylene, and
trinaphthalene. Particularly
aryl groups include phenyl, naphthyl, indenyl, and tetrahydronaphthyl. Unless
otherwise
specified, each instance of an aryl group is independently optionally
substituted, i.e.,
unsubstituted (an "unsubstituted aryl") or substituted (a "substituted aryl")
with one or more
substituents. In certain embodiments, the aryl group is unsubstituted C6_14
aryl. In certain
embodiments, the aryl group is substituted C6_14 aryl.
[0042] In certain embodiments, an aryl group substituted with one or more
of groups
selected from halo, C1-C8 alkyl, C1-C8 haloalkyl, cyano, hydroxy, C1-C8
alkoxy, and amino.
[0043] Examples of representative substituted aryls include the following
R56
R57
R56 and e R56
, 01
R57 R57 =
In these formulae one of R56 and R57 may be hydrogen and at least one of R56
and R57 is each
independently selected from C1-C8 alkyl, C1-C8 haloalkyl, 4-10 membered
heterocyclyl,
alkanoyl, C1-C8 alkoxy, heteroaryloxy, alkylamino, arylamino, heteroarylamino,
NR58C0R59,
NR58S0R59NR58S02R59, COOalkyl, COOaryl, C0NR58R59, C0NR580R59, NR58R59,
S02NR58R59, S-alkyl, SOalkyl, SO2alkyl, Saryl, SOaryl, SO2aryl; or R56 and R57
may be joined to
form a cyclic ring (saturated or unsaturated) from 5 to 8 atoms, optionally
containing one or more
heteroatoms selected from the group N, 0, or S. R6 and R61 are independently
hydrogen, C1-C8
alkyl, Ci-C4 haloalkyl, C3-C10 cycloalkyl, 4-10 membered heterocyclyl, C6-C10
aryl, substituted
C6-C10 aryl, 5-10 membered heteroaryl, or substituted 5-10 membered
heteroaryl.
[0044] "Fused aryl" refers to an aryl having two of its ring carbon in
common with a
second aryl ring or with an aliphatic ring.
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[0045] "Aralkyl" is a subset of alkyl and aryl, as defined herein, and
refers to an
optionally substituted alkyl group substituted by an optionally substituted
aryl group.
[0046] "Heteroaryl" refers to a radical of a 5-10 membered monocyclic or
bicyclic 4n+2
aromatic ring system (e.g., having 6 or 10 it electrons shared in a cyclic
array) having ring carbon
atoms and 1-4 ring heteroatoms provided in the aromatic ring system, wherein
each heteroatom
is independently selected from nitrogen, oxygen and sulfur ("5-10 membered
heteroaryl"). In
heteroaryl groups that contain one or more nitrogen atoms, the point of
attachment can be a
carbon or nitrogen atom, as valency permits. Heteroaryl bicyclic ring systems
can include one or
more heteroatoms in one or both rings. "Heteroaryl" includes ring systems
wherein the
heteroaryl ring, as defined above, is fused with one or more carbocyclyl or
heterocyclyl groups
wherein the point of attachment is on the heteroaryl ring, and in such
instances, the number of
ring members continue to designate the number of ring members in the
heteroaryl ring system.
"Heteroaryl" also includes ring systems wherein the heteroaryl ring, as
defined above, is fused
with one or more aryl groups wherein the point of attachment is either on the
aryl or heteroaryl
ring, and in such instances, the number of ring members designates the number
of ring members
in the fused (aryl/heteroaryl) ring system. Bicyclic heteroaryl groups wherein
one ring does not
contain a heteroatom (e.g., indolyl, quinolinyl, carbazolyl, and the like) the
point of attachment
can be on either ring, i.e., either the ring bearing a heteroatom (e.g.,
2¨indoly1) or the ring that
does not contain a heteroatom (e.g., 5¨indoly1).
[0047] In some embodiments, a heteroaryl group is a 5-10 membered aromatic
ring
system having ring carbon atoms and 1-4 ring heteroatoms provided in the
aromatic ring system,
wherein each heteroatom is independently selected from nitrogen, oxygen, and
sulfur ("5-10
membered heteroaryl"). In some embodiments, a heteroaryl group is a 5-8
membered aromatic
ring system having ring carbon atoms and 1-4 ring heteroatoms provided in the
aromatic ring
system, wherein each heteroatom is independently selected from nitrogen,
oxygen, and sulfur
("5-8 membered heteroaryl"). In some embodiments, a heteroaryl group is a 5-6
membered
aromatic ring system having ring carbon atoms and 1-4 ring heteroatoms
provided in the
aromatic ring system, wherein each heteroatom is independently selected from
nitrogen, oxygen,
and sulfur ("5-6 membered heteroaryl"). In some embodiments, the 5-6 membered
heteroaryl
has 1-3 ring heteroatoms selected from nitrogen, oxygen, and sulfur. In some
embodiments, the
5-6 membered heteroaryl has 1-2 ring heteroatoms selected from nitrogen,
oxygen, and sulfur.
In some embodiments, the 5-6 membered heteroaryl has 1 ring heteroatom
selected from
nitrogen, oxygen, and sulfur. Unless otherwise specified, each instance of a
heteroaryl group is
independently optionally substituted, i.e., unsubstituted (an "unsubstituted
heteroaryl") or
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substituted (a "substituted heteroaryl") with one or more substituents. In
certain embodiments,
the heteroaryl group is unsubstituted 5-14 membered heteroaryl. In certain
embodiments, the
heteroaryl group is substituted 5-14 membered heteroaryl.
[0048] Exemplary 5¨membered heteroaryl groups containing one heteroatom
include,
without limitation, pyrrolyl, furanyl and thiophenyl. Exemplary 5¨membered
heteroaryl groups
containing two heteroatoms include, without limitation, imidazolyl, pyrazolyl,
oxazolyl,
isoxazolyl, thiazolyl, and isothiazolyl. Exemplary 5¨membered heteroaryl
groups containing
three heteroatoms include, without limitation, triazolyl, oxadiazolyl, and
thiadiazolyl. Exemplary
5¨membered heteroaryl groups containing four heteroatoms include, without
limitation,
tetrazolyl. Exemplary 6¨membered heteroaryl groups containing one heteroatom
include,
without limitation, pyridinyl. Exemplary 6¨membered heteroaryl groups
containing two
heteroatoms include, without limitation, pyridazinyl, pyrimidinyl, and
pyrazinyl. Exemplary 6¨
membered heteroaryl groups containing three or four heteroatoms include,
without limitation,
triazinyl and tetrazinyl, respectively. Exemplary 7¨membered heteroaryl groups
containing one
heteroatom include, without limitation, azepinyl, oxepinyl, and thiepinyl.
Exemplary 5,6¨
bicyclic heteroaryl groups include, without limitation, indolyl, isoindolyl,
indazolyl,
benzotriazolyl, benzothiophenyl, isobenzothiophenyl, benzofuranyl,
benzoisofuranyl,
benzimidazolyl, benzoxazolyl, benzisoxazolyl, benzoxadiazolyl, benzthiazolyl,
benzisothiazolyl,
benzthiadiazolyl, indolizinyl, and purinyl. Exemplary 6,6¨bicyclic heteroaryl
groups include,
without limitation, naphthyridinyl, pteridinyl, quinolinyl, isoquinolinyl,
cinnolinyl, quinoxalinyl,
phthalazinyl, and quinazolinyl.
[0049] Examples of representative heteroaryls include the following:
0 & ,\\NJ rN,J, fr\\i,,,\ r' 1
....., ,i., N,... ,..,..... ..,
Y Y N N N
N
I I
(
I 0
e
N / l N%
N N
N
i =N 0\
,
N
1110 \
( 0
Y Y Y
N
wherein each Y is selected from carbonyl, N, NR65, 0, and S; and R65 is
independently hydrogen,
C1-C8 alkyl, C3-C10 cycloalkyl, 4-10 membered heterocyclyl, C6-Cio aryl, and 5-
10 membered
heteroaryl.
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[0050] Examples of representative aryl having hetero atoms containing
substitution
include the following:
w w
. w) 401 Y and . Y>
Y' ,
wherein each W is selected from C(R66)2, NR66, 0, and S; and each Y is
selected from carbonyl,
NR66, 0 and S; and R66 is independently hydrogen, Ci-C8 alkyl, C3-Cio
cycloalkyl, 4-10
membered heterocyclyl, C6-C10 aryl, and 5-10 membered heteroaryl.
[0051] "Heteroaralkyl" is a subset of alkyl and heteroaryl, as defined
herein, and refers to
an optionally substituted alkyl group substituted by an optionally substituted
heteroaryl group.
[0052] "Carbocycly1" or "carbocyclic" refers to a radical of a non¨aromatic
cyclic
hydrocarbon group having from 3 to 10 ring carbon atoms ("C3_10 carbocyclyl")
and zero
heteroatoms in the non¨aromatic ring system. In some embodiments, a
carbocyclyl group has 3
to 8 ring carbon atoms ("C3_8 carbocyclyl"). In some embodiments, a
carbocyclyl group has 3 to
6 ring carbon atoms ("C3_6 carbocyclyl"). In some embodiments, a carbocyclyl
group has 3 to 6
ring carbon atoms ("C3_6 carbocyclyl"). In some embodiments, a carbocyclyl
group has 5 to 10
ring carbon atoms ("C5_10 carbocyclyl"). Exemplary C3_6 carbocyclyl groups
include, without
limitation, cyclopropyl (C3), cyclopropenyl (C3), cyclobutyl (C4),
cyclobutenyl (C4), cyclopentyl
(C5), cyclopentenyl (C5), cyclohexyl (C6), cyclohexenyl (C6), cyclohexadienyl
(C6), and the like.
Exemplary C3_8 carbocyclyl groups include, without limitation, the
aforementioned C3-6
carbocyclyl groups as well as cycloheptyl (C7), cycloheptenyl (C7),
cycloheptadienyl (C7),
cycloheptatrienyl (C7), cyclooctyl (C8), cyclooctenyl (C8), bicyclo[2.2.1
]heptanyl (C7),
bicyclo[2.2.2]octanyl (C8), and the like. Exemplary C3_10 carbocyclyl groups
include, without
limitation, the aforementioned C3_8 carbocyclyl groups as well as cyclononyl
(C9), cyclononenyl
(C9), cyclodecyl (C10), cyclodecenyl (C10), octahydro-1H¨indenyl (C9),
decahydronaphthalenyl
(C10), spiro[4.5]decanyl (C10), and the like. As the foregoing examples
illustrate, in certain
embodiments, the carbocyclyl group is either monocyclic ("monocyclic
carbocyclyl") or contain
a fused, bridged or spiro ring system such as a bicyclic system ("bicyclic
carbocyclyl") and can
be saturated or can be partially unsaturated. "Carbocycly1" also includes ring
systems wherein the
carbocyclyl ring, as defined above, is fused with one or more aryl or
heteroaryl groups wherein
the point of attachment is on the carbocyclyl ring, and in such instances, the
number of carbons
continue to designate the number of carbons in the carbocyclic ring system.
Unless otherwise
specified, each instance of a carbocyclyl group is independently optionally
substituted, i.e.,
unsubstituted (an "unsubstituted carbocyclyl") or substituted (a "substituted
carbocyclyl") with
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one or more substituents. In certain embodiments, the carbocyclyl group is
unsubstituted C3-10
carbocyclyl. In certain embodiments, the carbocyclyl group is a substituted C3-
10 carbocyclyl.
[0053] In some embodiments, "carbocyclyl" is a monocyclic, saturated
carbocyclyl group
having from 3 to 10 ring carbon atoms ("C3_10 cycloalkyl"). In some
embodiments, a cycloalkyl
group has 3 to 8 ring carbon atoms ("C3_8 cycloalkyl"). In some embodiments, a
cycloalkyl
group has 3 to 6 ring carbon atoms ("C3_6 cycloalkyl"). In some embodiments, a
cycloalkyl
group has 5 to 6 ring carbon atoms ("C5_6 cycloalkyl"). In some embodiments, a
cycloalkyl
group has 5 to 10 ring carbon atoms ("C5_10 cycloalkyl"). Examples of C5_6
cycloalkyl groups
include cyclopentyl (C5) and cyclohexyl (C5). Examples of C3_6 cycloalkyl
groups include the
aforementioned C5_6 cycloalkyl groups as well as cyclopropyl (C3) and
cyclobutyl (C4).
Examples of C3_8 cycloalkyl groups include the aforementioned C3_6 cycloalkyl
groups as well as
cycloheptyl (C7) and cyclooctyl (Cs). Unless otherwise specified, each
instance of a cycloalkyl
group is independently unsubstituted (an "unsubstituted cycloalkyl") or
substituted (a
"substituted cycloalkyl") with one or more substituents. In certain
embodiments, the cycloalkyl
group is unsubstituted C3_10 cycloalkyl. In certain embodiments, the
cycloalkyl group is
substituted C3_10 cycloalkyl.
[0054] "Heterocycly1" or "heterocyclic" refers to a radical of a 3¨ to
10¨membered non¨
aromatic ring system having ring carbon atoms and 1 to 4 ring heteroatoms,
wherein each
heteroatom is independently selected from nitrogen, oxygen, sulfur, boron,
phosphorus, and
silicon ("3-10 membered heterocyclyl"). In heterocyclyl groups that contain
one or more
nitrogen atoms, the point of attachment can be a carbon or nitrogen atom, as
valency permits. A
heterocyclyl group can either be monocyclic ("monocyclic heterocyclyl") or a
fused, bridged or
spiro ring system such as a bicyclic system ("bicyclic heterocyclyl"), and can
be saturated or can
be partially unsaturated. Heterocyclyl bicyclic ring systems can include one
or more heteroatoms
in one or both rings. "Heterocycly1" also includes ring systems wherein the
heterocyclyl ring, as
defined above, is fused with one or more carbocyclyl groups wherein the point
of attachment is
either on the carbocyclyl or heterocyclyl ring, or ring systems wherein the
heterocyclyl ring, as
defined above, is fused with one or more aryl or heteroaryl groups, wherein
the point of
attachment is on the heterocyclyl ring, and in such instances, the number of
ring members
continue to designate the number of ring members in the heterocyclyl ring
system. Unless
otherwise specified, each instance of heterocyclyl is independently optionally
substituted, i.e.,
unsubstituted (an "unsubstituted heterocyclyl") or substituted (a "substituted
heterocyclyl") with
one or more substituents. In certain embodiments, the heterocyclyl group is
unsubstituted 3-10
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membered heterocyclyl. In certain embodiments, the heterocyclyl group is
substituted 3-10
membered heterocyclyl.
[0055] In some embodiments, a heterocyclyl group is a 5-10 membered
non¨aromatic
ring system having ring carbon atoms and 1-4 ring heteroatoms, wherein each
heteroatom is
independently selected from nitrogen, oxygen, sulfur, boron, phosphorus, and
silicon ("5-10
membered heterocyclyl"). In some embodiments, a heterocyclyl group is a 5-8
membered non¨
aromatic ring system having ring carbon atoms and 1-4 ring heteroatoms,
wherein each
heteroatom is independently selected from nitrogen, oxygen, and sulfur ("5-8
membered
heterocyclyl"). In some embodiments, a heterocyclyl group is a 5-6 membered
non¨aromatic
ring system having ring carbon atoms and 1-4 ring heteroatoms, wherein each
heteroatom is
independently selected from nitrogen, oxygen, and sulfur ("5-6 membered
heterocyclyl"). In
some embodiments, the 5-6 membered heterocyclyl has 1-3 ring heteroatoms
selected from
nitrogen, oxygen, and sulfur. In some embodiments, the 5-6 membered
heterocyclyl has 1-2 ring
heteroatoms selected from nitrogen, oxygen, and sulfur. In some embodiments,
the 5-6
membered heterocyclyl has one ring heteroatom selected from nitrogen, oxygen,
and sulfur.
[0056] Exemplary 3¨membered heterocyclyl groups containing one heteroatom
include,
without limitation, azirdinyl, oxiranyl, thiorenyl. Exemplary 4¨membered
heterocyclyl groups
containing one heteroatom include, without limitation, azetidinyl, oxetanyl
and thietanyl.
Exemplary 5¨membered heterocyclyl groups containing one heteroatom include,
without
limitation, tetrahydrofuranyl, dihydrofuranyl, tetrahydrothiophenyl,
dihydrothiophenyl,
pyrrolidinyl, dihydropyrrolyl and pyrroly1-2,5¨dione. Exemplary 5¨membered
heterocyclyl
groups containing two heteroatoms include, without limitation, dioxolanyl,
oxasulfuranyl,
disulfuranyl, and oxazolidin-2-one. Exemplary 5¨membered heterocyclyl groups
containing three
heteroatoms include, without limitation, triazolinyl, oxadiazolinyl, and
thiadiazolinyl.
Exemplary 6¨membered heterocyclyl groups containing one heteroatom include,
without
limitation, piperidinyl, tetrahydropyranyl, dihydropyridinyl, and thianyl.
Exemplary 6¨
membered heterocyclyl groups containing two heteroatoms include, without
limitation,
piperazinyl, morpholinyl, dithianyl, dioxanyl. Exemplary 6¨membered
heterocyclyl groups
containing two heteroatoms include, without limitation, triazinanyl. Exemplary
7¨membered
heterocyclyl groups containing one heteroatom include, without limitation,
azepanyl, oxepanyl
and thiepanyl. Exemplary 8¨membered heterocyclyl groups containing one
heteroatom include,
without limitation, azocanyl, oxecanyl and thiocanyl. Exemplary 5-membered
heterocyclyl
groups fused to a C6 aryl ring (also referred to herein as a 5,6-bicyclic
heterocyclic ring) include,
without limitation, indolinyl, isoindolinyl, dihydrobenzofuranyl,
dihydrobenzothienyl,
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benzoxazolinonyl, and the like. Exemplary 6-membered heterocyclyl groups fused
to an aryl ring
(also referred to herein as a 6,6-bicyclic heterocyclic ring) include, without
limitation,
tetrahydroquinolinyl, tetrahydroisoquinolinyl, and the like.
[0057] Particular examples of heterocyclyl groups are shown in the
following illustrative
examples:
y) W) ,\X N-N 0 11\/,
Y
Y Y Y3
w
r ,i w/)
Y W N Y
-lil( a
W Y
[0058] wherein each W is selected from CR67, C(R67)2, NR67, 0, and S; and
each Y is
selected from NR67, 0, and S; and R67 is independently hydrogen, Ci-C8 alkyl,
C3-Cio cycloalkyl,
4-10 membered heterocyclyl, C6-Cio aryl, 5-10 membered heteroaryl. These
heterocyclyl rings
may be optionally substituted with one or more substituents selected from the
group consisting of
the group consisting of acyl, acylamino, acyloxy, alkoxy, alkoxycarbonyl,
alkoxycarbonylamino,
amino, substituted amino, aminocarbonyl (carbamoyl or amido),
aminocarbonylamino,
aminosulfonyl, sulfonylamino, aryl, aryloxy, azido, carboxyl, cyano,
cycloalkyl, halogen,
hydroxy, keto, nitro, thiol, -S-alkyl, ¨S-aryl, -S(0)-alkyl,¨S(0)-aryl, ¨S(0)2-
alkyl, and -S(0)2-
aryl. Substituting groups include carbonyl or thiocarbonyl which provide, for
example, lactam
and urea derivatives.
[0059] "Hetero" when used to describe a compound or a group present on a
compound
means that one or more carbon atoms in the compound or group have been
replaced by a
nitrogen, oxygen, or sulfur heteroatom. Hetero may be applied to any of the
hydrocarbyl groups
described above such as alkyl, e.g., heteroalkyl, cycloalkyl, e.g.,
heterocyclyl, aryl, e.g,.
heteroaryl, cycloalkenyl, e.g,. cycloheteroalkenyl, and the like having from 1
to 5, and
particularly from 1 to 3 heteroatoms.
[0060] "Acyl" refers to a radical -C(0)R20, where R2 is hydrogen,
substituted or
unsubstitued alkyl, substituted or unsubstitued alkenyl, substituted or
unsubstitued alkynyl,
substituted or unsubstitued carbocyclyl, substituted or unsubstituted
heterocyclyl, substituted or
unsubstituted aryl, or substituted or unsubstitued heteroaryl, as defined
herein. "Alkanoyl" is an
acyl group wherein R2 is a group other than hydrogen. Representative acyl
groups include, but
are not limited to, formyl (-CHO), acetyl (-C(=0)CH3), cyclohexylcarbonyl,
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cyclohexylmethylcarbonyl, benzoyl (-C(=0)Ph), benzylcarbonyl (-C(=0)CH2Ph),
¨C(0)-C1-C8
alkyl, ¨C(0)-(CH2)t(C6-C10 aryl), ¨C(0)-(CH2)t(5-10 membered heteroaryl),
¨C(0)-(CH2)t(C3-
Cio cycloalkyl), and ¨C(0)-(CH2)t(4-1 0 membered heterocyclyl), wherein t is
an integer from 0
to 4. In certain embodiments, R21 is C1-C8 alkyl, substituted with halo or
hydroxy; or C3-C10
cycloalkyl, 4-10 membered heterocyclyl, C6-Cio aryl, arylalkyl, 5-10 membered
heteroaryl or
heteroarylalkyl, each of which is substituted with unsubstituted C1-C4 alkyl,
halo, unsubstituted
C1-C4 alkoxy, unsubstituted Ci-C4 haloalkyl, unsubstituted Ci-C4 hydroxyalkyl,
or unsubstituted
Ci-C4 haloalkoxy or hydroxy.
[0061]
"Acylamino" refers to a radical -NR22c(0)R23, where each instance of R22 and
R23 is independently hydrogen, substituted or unsubstitued alkyl, substituted
or unsubstitued
alkenyl, substituted or unsubstitued alkynyl, substituted or unsubstitued
carbocyclyl, substituted
or unsubstituted heterocyclyl, substituted or unsubstituted aryl, or
substituted or unsubstitued
heteroarylõ as defined herein, or R22 is an amino protecting group. Exemplary
"acylamino"
groups include, but are not limited to, formylamino, acetylamino,
cyclohexylcarbonylamino,
cyclohexylmethyl-carbonylamino, benzoylamino and benzylcarbonylamino.
Particular
exemplary "acylamino" groups are ¨NR24C(0)-C1-C8 alkyl, ¨NR24C(0)-(CH2)t(C6-
C10 aryl), ¨
NR24C(0)-(CH2)t(5-10 membered heteroaryl), ¨NR24C(0)-(CH2)t(C3-C10
cycloalkyl), and ¨
NR24C(0)-(CH2)t(4-10 membered heterocyclyl), wherein t is an integer from 0 to
4, and each R24
independently represents H or Ci-C8 alkyl. In certain embodiments, R25 is H,
C1-C8 alkyl,
substituted with halo or hydroxy; C3-Cio cycloalkyl, 4-10 membered
heterocyclyl, C6-Cio aryl,
arylalkyl, 5-10 membered heteroaryl or heteroarylalkyl, each of which is
substituted with
unsubstituted C1-C4 alkyl, halo, unsubstituted C1-C4 alkoxy, unsubstituted Ci-
C4 haloalkyl,
unsubstituted C1-C4 hydroxyalkyl, or unsubstituted Ci-C4 haloalkoxy or
hydroxy; and R26 is H,
C1-C8 alkyl, substituted with halo or hydroxy;
C3-Cio cycloalkyl, 4-10 membered heterocyclyl, C6-Cio aryl, arylalkyl, 5-10
membered
heteroaryl or heteroarylalkyl, each of which is substituted with unsubstituted
CI-CI alkyl, halo,
unsubstituted C1-C4 alkoxy, unsubstituted CI-CI haloalkyl, unsubstituted CI-CI
hydroxyalkyl, or
unsubstituted C1_C4 haloalkoxy or hydroxyl; provided that at least one of R25
and R26 is other than
H.
[0062]
"Acyloxy" refers to a radical -0C(0)R27, where R27 is hydrogen, substituted or
unsubstitued alkyl, substituted or unsubstitued alkenyl, substituted or
unsubstituted alkynyl,
substituted or unsubstituted carbocyclyl, substituted or unsubstituted
heterocyclyl, substituted or
unsubstituted aryl, or substituted or unsubstituted heteroaryl, as defined
herein. Representative
examples include, but are not limited to, formyl, acetyl, cyclohexylcarbonyl,
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cyclohexylmethylcarbonyl, benzoyl and benzylcarbonyl. In certain embodiments,
R28 is Ci-Cs
alkyl, substituted with halo or hydroxy; C3-C10 cycloalkyl, 4-10 membered
heterocyclyl, C6-C10
aryl, arylalkyl, 5-10 membered heteroaryl or heteroarylalkyl, each of which is
substituted with
unsubstituted C1-C4 alkyl, halo, unsubstituted C1-C4 alkoxy, unsubstituted Ci-
C4 haloalkyl,
unsubstituted C1-C4 hydroxyalkyl, or unsubstituted Ci-C4 haloalkoxy or
hydroxy.
[0063] "Alkoxy" refers to the group ¨0R29 where R29 is substituted or
unsubstituted
alkyl, substituted or unsubstituted alkenyl, substituted or unsubstituted
alkynyl, substituted or
unsubstituted carbocyclyl, substituted or unsubstituted heterocyclyl,
substituted or unsubstituted
aryl, or substituted or unsubstituted heteroaryl. Particular alkoxy groups are
methoxy, ethoxy, n-
propoxy, isopropoxy, n-butoxy, tert-butoxy, sec-butoxy, n-pentoxy, n-hexoxy,
and 1,2-
dimethylbutoxy. Particular alkoxy groups are lower alkoxy, i.e. with between 1
and 6 carbon
atoms. Further particular alkoxy groups have between 1 and 4 carbon atoms.
[0064] In certain embodiments, R29 is a group that has 1 or more
substituents, for
instance, from 1 to 5 substituents, and particularly from 1 to 3 substituents,
in particular 1
substituent, selected from the group consisting of amino, substituted amino,
C6-Cio aryl, aryloxy,
carboxyl, cyano, C3-Cio cycloalkyl, 4-10 membered heterocyclyl, halogen, 5-10
membered
heteroaryl, hydroxyl, nitro, thioalkoxy, thioaryloxy, thiol, alkyl-S(0)-,
aryl¨S(0)-, alkyl¨S(0)2-
and aryl-S(0)2-. Exemplary 'substituted alkoxy' groups include, but are not
limited to, ¨0-
(CH2)t(C6-C10 aryl), ¨0-(CH2)(5-1 0 membered heteroaryl), ¨0-(CH2)t(C3-C10
cycloalkyl), and ¨
0-(CH2)(4-10 membered heterocyclyl), wherein t is an integer from 0 to 4 and
any aryl,
heteroaryl, cycloalkyl or heterocyclyl groups present, may themselves be
substituted by
unsubstituted C1-C4 alkyl, halo, unsubstituted C1-C4 alkoxy, unsubstituted Ci-
C4 haloalkyl,
unsubstituted C1-C4 hydroxyalkyl, or unsubstituted Ci-C4 haloalkoxy or
hydroxy. Particular
exemplary 'substituted alkoxy' groups are -0CF3, -OCH2CF3, -OCH2Ph, -OCH2-
cyclopropyl, -
OCH2CH2OH, and -OCH2CH2NMe2.
[0065] "Amino" refers to the radical -NH2.
[0066] "Substituted amino" refers to an amino group of the formula -N(R38)2
wherein R38
is hydrogen, substituted or unsubstituted alkyl, substituted or unsubstitued
alkenyl, substituted or
unsubstitued alkynyl, substituted or unsubstitued carbocyclyl, substituted or
unsubstituted
heterocyclyl, substituted or unsubstituted aryl, substituted or unsubstitued
heteroaryl, or an amino
protecting group, wherein at least one of R38 is not a hydrogen. In certain
embodiments,each R38
is independently selected from: hydrogen, Ci-C8 alkyl, C3-C8 alkenyl, C3-C8
alkynyl, C6-Cio aryl,
5-10 membered heteroaryl, 4-10 membered heterocyclyl, or C3-Cio cycloalkyl; or
C1-C8 alkyl,
substituted with halo or hydroxy; C3-C8 alkenyl, substituted with halo or
hydroxy; C3-C8 alkynyl,
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substituted with halo or hydroxy, or -(CH2)t(C6-C10 aryl), -(CH2)t(5-1 0
membered heteroaryl), -
(CH2)t(C3-C10 cycloalkyl), or -(CH2)t(4-1 0 membered heterocyclyl), wherein t
is an integer
between 0 and 8, each of which is substituted by unsubstituted CI-CI alkyl,
halo, unsubstituted
C1-C4 alkoxy, unsubstituted
haloalkyl, unsubstituted hydroxyalkyl, or unsubstituted
C1-C4 haloalkoxy or hydroxy; or both R38 groups are joined to form an alkylene
group.
[0067] Exemplary 'substituted amino' groups are ¨NR39-C1-C8 alkyl, ¨NR39-
(CH2)t(C6-
C10 aryl), ¨NR39-(CH2)t(5-10 membered heteroaryl), ¨NR39-(CH2)t(C3-C10
cycloalkyl), and ¨
NR39-(CH2)t(4-1 0 membered heterocyclyl), wherein t is an integer from 0 to 4,
for instance 1 or
2, each R39 independently represents H or CI-Cs alkyl, and any alkyl groups
present, may
themselves be substituted by halo, substituted or unsubstituted amino, or
hydroxy; and any aryl,
heteroaryl, cycloalkyl, or heterocyclyl groups present, may themselves be
substituted by
unsubstituted CI-CI alkyl, halo, unsubstituted alkoxy,
unsubstituted C1-C4 haloalkyl,
unsubstituted C1-C4 hydroxyalkyl, or unsubstituted haloalkoxy or hydroxy.
For the
avoidance of doubt the term 'substituted amino' includes the groups
alkylamino, substituted
alkylamino, alkylarylamino, substituted alkylarylamino, arylamino, substituted
arylamino,
dialkylamino, and substituted dialkylamino as defined below. Substituted amino
encompasses
both monosubstituted amino and disubstituted amino groups.
[0068] "Azido" refers to the radical -N3.
[0069] "Carbamoyl" or "amido" refers to the radical -C(0)NF12.
[0070] "Substituted carbamoyl" or "substituted amido" refers to the radical
-C(0)N(R62)2
wherein each R62 is independently hydrogen, substituted or unsubstituted
alkyl, substituted or
unsubstitued alkenyl, substituted or unsubstitued alkynyl, substituted or
unsubstitued carbocyclyl,
substituted or unsubstituted heterocyclyl, substituted or unsubstituted aryl,
substituted or
unsubstitued heteroaryl, or an amino protecting group, wherein at least one of
R62 is not a
hydrogen. In certain embodiments, R62 is selected from H, CI-Cs alkyl, C3-C10
cycloalkyl, 4-10
membered heterocyclyl, C6-C10 aryl, aralkyl, 5-10 membered heteroaryl, and
heteroaralkyl; or
C1-C8 alkyl substituted with halo or hydroxy; or C3-C10 cycloalkyl, 4-10
membered heterocyclyl,
C6-C10 aryl, aralkyl, 5-10 membered heteroaryl, or heteroaralkyl, each of
which is substituted by
unsubstituted C1-C4 alkyl, halo, unsubstituted C1-C4 alkoxy, unsubstituted
haloalkyl,
unsubstituted C1-C4 hydroxyalkyl, or unsubstituted haloalkoxy or hydroxy;
provided that
at least one R62 is other than H.
[0071] Exemplary 'substituted carbamoyl' groups include, but are not
limited to, ¨C(0)
NR64-C1-C8 alkyl, ¨C(0)NR64-(CH2)t(C6-C10 aryl), ¨C(0)N64-(CH2)t(5-1 0
membered heteroaryl),
¨C(0)NR64-(CH2)t(C3-Cto cycloalkyl), and ¨C(0)NR64-(CH2)t(4-10 membered
heterocyclyl),
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wherein t is an integer from 0 to 4, each R64 independently represents H or Ci-
C8 alkyl and any
aryl, heteroaryl, cycloalkyl or heterocyclyl groups present, may themselves be
substituted by
unsubstituted C1-C4 alkyl, halo, unsubstituted C1-C4 alkoxy, unsubstituted Ci-
C4 haloalkyl,
unsubstituted C1-C4 hydroxyalkyl, or unsubstituted Ci-C4 haloalkoxy or
hydroxy.
[0072] `Carboxy' refers to the radical -C(0)0H.
[0073] "Cyano" refers to the radical -CN.
[0074] "Halo" or "halogen" refers to fluoro (F), chloro (Cl), bromo (Br),
and iodo (I). In
certain embodiments, the halo group is either fluoro or chloro. In further
embodiments, the halo
group is iodo.
[0075] "Hydroxy" refers to the radical -OH.
[0076] "Nitro" refers to the radical ¨NO2.
[0077] "Cycloalkylalkyl" refers to an alkyl radical in which the alkyl
group is substituted
with a cycloalkyl group. Typical cycloalkylalkyl groups include, but are not
limited to,
cyclopropylmethyl, cyclobutylmethyl, cyclopentylmethyl, cyclohexylmethyl,
cycloheptylmethyl,
cyclooctylmethyl, cyclopropylethyl, cyclobutylethyl, cyclopentylethyl,
cyclohexylethyl,
cycloheptylethyl, and cyclooctylethyl, and the like.
[0078] "Heterocyclylalkyl" refers to an alkyl radical in which the alkyl
group is
substituted with a heterocyclyl group. Typical heterocyclylalkyl groups
include, but are not
limited to, pyrrolidinylmethyl, piperidinylmethyl, piperazinylmethyl,
morpholinylmethyl,
pyrrolidinylethyl, piperidinylethyl, piperazinylethyl, morpholinylethyl, and
the like.
[0079] "Cycloalkenyl" refers to substituted or unsubstituted carbocyclyl
group having
from 3 to 10 carbon atoms and having a single cyclic ring or multiple
condensed rings, including
fused and bridged ring systems and having at least one and particularly from 1
to 2 sites of
olefinic unsaturation. Such cycloalkenyl groups include, by way of example,
single ring
structures such as cyclohexenyl, cyclopentenyl, cyclopropenyl, and the like.
[0080] "Fused cycloalkenyl" refers to a cycloalkenyl having two of its
ring carbon atoms
in common with a second aliphatic or aromatic ring and having its olefinic
unsaturation located
to impart aromaticity to the cycloalkenyl ring.
[0081] "Ethenyl" refers to substituted or unsubstituted ¨(C=C)-.
[0082] "Ethylene" refers to substituted or unsubstituted ¨(C-C)-.
[0083] "Ethynyl" refers to ¨(CC)-.
[0084] "Nitrogen-containing heterocyclyl" group means a 4- to 7- membered
non-
aromatic cyclic group containing at least one nitrogen atom, for example, but
without limitation,
morpholine, piperidine (e.g. 2-piperidinyl, 3-piperidinyl and 4-piperidinyl),
pyrrolidine (e.g. 2-
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pyrrolidinyl and 3-pyrrolidinyl), azetidine, pyrrolidone, imidazoline,
imidazolidinone, 2-
pyrazoline, pyrazolidine, piperazine, and N-alkyl piperazines such as N-methyl
piperazine.
Particular examples include azetidine, piperidone and piperazone.
[0085] "Thioketo" refers to the group =S.
[0086] Alkyl, alkenyl, alkynyl, carbocyclyl, heterocyclyl, aryl, and
heteroaryl groups, as
defined herein, are optionally substituted (e.g., "substituted" or
"unsubstituted" alkyl,
"substituted" or "unsubstituted" alkenyl, "substituted" or "unsubstituted"
alkynyl, "substituted"
or "unsubstituted" carbocyclyl, "substituted" or "unsubstituted" heterocyclyl,
"substituted" or
"unsubstituted" aryl or "substituted" or "unsubstituted" heteroaryl group). In
general, the term
"substituted", whether preceded by the term "optionally" or not, means that at
least one hydrogen
present on a group (e.g., a carbon or nitrogen atom) is replaced with a
permissible substituent,
e.g., a substituent which upon substitution results in a stable compound,
e.g., a compound which
does not spontaneously undergo transformation such as by rearrangement,
cyclization,
elimination, or other reaction. Unless otherwise indicated, a "substituted"
group has a substituent
at one or more substitutable positions of the group, and when more than one
position in any given
structure is substituted, the substituent is either the same or different at
each position. The term
"substituted" is contemplated to include substitution with all permissible
substituents of organic
compounds, any of the substituents described herein that results in the
formation of a stable
compound. The present invention contemplates any and all such combinations in
order to arrive
at a stable compound. For purposes of this invention, heteroatoms such as
nitrogen may have
hydrogen substituents and/or any suitable substituent as described herein
which satisfy the
valencies of the heteroatoms and results in the formation of a stable moiety.
[0087] Exemplary carbon atom substituents include, but are not limited to,
halogen, -CN,
-NO2, -N3, -S02H, -S03H, -OH, -OR', -0N(Rbb)2, -N(Rbb)2, -N(Rbb)3+X-, -N(OR)R,
_
SH, -SR', -SSR", -C(=0)R', -0O2H, -CHO, -c(OR)2, -CO2R', -0C(=0)R', -00O2R', -
C(=0)N(Rbb)2, -0C(=0)N(Rbb)2, -NRbbC(=0)R', -NRbbCO2R', -NRbbC(=0)N(Rbb)2, -
C(=NRbb)Raa, -C(=NRbb)OR', -0C(=NRbb)R', -0C(=NRbb)OR', -C(=NRbb)N(Rbb)2, -
OC(=NRbb)N(Rbb)2, -NRbbC(=NRbb)N(Rbb)2, -C(=0)NRbbSO2R', -NRbbSO2R', -
S02N(Rbb)2, -
SO2R', -S020Raa, -0S02R', -S(=0)R', -0S(=0)R', -Si(R')3, -0Si(Raa)3 -
C(=S)N(Rbb)2, -
C(=0)SR', -C(=S)SR', -SC(=S)SR', -SC(0)SR', -0C(0)SR', -SC(=0)OR', -
SC(=0)R', -P(=0)2R', -0P(=0)2R', -P(=0)(Raa)2, -0P(=0)(Raa)2, -0P(=0)(0Rec)2, -
P(=0)2N(Rbb)2, -0P(=0)2N(Rbb)2, -P(=0)(NRbb)2, -0P(=0)(NRbb)2, -
NRbbP(=0)(0Ree)2, -
NRbbP(=0)(NRbb)2, -P(R)2, -P(R)3, -OP(R)2, -OP(R)3, -B(R')2, -B(OR)2, -
BR'(ORee),
Ci_io alkyl, Ci_io perhaloalkyl, C2_10 alkenyl, C2-10 alkynyl, C3-10
carbocyclyl, 3-14 membered
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heterocyclyl, C6_14 aryl, and 5-14 membered heteroaryl, wherein each alkyl,
alkenyl, alkynyl,
carbocyclyl, heterocyclyl, aryl, and heteroaryl is independently substituted
with 0, 1, 2, 3, 4, or 5
Rdd groups;
or two geminal hydrogens on a carbon atom are replaced with the group =0, =S,
=NN(R)2,
NRbbc 0)Raa, NNRbbC( O)ORaa,= bNNR bs( 0)2-xa a, hh
NR-, or =NOR";
each instance of Raa is, independently, selected from Ci_io alkyl, Ci_io
perhaloalkyl, C2_10
alkenyl, C2_10 alkynyl, C3_10 carbocyclyl, 3-14 membered heterocyclyl, C6_14
aryl, and 5-14
membered heteroaryl, or two Raa groups are joined to form a 3-14 membered
heterocyclyl or 5-
14 membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl,
carbocyclyl, heterocyclyl,
aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 Rdd
groups;
each instance of Rbb is, independently, selected from hydrogen, -OH, -
N(Ree)2, -CN, -
C(=0)Raa, -C(=0)N(Ree)2, -CO2Raa, -SO2Raa, -C(=NR")0Raa, -C(=NR")N(Ree)2, -
SO2N(Ree)2,
-SO2Ree, -S020R", -SORaa, -C(=S)N(Ree)2, -C(=0)SR", -C(=S)SR", -P(=0)2Raa, -
P(=0)(Raa)2, -P(=0)2N(Ree)2, -P(=0)(NR")2, Ci_io alkyl, Ci_io perhaloalkyl,
C2_10 alkenyl, C2_10
alkynyl, C3_10 carbocyclyl, 3-14 membered heterocyclyl, C6_14 aryl, and 5-14
membered
heteroaryl, or two Rbb groups are joined to form a 3-14 membered heterocyclyl
or 5-14
membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl, carbocyclyl,
heterocyclyl, aryl,
and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 Rdd
groups;
each instance of Ree is, independently, selected from hydrogen, Ci_io alkyl,
Ci_io perhaloalkyl,
C2_10 alkenyl, C2_10 alkynyl, C3_10 carbocyclyl, 3-14 membered heterocyclyl,
C6_14 aryl, and 5-14
membered heteroaryl, or two Ree groups are joined to form a 3-14 membered
heterocyclyl or 5-
14 membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl,
carbocyclyl, heterocyclyl,
aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 Rdd
groups;
each instance of Rdd is, independently, selected from halogen, -CN, -NO2, -N3,
-S02H, -S03H,
-OH, -0Ree, -0N(Rff)2, -N(Rff)2, -N(Rff)3+X-, -N(ORee)Rff, -SH, -SRee, -SSRee,
-C(=0)Ree, -
CO2H, -CO2Ree, -0C(=0)Ree, -0CO2Ree, -C(=0)N(R1)2, -0C(=0)N(R1)2, -
NRffC(=0)Ree, -
NRffCO2Ree, -NRffC(=0)N(Rff)2, -C(=NRff)0Ree, -0C(=NRff)Ree, -0C(=NRff)0Ree, -
C(=NRff)N(Rff)2, -0C(=NRff)N(Rff)2, -NRffC(=NRff)N(Rff)2,-NRffS02Ree, -
SO2N(R)2, -
SO2Ree, -S020Ree, -0S02Ree, -S(=0)Ree, -Si(R)3, -0Si(Ree)3, -C(=S)N(Rff)2, -
C(=0)SRee, -
C(=S)SRee, -SC(=S)SRee, -P(=0)2Ree, -P(=0)(Ree)2, -0P(=0)(Ree)2, -
0P(=0)(0Ree)2, C1-6
alkyl, C1_6 perhaloalkyl, C2_6 alkenyl, C2_6 alkynyl, C3_10 carbocyclyl, 3-10
membered
heterocyclyl, C6_10 aryl, 5-10 membered heteroaryl, wherein each alkyl,
alkenyl, alkynyl,
carbocyclyl, heterocyclyl, aryl, and heteroaryl is independently substituted
with 0, 1, 2, 3, 4, or 5
Rgg groups, or two geminal Rdd substituents can be joined to form =0 or =S;
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each instance of Ree is, independently, selected from Ci_6 alkyl,
Ci_6perhaloalkyl, C2_6 alkenyl,
C2_6 alkynyl, C3_10 carbocyclyl, C6_10 aryl, 3-10 membered heterocyclyl, and 3-
10 membered
heteroaryl, wherein each alkyl, alkenyl, alkynyl, carbocyclyl, heterocyclyl,
aryl, and heteroaryl is
independently substituted with 0, 1, 2, 3, 4, or 5 Rgg groups;
each instance of e is, independently, selected from hydrogen, Ci_6 alkyl,
Ci_6perhaloalkyl, C2_6
alkenyl, C2_6 alkynyl, C3_10 carbocyclyl, 3-10 membered heterocyclyl, C6_1()
aryl and 5-10
membered heteroaryl, or two e groups are joined to form a 3-14 membered
heterocyclyl or 5-
14 membered heteroaryl ring, wherein each alkyl, alkenyl, alkynyl,
carbocyclyl, heterocyclyl,
aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4, or 5 Rgg
groups; and
each instance of Rgg is, independently, halogen, -CN, -NO2, -N3, -S02H, -S03H,
-OH, -0C1-6
alkyl, -0N(C1_6 alky1)2, -N(C1_6 alky1)2, -N(C1_6 alky1)3+X , -NH(C1_6
alky1)2+X , -NH2(C1_6
alkyl) +X-, -NH3+X-, -N(OCi_6 alkyl)(Ci_6 alkyl), -N(OH)(C1_6 alkyl), -NH(OH),
-SH, -SC1-6
alkyl, -SS(C1_6 alkyl), -C(=0)(C1_6 alkyl), -CO2H, -0O2(C1_6 alkyl), -
0C(=0)(C1_6 alkyl), -
00O2(C1_6 alkyl), -C(=0)NH2, -C(=0)N(C1_6 alky1)2, -0C(=0)NH(C1_6 alkyl), -
NHC(=0)( C1-
6 alkyl), -N(C1_6 alkyl)C(=0)( C1_6 alkyl), -NHCO2(C1_6 alkyl), -NHC(=0)N(C1_6
alky1)2, -
NHC(=0)NH(C1_6 alkyl), -NHC(=0)NH2, -C(=NH)0(C1_6 alkyl),-0C(=NH)(C1_6 alkyl),
-
OC(=NH)0C1_6 alkyl, -C(=NH)N(C1_6 alky1)2, -C(=NH)NH(C1_6 alkyl), -C(=NH)NH2, -
0C(=NH)N(C1_6 alky1)2, -0C(NH)NH(C1_6 alkyl), -0C(NH)NH2, -NHC(NH)N(C1_6
alky1)2, -
NHC(=NH)NH2, -NHS02(C1_6 alkyl), -SO2N(C1_6 alky1)2, -SO2NH(C1_6 alkyl), -
SO2NH2,-
S02C1_6 alkyl, -S020C1_6 alkyl, -0S02C1_6 alkyl, -SOC1_6 alkyl, -Si(Ci_6
alky1)3, -0Si(C1-6
alky1)3 -C(=S)N(Ci_6 alky1)2, C(=S)NH(C1_6 alkyl), C(=S)NH2, -C(=0)S(C1_6
alkyl), -
C(=S)SC1_6 alkyl, -SC(=S)SC1_6 alkyl, -P(=0)2(C1_6 alkyl), -P(=0)(C1_6
alky1)2, -0P(=0)(C1-6
alky1)2, -0P(=0)(0C1_6 alky1)2, C1_6 alkyl, Ci_6 perhaloalkyl, C2_6 alkenyl,
C2_6 alkynyl, C3-10
carbocyclyl, C6_10 aryl, 3-10 membered heterocyclyl, 5-10 membered heteroaryl;
or two geminal
Rgg substituents can be joined to form =0 or =S; wherein X- is a counterion.
[0088] A "counterion" or "anionic counterion" is a negatively charged group
associated
with a cationic quaternary amino group in order to maintain electronic
neutrality. Exemplary
counterions include halide ions (e.g., F, Cl-, Br-, F), NO3-, C104-, OW, H2PO4-
, HSO4-,
sulfonate ions (e.g., methansulfonate, trifluoromethanesulfonate, p-
toluenesulfonate,
benzenesulfonate, 10-camphor sulfonate, naphthalene-2-sulfonate, naphthalene-l-
sulfonic
acid-5-sulfonate, ethan-l-sulfonic acid-2-sulfonate, and the like), and
carboxylate ions (e.g.,
acetate, ethanoate, propanoate, benzoate, glycerate, lactate, tartrate,
glycolate, and the like).
[0089] Nitrogen atoms can be substituted or unsubstituted as valency
permits, and include
primary, secondary, tertiary, and quarternary nitrogen atoms. Exemplary
nitrogen atom
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substitutents include, but are not limited to, hydrogen, -OH, -OR', -N(R")2, -
CN, -C(=0)Raa,
-C(=0)N(Rec)2, -CO2Raa, -SO2Raa, -C(=NRbb)Raa, -C(=NRce)0Raa, -C(=NInN(Rec)2, -
SO2N(R")2, -SO2R", -S020R", -SORaa, -C(=S)N(R")2, -C(=0)SR", -C(=S)SR", -
P(=0)2Raa,
-P(=0)(Raa)2, -P(=0)2N(Rec)2, -P(=0)(NRec)2, Ci_io alkyl, Ci_io perhaloalkyl,
C2-10 alkenyl, C2-
alkynyl, C3_10 carbocyclyl, 3-14 membered heterocyclyl, C6_14 aryl, and 5-14
membered
heteroaryl, or two Ree groups attached to a nitrogen atom are joined to form a
3-14 membered
heterocyclyl or 5-14 membered heteroaryl ring, wherein each alkyl, alkenyl,
alkynyl,
carbocyclyl, heterocyclyl, aryl, and heteroaryl is independently substituted
with 0, 1, 2, 3, 4, or 5
Rdd groups, and wherein Raa, Rbb, Ree and Rdd are as defined above.
[0090] In certain embodiments, the substituent present on a nitrogen atom
is a nitrogen
protecting group (also referred to as an amino protecting group). Nitrogen
protecting groups
include, but are not limited to, -OH, -OR', -N(Rec)2, -C(=0)Raa, -C(=0)N(R")2,
-CO2Raa, -
SO2Raa, -C(=NRce)Raa, -C(=NRce)0Raa, -C(=NRce)N(Rec)2, -SO2N(Rec)2, -SO2Rec, -
S020Rec, -
SORaa, -C(=S)N(Rec)2, -C(=0)SR", -C(=S)SR", Ci_io alkyl (e.g., aralkyl,
heteroaralkyl), C2-10
alkenyl, C2_10 alkynyl, C3_10 carbocyclyl, 3-14 membered heterocyclyl, C6_14
aryl, and 5-14
membered heteroaryl groups, wherein each alkyl, alkenyl, alkynyl, carbocyclyl,
heterocyclyl,
aralkyl, aryl, and heteroaryl is independently substituted with 0, 1, 2, 3, 4,
or 5 Rdd groups, and
wherein Raa, Rbb, Re' and Rdd are as defined herein. Nitrogen protecting
groups are well known in
the art and include those described in detail in Protecting Groups in Organic
Synthesis, T. W.
Greene and P. G. M. Wuts, 3rd edition, John Wiley & Sons, 1999, incorporated
herein by
reference.
[0091] For example, nitrogen protecting groups such as amide groups (e.g., -
C(=0)Raa)
include, but are not limited to, formamide, acetamide, chloroacetamide,
trichloroacetamide,
trifluoroacetamide, phenylacetamide, 3-phenylpropanamide, picolinamide, 3-
pyridylcarboxamide, N-benzoylphenylalanyl derivative, benzamide, p-
phenylbenzamide, o-
nitophenylacetamide, o-nitrophenoxyacetamide, acetoacetamide, (N'-
dithiobenzyloxyacylamino)acetamide, 3-(p-hydroxyphenyl)propanamide, 3-(o-
nitrophenyl)propanamide, 2-methyl-2-(o-nitrophenoxy)propanamide, 2-methy1-2-(o-
phenylazophenoxy)propanamide, 4-chlorobutanamide, 3-methyl-3-nitrobutanamide,
o-
nitrocinnamide, N-acetylmethionine derivative, o-nitrobenzamide and o-
(benzoyloxymethyl)benzamide.
[0092] Nitrogen protecting groups such as carbamate groups (e.g., -
C(=0)0Raa) include,
but are not limited to, methyl carbamate, ethyl carbamante, 9-fluorenylmethyl
carbamate
(Fmoc), 9-(2-sulfo)fluorenylmethyl carbamate, 9-(2,7-dibromo)fluoroenylmethyl
carbamate,
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2,7¨di¨t¨butyl[9410,10¨dioxo-10,10,10,10¨tetrahydrothioxanthylAmethyl
carbamate (DBD¨
Tmoc), 4¨methoxyphenacyl carbamate (Phenoc), 2,2,2¨trichloroethyl carbamate
(Troc), 2¨
trimethylsilylethyl carbamate (Teoc), 2¨phenylethyl carbamate (hZ),
141¨adamanty1)-1¨
methylethyl carbamate (Adpoc), 1,1¨dimethy1-2¨haloethyl carbamate,
1,1¨dimethy1-2,2¨
dibromoethyl carbamate (DB¨t¨BOC), 1,1¨dimethy1-2,2,2¨trichloroethyl carbamate
(TCBOC),
1¨methyl-144¨biphenylyl)ethyl carbamate (Bpoc), 143,5¨di¨t¨butylpheny1)-
1¨methylethyl
carbamate (t¨Bumeoc), 2¨(2'¨ and 4'¨pyridyl)ethyl carbamate (Pyoc), 2¨(N,N¨
dicyclohexylcarboxamido)ethyl carbamate, t¨butyl carbamate (BOC), 1¨adamantyl
carbamate
(Adoc), vinyl carbamate (Voc), ally' carbamate (Alloc), 1¨isopropylally1
carbamate (Ipaoc),
cinnamyl carbamate (Coc), 4¨nitrocinnamyl carbamate (Noc), 8¨quinoly1
carbamate, N¨
hydroxypiperidinyl carbamate, alkyldithio carbamate, benzyl carbamate (Cbz),
p¨methoxybenzyl
carbamate (Moz), p¨nitobenzyl carbamate, p¨bromobenzyl carbamate,
p¨chlorobenzyl
carbamate, 2,4¨dichlorobenzyl carbamate, 4¨methylsulfinylbenzyl carbamate
(Msz), 9¨
anthrylmethyl carbamate, diphenylmethyl carbamate, 2¨methylthioethyl
carbamate, 2¨
methylsulfonylethyl carbamate, 2¨(p¨toluenesulfonyl)ethyl carbamate,
[241,3¨dithianylAmethyl
carbamate (Dmoc), 4¨methylthiophenyl carbamate (Mtpc), 2,4¨dimethylthiophenyl
carbamate
(Bmpc), 2¨phosphonioethyl carbamate (Peoc), 2¨triphenylphosphonioisopropyl
carbamate
(Ppoc), 1,1¨dimethy1-2¨cyanoethyl carbamate, m¨chloro¨p¨acyloxybenzyl
carbamate, p¨
(dihydroxyboryl)benzyl carbamate, 5¨benzisoxazolylmethyl carbamate,
24trifluoromethyl)-6¨
chromonylmethyl carbamate (Tcroc), m¨nitrophenyl carbamate,
3,5¨dimethoxybenzyl
carbamate, o¨nitrobenzyl carbamate, 3,4¨dimethoxy-6¨nitrobenzyl carbamate,
phenyl(o¨
nitrophenyl)methyl carbamate, t¨amyl carbamate, S¨benzyl thiocarbamate,
p¨cyanobenzyl
carbamate, cyclobutyl carbamate, cyclohexyl carbamate, cyclopentyl carbamate,
cyclopropylmethyl carbamate, p¨decyloxybenzyl carbamate,
2,2¨dimethoxyacylvinyl carbamate,
o¨(N,N¨dimethylcarboxamido)benzyl carbamate, 1,1¨dimethy1-3¨(N,N¨
dimethylcarboxamido)propyl carbamate, 1,1¨dimethylpropynyl carbamate,
di(2¨pyridyl)methyl
carbamate, 2¨furanylmethyl carbamate, 2¨iodoethyl carbamate, isoborynl
carbamate, isobutyl
carbamate, isonicotinyl carbamate, p¨(p '¨methoxyphenylazo)benzyl carbamate,
1¨
methylcyclobutyl carbamate, 1¨methylcyclohexyl carbamate,
1¨methyl¨l¨cyclopropylmethyl
carbamate, 1¨methyl-143,5¨dimethoxyphenyl)ethyl carbamate, 1¨methy1-14p¨
phenylazophenyl)ethyl carbamate, 1¨methyl¨l¨phenylethyl carbamate, 1¨methy1-
144¨
pyridyl)ethyl carbamate, phenyl carbamate,p¨(phenylazo)benzyl carbamate,
2,4,6¨tri¨t¨
butylphenyl carbamate, 4¨(trimethylammonium)benzyl carbamate, and
2,4,6¨trimethylbenzyl
carbamate.
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[0093] Nitrogen protecting groups such as sulfonamide groups (e.g.,
¨S(=0)2Raa) include,
but are not limited to, p¨toluenesulfonamide (Ts), benzenesulfonamide,
2,3,6,¨trimethy1-4¨
methoxybenzenesulfonamide (Mtr), 2,4,6¨trimethoxybenzenesulfonamide (Mtb),
2,6¨dimethy1-
4¨methoxybenzenesulfonamide (Pme), 2,3,5,6¨tetramethy1-
4¨methoxybenzenesulfonamide
(Mte), 4¨methoxybenzenesulfonamide (Mbs), 2,4,6¨trimethylbenzenesulfonamide
(Mts), 2,6¨
dimethoxy-4¨methylbenzenesulfonamide (iMds), 2,2,5,7,8¨pentamethylchroman-6¨
sulfonamide (Pmc), methanesulfonamide (Ms), P¨trimethylsilylethanesulfonamide
(SES), 9¨
anthracenesulfonamide, 4¨(4',8'¨dimethoxynaphthylmethyl)benzenesulfonamide
(DNMBS),
benzylsulfonamide, trifluoromethylsulfonamide, and phenacylsulfonamide.
[0094] Other nitrogen protecting groups include, but are not limited to,
phenothiazinyl¨
(10)¨acyl derivative, N'¨p¨toluenesulfonylaminoacyl derivative,
N'¨phenylaminothioacyl
derivative, N¨benzoylphenylalanyl derivative, N¨acetylmethionine derivative,
4,5¨dipheny1-3¨
oxazolin-2¨one, N¨phthalimide, N¨dithiasuccinimide (Dts), N-
2,3¨diphenylmaleimide, N-2,5¨
dimethylpyrrole, N-1,1,4,4¨tetramethyldisilylazacyclopentane adduct (STABASE),
5¨
substituted 1,3¨dimethy1-1,3,5¨triazacyclohexan-2¨one, 5¨substituted
1,3¨dibenzy1-1,3,5¨
triazacyclohexan-2¨one, 1¨substituted 3,5¨dinitro-4¨pyridone, N¨methylamine,
N¨allylamine,
N[2¨(trimethylsilyl)ethoxy]methylamine (SEM), N-3¨acetoxypropylamine,
N¨(1¨isopropy1-4¨
nitro-2¨oxo-3¨pyroolin-3¨yl)amine, quaternary ammonium salts, N¨benzylamine,
N¨di(4¨
methoxyphenyl)methylamine, N-5¨dibenzosuberylamine, N¨triphenylmethylamine
(Tr), N¨[(4¨
methoxyphenyl)diphenylmethyl]amine (MMTr), N-9¨phenylfluorenylamine (PhF), N-
2,7¨
dichloro-9¨fluorenylmethyleneamine, N¨ferrocenylmethylamino (Fcm), N-
2¨picolylamino N'¨
oxide, N-1,1¨dimethylthiomethyleneamine, N¨benzylideneamine, N¨p¨
methoxybenzylideneamine, N¨diphenylmethyleneamine, N¨[(2¨
pyridyl)mesityl]methyleneamine, N¨(N' ,N'¨dimethylaminomethylene)amine, N,N'¨
isopropylidenediamine, N¨p¨nitrobenzylideneamine, N¨salicylideneamine, N-5¨
chlorosalicylideneamine, N¨(5¨chloro-2¨hydroxyphenyl)phenylmethyleneamine, N¨
cyclohexylideneamine, N¨(5 ,5¨dimethy1-3¨oxo¨l¨cyclohexenyl)amine, N¨borane
derivative,
N¨diphenylborinic acid derivative, N¨[phenyl(pentaacylchromium¨ or
tungsten)acyl]amine, N¨
copper chelate, N¨zinc chelate, N¨nitroamine, N¨nitrosoamine, amine N¨oxide,
diphenylphosphinamide (Dpp), dimethylthiophosphinamide (Mpt),
diphenylthiophosphinamide
(Ppt), dialkyl phosphoramidates, dibenzyl phosphoramidate, diphenyl
phosphoramidate,
benzenesulfenamide, o¨nitrobenzenesulfenamide (Nps),
2,4¨dinitrobenzenesulfenamide,
pentachlorobenzenesulfenamide, 2¨nitro-4¨methoxybenzenesulfenamide,
triphenylmethylsulfenamide, and 3¨nitropyridinesulfenamide (Npys).
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100951 In certain embodiments, the substituent present on an oxygen atom is
an oxygen
protecting group (also referred to as a hydroxyl protecting group). Oxygen
protecting groups
include, but are not limited to, ¨R', ¨N(Rbb)2, _C(0)SR', ¨C(=0)R', ¨CO2R', ¨
C(=0)N(Rbb)2, ¨C(=NRbb)R', ¨C(=NRbb)OR', ¨C(=NRbb)N(Rbb)2, ¨S(=0)R', ¨SO2R', ¨
Si(R')3,¨P(Ree)2, ¨P(R)3, ¨P(=0)2R', ¨P(=0)(R')2, ¨P(=0)(0Ree)2,
¨P(=0)2N(Rbb)2, and ¨
p(=0"Rbb)2, wherein R', Rbb, and Ree are as defined herein. Oxygen protecting
groups are
well known in the art and include those described in detail in Protecting
Groups in Organic
Synthesis, T. W. Greene and P. G. M. Wuts, 3rd edition, John Wiley & Sons,
1999, incorporated
herein by reference.
[0096] Exemplary oxygen protecting groups include, but are not limited to,
methyl,
methoxylmethyl (MOM), methylthiomethyl (MTM), t¨butylthiomethyl,
(phenyldimethylsilyl)methoxymethyl (SMOM), benzyloxymethyl (BOM), p¨
methoxybenzyloxymethyl (PMBM), (4¨methoxyphenoxy)methyl (p¨AOM),
guaiacolmethyl
(GUM), t¨butoxymethyl, 4¨pentenyloxymethyl (POM), siloxymethyl,
2¨methoxyethoxymethyl
(MEM), 2,2,2¨trichloroethoxymethyl, bis(2¨chloroethoxy)methyl, 2¨
(trimethylsilyl)ethoxymethyl (SEMOR), tetrahydropyranyl (THP),
3¨bromotetrahydropyranyl,
tetrahydrothiopyranyl, 1¨methoxycyclohexyl, 4¨methoxytetrahydropyranyl (MTHP),
4¨
methoxytetrahydrothiopyranyl, 4¨methoxytetrahydrothiopyranyl S,S¨dioxide,
1¨[(2¨chloro-4¨
methyl)pheny1]-4¨methoxypiperidin-4¨y1 (CTMP), 1,4¨dioxan-2¨yl,
tetrahydrofuranyl,
tetrahydrothiofuranyl, 2,3,3 a,4,5,6,7,7a¨octahydro-7,8,8¨trimethy1-
4,7¨methanobenzofuran-2¨
yl, 1¨ethoxyethyl, 1¨(2¨chloroethoxy)ethyl, 1¨methyl¨l¨methoxyethyl, 1¨methy1-
1¨
benzyloxyethyl, 1¨methyl-1¨benzyloxy-2¨fluoroethyl, 2,2,2¨trichloroethyl, 2¨
trimethylsilylethyl, 2¨(phenylselenyl)ethyl, t¨butyl, allyl, p¨chlorophenyl,
p¨methoxyphenyl,
2,4¨dinitrophenyl, benzyl (Bn), p¨methoxybenzyl, 3,4¨dimethoxybenzyl,
o¨nitrobenzyl, p¨
nitrobenzyl, p¨halobenzyl, 2,6¨dichlorobenzyl, p¨cyanobenzyl, p¨phenylbenzyl,
2¨picolyl, 4¨
picolyl, 3¨methyl-2¨picoly1N¨oxido, diphenylmethyl, p,p '¨dinitrobenzhydryl,
5¨
dibenzosuberyl, triphenylmethyl, a¨naphthyldiphenylmethyl,
p¨methoxyphenyldiphenylmethyl,
di(p¨methoxyphenyl)phenylmethyl, tri(p¨methoxyphenyl)methyl, 4¨(4'¨
bromophenacyloxyphenyl)diphenylmethyl,
4,4',4"¨tris(4,5¨dichlorophthalimidophenyl)methyl,
4,4',4"¨tris(levulinoyloxyphenyl)methyl, 4,4',4"¨tris(benzoyloxyphenyl)methyl,
3¨(imidazol-1¨
yl)bis(4',4"¨dimethoxyphenyl)methyl, 1,1¨bis(4¨methoxypheny1)-
1'¨pyrenylmethyl, 9¨anthryl,
9¨(9¨phenyl)xanthenyl, 9(9¨pheny1-10¨oxo)anthryl, 1,3¨benzodisulfuran-2¨yl,
benzisothiazolyl 5,5¨dioxido, trimethylsilyl (TMS), triethylsilyl (TES),
triisopropylsilyl (TIPS),
dimethylisopropylsilyl (IPDMS), diethylisopropylsilyl (DEIPS),
dimethylthexylsilyl, t-
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butyldimethylsilyl (TBDMS), t¨butyldiphenylsilyl (TBDPS), tribenzylsilyl,
tri¨p¨xylylsilyl,
triphenylsilyl, diphenylmethylsilyl (DPMS), t¨butylmethoxyphenylsilyl (TBMPS),
formate,
benzoylformate, acetate, chloroacetate, dichloroacetate, trichloroacetate,
trifluoroacetate,
methoxyacetate, triphenylmethoxyacetate, phenoxyacetate,
p¨chlorophenoxyacetate, 3¨
phenylpropionate, 4¨oxopentanoate (levulinate), 4,4¨(ethylenedithio)pentanoate
(leyulinoyldithioacetal), pivaloate, adamantoate, crotonate,
4¨methoxycrotonate, benzoate, p¨
phenylbenzoate, 2,4,6¨trimethylbenzoate (mesitoate), alkyl methyl carbonate,
9¨fluorenylmethyl
carbonate (Fmoc), alkyl ethyl carbonate, alkyl 2,2,2¨trichloroethyl carbonate
(Troc), 2¨
(trimethylsilyl)ethyl carbonate (TMSEC), 2¨(phenylsulfonyl) ethyl carbonate
(Psec), 2¨
(triphenylphosphonio) ethyl carbonate (Peoc), alkyl isobutyl carbonate, alkyl
vinyl carbonate
alkyl ally' carbonate, alkyl p¨nitrophenyl carbonate, alkyl benzyl carbonate,
alkyl p¨
methoxybenzyl carbonate, alkyl 3,4¨dimethoxybenzyl carbonate, alkyl
o¨nitrobenzyl carbonate,
alkyl p¨nitrobenzyl carbonate, alkyl S¨benzyl thiocarbonate, 4¨ethoxy-
1¨napththyl carbonate,
methyl dithiocarbonate, 2¨iodobenzoate, 4¨azidobutyrate, 4¨nitro-
4¨methylpentanoate, o¨
(dibromomethyl)benzoate, 2¨formylbenzenesulfonate, 2¨(methylthiomethoxy)ethyl,
4¨
(methylthiomethoxy)butyrate, 2¨(methylthiomethoxymethyl)benzoate, 2,6¨dichloro-
4¨
methylphenoxyacetate, 2,6¨dichloro-4¨(1,1,3,3¨tetramethylbutyl)phenoxyacetate,
2,4¨bis(1,1¨
dimethylpropyl)phenoxyacetate, chlorodiphenylacetate, isobutyrate,
monosuccinoate, (E)-2¨
methy1-2¨butenoate, o¨(methoxyacyl)benzoate, a¨naphthoate, nitrate, alkyl
N,N,N',N'¨
tetramethylphosphorodiamidate, alkyl N¨phenylcarbamate, borate,
dimethylphosphinothioyl,
alkyl 2,4¨dinitrophenylsulfenate, sulfate, methanesulfonate (mesylate),
benzylsulfonate, and
tosylate (Ts).
[0097] In certain embodiments, the substituent present on an sulfur atom is
an sulfur
protecting group (also referred to as a thiol protecting group). Sulfur
protecting groups include,
but are not limited to, ¨Raa, ¨N(Rbb)2, ¨C(=0)SRaa, ¨C(=0)Raa, ¨CO2Raa,
¨C(=0)N(Rbb)2, ¨
C(=NRbb)Raa, ¨C(=NRbb)0Ra1, ¨C(=NRbb)N(Rbb)2, ¨S(=0)Raa, ¨SO2Raa,
¨Si(Raa)3,¨P(Rec)2, ¨
P(R)3, ¨P(=0)2Raa, ¨P(=0)(Raa)2, ¨P(=0)(0Ree)2, ¨P(=0)2N(Rbb)2, and
¨P(=0)(NRbb)2, wherein
Raa, Rbb, and Re' are as defined herein. Sulfur protecting groups are well
known in the art and
include those described in detail in Protecting Groups in Organic Synthesis,
T. W. Greene and P.
G. M. Wuts, 3rd edition, John Wiley & Sons, 1999, incorporated herein by
reference.
[0098] "Compounds of the present invention", and equivalent expressions,
are meant to
embrace the compounds as hereinbefore described, in particular compounds
according to any of
the Formula herein recited and/or described, which expression includes the
prodrugs, the
pharmaceutically acceptable salts, and the solvates, e.g., hydrates, where the
context so permits.
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Similarly, reference to intermediates, whether or not they themselves are
claimed, is meant to
embrace their salts, and solvates, where the context so permits.
[0099] These and other exemplary substituents are described in more detail
in the
Detailed Description, Examples, and claims. The invention is not intended to
be limited in any
manner by the above exemplary listing of substituents.
Other definitions
[00100] "Pharmaceutically acceptable" means approved or approvable by a
regulatory
agency of the Federal or a state government or the corresponding agency in
countries other than
the United States, or that is listed in the U.S. Pharmacopoeia or other
generally recognized
pharmacopoeia for use in animals, and more particularly, in humans.
[00101] "Pharmaceutically acceptable salt" refers to a salt of a compound
of the invention
that is pharmaceutically acceptable and that possesses the desired
pharmacological activity of the
parent compound. In particular, such salts are non-toxic may be inorganic or
organic acid
addition salts and base addition salts. Specifically, such salts include: (1)
acid addition salts,
formed with inorganic acids such as hydrochloric acid, hydrobromic acid,
sulfuric acid, nitric
acid, phosphoric acid, and the like; or formed with organic acids such as
acetic acid, propionic
acid, hexanoic acid, cyclopentanepropionic acid, glycolic acid, pyruvic acid,
lactic acid, malonic
acid, succinic acid, malic acid, maleic acid, fumaric acid, tartaric acid,
citric acid, benzoic acid,
3-(4-hydroxybenzoyl) benzoic acid, cinnamic acid, mandelic acid,
methanesulfonic acid,
ethanesulfonic acid, 1,2-ethane-disulfonic acid, 2-hydroxyethanesulfonic acid,
benzenesulfonic
acid, 4-chlorobenzenesulfonic acid, 2-naphthalenesulfonic acid, 4-
toluenesulfonic acid,
camphorsulfonic acid, 4-methylbicyclo[2.2.2]-oct-2-ene-1-carboxylic acid,
glucoheptonic acid,
3-phenylpropionic acid, trimethylacetic acid, tertiary butylacetic acid,
lauryl sulfuric acid,
gluconic acid, glutamic acid, hydroxynaphthoic acid, salicylic acid, stearic
acid, muconic acid,
and the like; or (2) salts formed when an acidic proton present in the parent
compound either is
replaced by a metal ion, e.g., an alkali metal ion, an alkaline earth ion, or
an aluminum ion; or
coordinates with an organic base such as ethanolamine, diethanolamine,
triethanolamine, N-
methylglucamine and the like. Salts further include, by way of example only,
sodium,
potassium, calcium, magnesium, ammonium, tetraalkylammonium, and the like; and
when the
compound contains a basic functionality, salts of non toxic organic or
inorganic acids, such as
hydrochloride, hydrobromide, tartrate, mesylate, acetate, maleate, oxalate and
the like. The term
"pharmaceutically acceptable cation" refers to an acceptable cationic counter-
ion of an acidic
functional group. Such cations are exemplified by sodium, potassium, calcium,
magnesium,
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ammonium, tetraalkylammonium cations, and the like (see, e.g., Berge, et al.,
J. Pharm. Sci.
66(1): 1-79 (Jan.'77).
[00102]
"Pharmaceutically acceptable vehicle" refers to a diluent, adjuvant, excipient
or
carrier with which a compound of the invention is administered.
[00103]
"Pharmaceutically acceptable metabolically cleavable group" refers to a group
which is cleaved in vivo to yield the parent molecule of the structural
Formula indicated herein.
Examples of metabolically cleavable groups include -COR, -COOR,-CONRR and
¨CH2OR
radicals, where R is selected independently at each occurrence from alkyl,
trialkylsilyl,
carbocyclic aryl or carbocyclic aryl substituted with one or more of alkyl,
halogen, hydroxy or
alkoxy. Specific examples of representative metabolically cleavable groups
include acetyl,
methoxycarbonyl, benzoyl, methoxymethyl and trimethylsilyl groups.
[00104]
"Prodrugs" refers to compounds, including derivatives of the compounds of the
invention,which have cleavable groups and become by solvolysis or under
physiological
conditions the compounds of the invention that are pharmaceutically active in
vivo. Such
examples include, but are not limited to, choline ester derivatives and the
like, N-
alkylmorpholine esters and the like. Other derivatives of the compounds of
this invention have
activity in both their acid and acid derivative forms, but in the acid
sensitive form often offers
advantages of solubility, tissue compatibility, or delayed release in the
mammalian organism
(see, Bundgard, H., Design of Prodrugs, pp. 7-9, 21-24, Elsevier, Amsterdam
1985). Prodrugs
include acid derivatives well know to practitioners of the art, such as, for
example, esters
prepared by reaction of the parent acid with a suitable alcohol, or amides
prepared by reaction of
the parent acid compound with a substituted or unsubstituted amine, or acid
anhydrides, or mixed
anhydrides. Simple aliphatic or aromatic esters, amides and anhydrides derived
from acidic
groups pendant on the compounds of this invention are particular prodrugs. In
some cases it is
desirable to prepare double ester type prodrugs such as (acyloxy)alkyl esters
or
((alkoxycarbonyl)oxy)alkylesters. Particularly the Ci to C8 alkyl, C2-C8
alkenyl, C2-C8 alkynyl,
aryl, C2-C12 substituted aryl, and C2-C12 arylalkyl esters of the compounds of
the invention.
[00105]
"Solvate" refers to forms of the compound that are associated with a solvent
or
water (also referred to as "hydrate"), usually by a solvolysis reaction. This
physical association
includes hydrogen bonding. Conventional solvents include water, ethanol,
acetic acid and the
like. The compounds of the invention may be prepared e.g. in crystalline form
and may be
solvated or hydrated. Suitable solvates include pharmaceutically acceptable
solvates, such as
hydrates, and further include both stoichiometric solvates and non-
stoichiometric solvates. In
certain instances the solvate will be capable of isolation, for example when
one or more solvent
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molecules are incorporated in the crystal lattice of the crystalline solid.
"Solvate" encompasses
both solution-phase and isolable solvates. Representative solvates include
hydrates, ethanolates
and methanolates.
[00106] A "subject" to which administration is contemplated includes, but
is not limited
to, humans (i.e., a male or female of any age group, e.g., a pediatric subject
(e.g, infant, child,
adolescent) or adult subject (e.g., young adult, middle¨aged adult or senior
adult)) and/or a non-
human animal, e.g., a mammal such as primates (e.g., cynomolgus monkeys,
rhesus monkeys),
cattle, pigs, horses, sheep, goats, rodents, cats, and/or dogs. In certain
embodiments, the subject
is a human. In certain embodiments, the subject is a non-human animal. The
terms "human",
"patient" and "subject" are used interchangeably herein.
[00107] "Therapeutically effective amount" means the amount of a compound
that, when
administered to a subject for treating a disease, is sufficient to effect such
treatment for the
disease. The "therapeutically effective amount" can vary depending on the
compound, the
disease and its severity, and the age, weight, etc., of the subject to be
treated.
[00108] "Preventing" or "prevention" refers to a reduction in risk of
acquiring or
developing a disease or disorder (i.e., causing at least one of the clinical
symptoms of the disease
not to develop in a subject not yet exposed to a disease-causing agent, or
predisposed to the
disease in advance of disease onset.
[00109] The term "prophylaxis" is related to "prevention", and refers to a
measure or
procedure the purpose of which is to prevent, rather than to treat or cure a
disease. Non-limiting
examples of prophylactic measures may include the administration of vaccines;
the
administration of low molecular weight heparin to hospital patients at risk
for thrombosis due, for
example, to immobilization; and the administration of an anti-malarial agent
such as chloroquine,
in advance of a visit to a geographical region where malaria is endemic or the
risk of contracting
malaria is high.
[00110] "Treating" or "treatment" of any disease or disorder refers, in
certain
embodiments, to ameliorating the disease or disorder (i.e., arresting the
disease or reducing the
manifestation, extent or severity of at least one of the clinical symptoms
thereof). In another
embodiment "treating" or "treatment" refers to ameliorating at least one
physical parameter,
which may not be discernible by the subject. In yet another embodiment,
"treating" or
"treatment" refers to modulating the disease or disorder, either physically,
(e.g., stabilization of a
discernible symptom), physiologically, (e.g., stabilization of a physical
parameter), or both. In a
further embodiment, "treating" or "treatment" relates to slowing the
progression of the disease.
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[00111] As used herein, the term "isotopic variant" refers to a compound
that contains
unnatural proportions of isotopes at one or more of the atoms that constitute
such compound.
For example, an "isotopic variant" of a compound can contain one or more non-
radioactive
isotopes, such as for example, deuterium (2H or D), carbon-13 (13C), nitrogen-
15 (15N), or the
like. It will be understood that, in a compound where such isotopic
substitution is made, the
following atoms, where present, may vary, so that for example, any hydrogen
may be 2H/D, any
carbon may be 13C, or any nitrogen may be 15N, and that the presence and
placement of such
atoms may be determined within the skill of the art. Likewise, the invention
may include the
preparation of isotopic variants with radioisotopes, in the instance for
example, where the
resulting compounds may be used for drug and/or substrate tissue distribution
studies. The
radioactive isotopes tritium, i.e., 3H, and carbon-14, i.e., 14C, are
particularly useful for this
purpose in view of their ease of incorporation and ready means of detection.
Further, compounds
may be prepared that are substituted with positron emitting isotopes, such as
11C, 18F, 150 and
13N, and would be useful in Positron Emission Topography (PET) studies for
examining substrate
receptor occupancy. All isotopic variants of the compounds provided herein,
radioactive or not,
are intended to be encompassed within the scope of the invention.
[00112] It is also to be understood that compounds that have the same
molecular formula
but differ in the nature or sequence of bonding of their atoms or the
arrangement of their atoms in
space are termed "isomers". Isomers that differ in the arrangement of their
atoms in space are
termed "stereoisomers".
[00113] Stereoisomers that are not mirror images of one another are termed
"diastereomers" and those that are non-superimposable mirror images of each
other are termed
"enantiomers". When a compound has an asymmetric center, for example, when it
is bonded to
four different groups, a pair of enantiomers is possible. An enantiomer can be
characterized by
the absolute configuration of its asymmetric center and is described by the R-
and S-sequencing
rules of Cahn and Prelog, or by the manner in which the molecule rotates the
plane of polarized
light and designated as dextrorotatory or leyorotatory (i.e., as (+) or (-)-
isomers respectively). A
chiral compound can exist as either individual enantiomer or as a mixture
thereof A mixture
containing equal proportions of the enantiomers is called a "racemic mixture".
[00114] "Tautomers" refer to compounds that are interchangeable forms of a
particular
compound structure, and that vary in the displacement of hydrogen atoms and
electrons. Thus,
two structures may be in equilibrium through the movement of 7I electrons and
an atom (usually
H). For example, enols and ketones are tautomers because they are rapidly
interconyerted by
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treatment with either acid or base. Another example of tautomerism is the aci-
and nitro- forms
of phenylnitromethane, which are likewise formed by treatment with acid or
base.
Tautomeric forms may be relevant to the attainment of the optimal chemical
reactivity and
biological activity of a compound of interest.
[00115] As used herein a pure enantiomeric compound is substantially free
from other
enantiomers or stereoisomers of the compound (i.e., in enantiomeric excess).
In other words, an
"S" form of the compound is substantially free from the "R" form of the
compound and is, thus,
in enantiomeric excess of the "R" form. The term "enantiomerically pure" or
"pure enantiomer"
denotes that the compound comprises more than 75% by weight, more than 80% by
weight, more
than 85% by weight, more than 90% by weight, more than 91% by weight, more
than 92% by
weight, more than 93% by weight, more than 94% by weight, more than 95% by
weight, more
than 96% by weight, more than 97% by weight, more than 98% by weight, more
than 98.5% by
weight, more than 99% by weight, more than 99.2% by weight, more than 99.5% by
weight,
more than 99.6% by weight, more than 99.7% by weight, more than 99.8% by
weight or more
than 99.9% by weight, of the enantiomer. In certain embodiments, the weights
are based upon
total weight of all enantiomers or stereoisomers of the compound.
[00116] As used herein and unless otherwise indicated, the term
"enantiomerically pure R-
compound" refers to at least about 80% by weight R-compound and at most about
20% by
weight S-compound, at least about 90% by weight R-compound and at most about
10% by
weight S-compound, at least about 95% by weight R-compound and at most about
5% by weight
S-compound, at least about 99% by weight R-compound and at most about 1% by
weight 5-
compound, at least about 99.9% by weight R-compound or at most about 0.1% by
weight 5-
compound. In certain embodiments, the weights are based upon total weight of
compound.
[00117] As used herein and unless otherwise indicated, the term
"enantiomerically pure 5-
compound" or "S-compound" refers to at least about 80% by weight S-compound
and at most
about 20% by weight R-compound, at least about 90% by weight S-compound and at
most about
10% by weight R-compound, at least about 95% by weight S-compound and at most
about 5%
by weight R-compound, at least about 99% by weight S-compound and at most
about 1% by
weight R-compound or at least about 99.9% by weight S-compound and at most
about 0.1% by
weight R-compound. In certain embodiments, the weights are based upon total
weight of
compound.
[00118] In the compositions provided herein, an enantiomerically pure
compound or a
pharmaceutically acceptable salt, solvate, hydrate or prodrug thereof can be
present with other
active or inactive ingredients. For example, a pharmaceutical composition
comprising
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enantiomerically pure R-compound can comprise, for example, about 90%
excipient and about
10% enantiomerically pure R-compound. In certain embodiments, the
enantiomerically pure R-
compound in such compositions can, for example, comprise, at least about 95%
by weight R-
compound and at most about 5% by weight S-compound, by total weight of the
compound. For
example, a pharmaceutical composition comprising enantiomerically pure S-
compound can
comprise, for example, about 90% excipient and about 10% enantiomerically pure
S-compound.
In certain embodiments, the enantiomerically pure S-compound in such
compositions can, for
example, comprise, at least about 95% by weight S-compound and at most about
5% by weight
R-compound, by total weight of the compound. In certain embodiments, the
active ingredient
can be formulated with little or no excipient or carrier.
[00119] The compounds of this invention may possess one or more asymmetric
centers;
such compounds can therefore be produced as individual (R)- or (S)-
stereoisomers or as
mixtures thereof
[00120] Unless indicated otherwise, the description or naming of a
particular compound in
the specification and claims is intended to include both individual
enantiomers and mixtures,
racemic or otherwise, thereof The methods for the determination of
stereochemistry and the
separation of stereoisomers are well-known in the art.
[00121] One having ordinary skill in the art of organic synthesis will
recognize that the
maximum number of heteroatoms in a stable, chemically feasible heterocyclic
ring, whether it is
aromatic or non aromatic, is determined by the size of the ring, the degree of
unsaturation and the
valence of the heteroatoms. In general, a heterocyclic ring may have one to
four heteroatoms so
long as the heteroaromatic ring is chemically feasible and stable.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
[00122] In certain aspects, provided herein are pharmaceutical compositions
comprising a
bolaamphiphile vesicle complex.
[00123] In certain aspects, the bolaamphiphile vesicle complexes comprise
one or more
bolaamphiphilic compounds and a biologically active compound. In a particular
embodiment, the
biologically active compound is siRNA.
[00124] In further aspects, provided herein are novel siRNA and
bolamphiphilic complex
comprising siRNA and one or more bolaamphiphilic compounds.
[00125] In further aspects, provided herein are novel formulations of siRNA
with
bolaamphiphilic compounds or with bolaamphiphile vesicles.
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[00126] In another aspect, provided here are methods of delivering siRNA
into animal or
human cell comprising the step of administering to the animal or human a
pharmaceutical
composition comprising a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA. In
one
embodiment, the cell is brain cell, liver cell, gall bladder cell, or a lung
cell. In other
embodiments, the cells are are cells of a lymph node, a CD4+ lymphocyte, or a
cell of the
mononuclear phagocyte system, including, without limitation, a monocyte,
macrophage, a
resident brain microglial cell and a dendritic cell.
[00127] In another aspect, provided here are methods of delivering siRNA
into animal or
human organs comprising the step of administering to the animal or human a
pharmaceutical
composition comprising of a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA. In
one
embodiment, the organ is brain, liver, gall bladder, or a lung.
[00128] In one embodiment, the bolaamphiphilic complex comprises one
bolaamphiphilic
compound. In another embodiment, the bolaamphiphilic complex comprises two
bolaamphiphilic
compounds.
[00129] In one embodiment, the bolaamphiphilic compound consists of two
hydrophilic
headgroups linked through a long hydrophobic chain. In another embodiment, the
hydrophilic
headgroup is an amino containing group. In a specific embodiment, the
hydrophilic headgroup is
a tertiary or quaternary amino containing group.
[00130] In one particular embodiment, the bolaamphiphilic compound is a
compound
according to formula I:
HG2 ¨L1 ¨HG1
I
or a pharmaceutically acceptable salt, solvate, hydrate, prodrug,
stereoisomer, tautomer, isotopic
variant, or N¨oxide thereof, or a combination thereof;
wherein:
each HG1 and HG2 is independently a hydrophilic head group; and
L1 is alkylene, alkenyl, heteroalkylene, or heteroalkenyl linker;
unsubstituted or
substituted with Ci-C20 alkyl, hydroxyl, or oxo.
[00131] In one embodiment, the pharmaceutically acceptable salt is a
quaternary
ammonium salt.
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[00132] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
Li is heteroalkylene, or heteroalkenyl linker comprising C, N, and 0 atoms;
unsubstituted or
substituted with Ci-C20 alkyl, hydroxyl, or oxo.
[00133] In another embodiment, with respect to the bolaamphiphilic compound
of formula
I, Li is
¨0-L2¨C(0)-0-(CH2).4-0-C(0)-L3-0-, or
¨0-L2¨C(0)-0-(CH2).5-0-C(0)-(CH2)06-,
and wherein each L2 and L3 is C4-C20 alkenyl linker; unsubstituted or
substituted with C1-C8
alkyl or hydroxy;
and n4, n5, and n6 is independently an integer from 4-20.
[00134] In one embodiment, each L2 and L3 is independently ¨C(R1)-C(0H)-CH2-
(CH=CH)-(CH2).2-; Ri is Ci-C8 alkyl, and n7 is independently an integer from 4-
20.
[00135] In another embodiment, with respect to the bolaamphiphilic compound
of formula
I, Li is ¨0-(CH2).1-0-C(0)-(CH2).2-C(0)-0-(CH2).3-0-.
[00136] In another embodiment, with respect to the bolaamphiphilic compound
of
formula I, Li is
,ZI
r
0 0
Linker AA
0 0
Zi Z2
k
Linker BB
R
____ ORia ib 0 _________
0 0
R2a Zi Z2 1( )12R2b
Linker CC
or
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__________________________ ORla
0
R2a ')n9 ZI
Linker DD
wherein:
each Z1 and Z2 is independently -C(R3)2-, -N(R3)- or -0-;
each R", R3, and R4 is independently H or Ci-C8 alkyl;
each R2a and R2b is independently H, Ci-C8 alkyl, OH, or alkoxy;
each n8, n9, n11, and n12 is independently an integer from 1-20;
n10 is an integer from 2-20; and
each dotted bond is independently a single or a double bond.
and wherein each methylene carbon is unsubstituted or substituted with Ci-C4
alkyl; and each
nl, n2, and n3 is independently an integer from 4-20.
[00137] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
the bolaamphiphilic compound is a compound according to formula II, III, IV,
V, or VI:
-)nio Z2
HG2 _______________________ (-')n9 (-),,õ HG'
0 0
0 0
HG2 ____________________ -')n9 22 --)nl 1 HG1
III
HG2 ¨0 Rlb
0¨HG1
0 0
R2a( )n8 Z1 Z2 R2b
Iv
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HG2-0Rla
0
.=,( µ')n10
R2a ( )1-1--8 - - ¨HG1
V ,or
HG2-0Rla
0
.... ,.., R4
--- - -
R2a (')n8 n9 Zi
VI
or a pharmaceutically acceptable salt, solvate, hydrate, prodrug,
stereoisomer, tautomer, isotopic
variant, or N¨oxide thereof, or a combination thereof;
wherein:
each HG1 and HG2 is independently a hydrophilic head group;
each Z1 and Z2 is independently -C(R3)2-, -N(R3)- or ¨0-;
¨ lb,
each Ria, xR3, and R4 is independently H or Ci-C8 alkyl;
each R2a and R2b is independently H, Ci-C8 alkyl, OH, alkoxy, or 0-HG1 or 0-
HG2;
each n8, n9, n11, and n12 is independently an integer from 1-20;
n10 is an integer from 2-20; and
each dotted bond is independently a single or a double bond.
[00138] In one embodiment, with respect to the bolaamphiphilic compound of
formula II,
III, IV, V, or VI, each n9 and n11 is independently an integer from 2-12. In
another embodiment,
n9 and n11 is independently an integer from 4-8. In a particular embodiment,
each n9 and n11 is
7 or 11.
[00139] In one embodiment, with respect to the bolaamphiphilic compound of
formula II,
III, IV, V, or VI, each n8 and n12 is independently 1, 2, 3, or 4. In a
particular embodiment, each
n8 and n12 is 1.
[00140] In one embodiment, with respect to the bolaamphiphilic compound of
formula II,
III, IV, V, or VI, each R2a and R2b is independently H, OH, or alkoxy. In
another embodiment,
each R2a and R2b is independently H, OH, or OMe. In another embodiment, each
R2a and R2b is
independently-O-HG1 or 0-HG2. In a particular embodiment, each R2a and R2b is
OH.
[00141] In one embodiment, with respect to the bolaamphiphilic compound of
formula II,
III, IV, V, or VI, each Ria and Rib is independently H, Me, Et, n-Pr, i-Pr, n-
Bu, i-Bu, sec-Bu, n-
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pentyl, isopentyl, n-hexyl, n-heptyl, or n-octyl. In a particular embodiment,
each Ria and Rib is
independently n-pentyl.
[00142] In one embodiment, with respect to the bolaamphiphilic compound of
formula II,
III, IV, V, or VI, each dotted bond is a single bond. In another embodiment,
each dotted bond is a
double bond.
[00143] In one embodiment, with respect to the bolaamphiphilic compound of
formula II,
III, IV, V, or VI, n10 is an integer from 2-16. In another embodiment, n10 is
an integer from 2-
12. In a particular embodiment, n10 is 2, 4, 6, 8, 10, 12, or 16.
[00144] In one embodiment, with respect to the bolaamphiphilic compound of
formula IV,
R4 is H, Me, Et, n-Pr, i-Pr, n-Bu, i-Bu, sec-Bu, n-pentyl, or isopentyl. In
another embodiment, R4
is Me, or Et. In a particular embodiment, R4 is Me.
[00145] In one embodiment, with respect to the bolaamphiphilic compound of
formula II,
III, IV, V, or VI, each Zi and Z2 is independently C(R3)2-, or -N(R3)-. In
another embodiment,
each Zi and Z2 is independently C(R3)2-, or -N(R3)-; and each R3 is
independently H, Me, Et, n-
Pr, i-Pr, n-Bu, i-Bu, sec-Bu, n-pentyl, or isopentyl. In a particular
embodiment, R3 is H.
[00146] In one embodiment, with respect to the bolaamphiphilic compound of
formula II,
III, IV, V, or VI, each Zi and Z2 is ¨0-.
[00147] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, or IV, each HG1 and HG2 is independently selected from:
0 0 0 R8
(A\
/k iirli X
( \ A
/k iir11 ))X
X
13
'
0
( A \
k iirll 0
( irml
)X and
(')ri13 0 0
wherein:
X is ¨NR5aR5b, or ¨N+R5aR5bR5e; each R5a, and R5b is independently H or
substituted or
unsubstituted Ci-C20 alkyl or R5a and R5b may join together to form an N
containing
substituted or unsubstituted heteroaryl, or substituted or unsubstituted
heterocyclyl;
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each R5' is independently substituted or unsubstituted Ci-C20 alkyl; each R8
is independently
H, substituted or unsubstituted Ci-C20 alkyl, alkoxy, or carboxy;
ml is 0 or 1; and
each n13, n14, and n15 is independently an integer from 1-20.
[00148] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, or IV, HG1 and HG2 are as defined above, and each ml is 0.
[00149] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, or IV, HG1 and HG2 are as defined above, and each ml is 1.
[00150] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, or IV, HG1 and HG2 are as defined above, and each n13 is 1 or 2.
[00151] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, or IV, HG1 and HG2 are as defined above, and each n14 and n15 is
independently 1, 2, 3,
4, or 5. In another embodiment, each n14 and n15 is independently 2 or 3.
[00152] In one particular embodiment, the bolaamphiphilic compound is a
compound
according to formula VIIa, VIIb, VIIc, or VIId:
c) 0 0
0
VI la
(:) 0
0 _ (inio
H 0 N( N 0 Ho 0
VI lb
0 y, )3 Sir 0 0 ) 3 0 X
0 '')
0 H0 N()7 n10-N)*L()70 Ho 0
VI lc
or
( `")i
0 0
VI Id
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or a pharmaceutically acceptable salt, solvate, hydrate, prodrug,
stereoisomer, tautomer, isotopic
variant, or N¨oxide thereof, or a combination thereof;
wherein:
each X is ¨NR5aR5b, or ¨N+R5aR5bR5'; each R5a, and R5b is independently H or
substituted or
unsubstituted C1-C20 alkyl or R5a and R5b may join together to form an N
containing
substituted or unsubstituted heteroaryl, or substituted or unsubstituted
heterocyclyl;
each R5' is independently substituted or unsubstituted Ci-C20 alkyl;;
n10 is an integer from 2-20; and
each dotted bond is independently a single or a double bond.
[00153] In
another particular embodiment, the bolaamphiphilic compound is a compound
according to formula Villa, VIIIb, VIIIc, or VIIId:
x.rc) 0 0
x
Villa ,
Xr0..........õ--...õ,,,,,....., 0
HO-Th( '"-)7 O '0 ( '"--)7.-- - - -------------
OH 0
VIllb
'
x y(--')3 lr \W 0 0 0 (
"=)3 0 x
0 i \ )L ( ¨)n10 [I ---
--ir
0
HO-Thk /7 0-''' .-'0 ( '---)7---- - - ---
--."----0 IP 0
VIIIc
or
X,......., ....,,o...,....)____7()nlyo''',,
N411
0 0
VII Id
or a pharmaceutically acceptable salt, solvate, hydrate, prodrug,
stereoisomer, tautomer, isotopic
variant, or N¨oxide thereof, or a combination thereof;
wherein:
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each X is ¨NR5aR5b, or ¨N+R5aR5bR5e; each R5a, and R5b is independently H or
substituted or
unsubstituted C1-C20 alkyl or R5a and R5b may join together to form an N
containing
substituted or unsubstituted heteroaryl, or substituted or unsubstituted
heterocyclyl;
each R5' is independently substituted or unsubstituted Ci-C20 alkyl;;
n10 is an integer from 2-20; and
each dotted bond is independently a single or a double bond.
[00154] In another particular embodiment, the bolaamphiphilic compound is a
compound
according to formula IXa, IXb, or IXc:
xr0..õ.....,,,..--,õ,... 0 0
0 Nnio )-x
H H
IXa ,
XThrO.,.õ,,,,...õ---,õ_,,- 0 0 0
H H
IXb ,
( )31((:)\ 0 0 0
0
H H
IXc
or a pharmaceutically acceptable salt, solvate, hydrate, prodrug,
stereoisomer, tautomer, isotopic
variant, or N¨oxide thereof, or a combination thereof;
wherein:
each X is ¨NR5aR5b, or ¨N+R5aR5bR5e; each R5a, and R5b is independently H or
substituted or
unsubstituted C1-C20 alkyl or R5a and R5b may join together to form an N
containing
substituted or unsubstituted heteroaryl, or substituted or unsubstituted
heterocyclyl;
each R5' is independently substituted or unsubstituted Ci-C20 alkyl;;
n10 is an integer from 2-20; and
each dotted bond is independently a single or a double bond.
[00155] In another particular embodiment, the bolaamphiphilic compound is a
compound
according to formula Xa, Xb, or Xc:
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x=rC) 0 0
0X
HO
Xa
X(C) 0 0 0
0 (-)nio
C) (.-)3 OX
Xb
)3,0 0 0
¨
0 oHO-Th(,--)7 ()ni
)LO
Xc
or a pharmaceutically acceptable salt, solvate, hydrate, prodrug,
stereoisomer, tautomer, isotopic
variant, or N¨oxide thereof, or a combination thereof;
wherein:
each X is ¨NR5aR5b, or ¨N+R5aR5bR5'; each R5a, and R5b is independently H or
substituted or
unsubstituted C1-C20 alkyl or R5a and R5b may join together to form an N
containing
substituted or unsubstituted heteroaryl, or substituted or unsubstituted
heterocyclyl;
each R5' is independently substituted or unsubstituted Ci-C20 alkyl;;
n10 is an integer from 2-20; and
each dotted bond is independently a single or a double bond.
[00156] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXe, or Xa-Xc, each dotted bond is a single bond.
In another
embodiment, each dotted bond is a double bond.
[00157] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXe, or Xa-Xc, n10 is an integer from 2-16.
[00158] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXe, or Xa-Xc, n10 is an integer from 2-12.
[00159] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXe, or Xa-Xc, n10 is 2, 4, 6, 8, 10, 12, or 16.
[00160] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXe, or Xa-Xc, each R5a, R5b, and R5' is
independently substituted or
unsubstituted Ci-C20 alkyl.
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[00161] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, each R5a, R5b, and R5e is
independently unsubstituted
C1-C20 alkyl.
[00162] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, one of R5a, R5b, and R5e is Ci-C20
alkyl substituted
with ¨0C(0)R6; and R6 is Ci-C20 alkyl.
[00163] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, two of R5a, R5b, and R5e are
independently Ci-C20
alkyl substituted with ¨0C(0)R6; and R6 is Ci-C20 alkyl. In one embodiment, R6
is Me, Et, n-Pr,
i-Pr, n-Bu, i-Bu, sec-Bu, n-pentyl, isopentyl, n-hexyl, n-heptyl, or n-octyl.
In a particular
embodiment, R6 is Me.
[00164] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, one of R5a, R5b, and R5e is Ci-C20
alkyl substituted
with amino, alkylamino or dialkylamino.
[00165] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, two of R5a, R5b, and R5e are
independently Ci-C20
alkyl substituted with amino, alkylamino or dialkylamino.
[00166] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, R5a, and R5b together with the N
they are attached to
form substituted or unsubstituted heteroaryl.
[00167] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, R5a, and R5b together with the N
they are attached to
form substituted or unsubstituted pyridyl.
[00168] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, R5a, and R5b together with the N
they are attached to
form substituted or unsubstituted monocyclic or bicyclic heterocyclyl.
[00169] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, X is substituted or unsubstituted
N
[00170] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, X is
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N
,
substituted with one or more groups selected from alkoxy, acetyl, and
substituted or
unsubstituted Ph.
[00171] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-D(c, or Xa-Xc, X is
0
N
ci--
Mµ
=
F .
[00172] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-D(c, or Xa-Xc, X is ¨NMe2 or ¨N+Me3.
[00173] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-D(c, or Xa-Xc, X is ¨N(Me)-CH2CH2-0Ac or ¨N+(Me)2.-
CH2CF12-
0Ac.
[00174] In one embodiment, with respect to the bolaamphiphilic compound of
formula
VIIa-VIId, VIIIa-VIIId, IXa-IXc, or Xa-Xc, X is a chitosanyl group; and the
chitosanyl group is
a poly-(D)glucosaminyl group with MW of 3800 to 20,000 Daltons, and is
attached to the core
via N.
[00175] In one embodiment, the chitosanyl group is
OH OH
H¨[ ¨04 ______________________ n
_____________________________ ._, r ,0
0
12¨OH
NH H
/N
R7a
=
,
and wherein each pl and p2 is independently an integer from 1-400; and each
R7a is H or acyl.
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[00176] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
a pharmaceutically acceptable salt.
[00177] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
in a form of a quaternary salt.
[00178] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
in a form of a quaternary salt with pharmaceutically acceptable alkyl halide
or alkyl tosylate.
[00179] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
any one of the bolaambphilic compounds listed in Table 1.
[00180] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
GLH-19.
[00181] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
GLH-20.
[00182] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
GLH-16.
[00183] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
GLH-26, 29, or 41.
[00184] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
other than Comound ID GLH-16, GLH-19, GLH-20, GLH-26, GLH-29, or GLH-41.
[00185] In one embodiment, with respect to the bolaamphiphilic compound of
formula I,
II, III, IV, V, VI, VIIa-VIIc, VIIIa-VIIIc, IXa-D(c and Xa-Xc, the
bolaamphiphilic compound is
other than Comound ID GLH-6, GLH-8, GLH-12, GLH-13, GLH-13a, or GLH-49 to GLH-
54
(all can be used as intermediates for bolaamphiphiles).
[00186] In another specific aspect, provided herein are composition of
novel
bolaamphiphilic compounds, wherein the bolaamphiphilic compound is selected
from the
bolaambphilic compounds listed in Table 1. In one embodiment, with respect to
the
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bolaamphiphilic compound, the bolaamphiphilic compound is other than Comound
ID GLH-16,
GLH-19, GLH-20, GLH-26, GLH-29, or GLH-41. In another embodiment, with respect
to the
bolaamphiphilic compound, the compound is other than compound with ID GLH-3,
GLH-4,
GLH-5, or GLH-21.
[00187] In one particular embodiment, bolaamphiphilic compound is selected
from the
bolaambphilic compounds listed in Table 1, and the compound is compound with
ID GLH-7,
GLH-9, GLH-10, GLH-11, GLH-14, GLH-15, GLH-17, GLH-18, GLH-22, GLH-23, GLH-24,
GLH-25, GLH-27, GLH-28, GLH-30 to GLH-48, GLH-55, GLH-56, or GLH-57.
[00188] In another specific aspect, provided herein are methods for
delivering siRNA
across the cell membrane. In one embodiment, the cell is brain cell, liver
cell, gall bladder cell, or
a lung cell. In other specific aspects, the cells are are cells of a lymph
node, a CD4+ lymphocyte,
or a cell of the mononuclear phagocyte system, including, without limitation,
a monocyte,
macrophage, a resident brain microglial cell and a dendritic cell.
[00189] In another aspect, provided here are methods of delivering siRNA
into animal or
human brain comprising the step of administering to the animal or human a
pharmaceutical
composition comprising of a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA.
[00190] In another aspect, provided here are methods of delivering siRNA
into animal or
human liver comprising the step of administering to the animal or human a
pharmaceutical
composition comprising of a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA.
[00191] In another aspect, provided here are methods of delivering siRNA
into animal or
human lungs comprising the step of administering to the animal or human a
pharmaceutical
composition comprising of a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA.
[00192] In another aspect, provided here are methods of delivering siRNA
into animal or
human gall bladder comprising the step of administering to the animal or human
a
pharmaceutical composition comprising of a bolaamphiphile vesicle complex; and
wherein the
bolaamphiphile vesicle complex comprises one or more bolaamphiphilic compounds
and siRNA.
[00193] In another specific aspect, provided herein are nano-particles,
comprising one or
more bolaamphiphilic compounds and siRNA. In one embodiment, the
bolaamphiphilic
compounds and siRNA are encapsulated within the nano-particle.
[00194] In particular embodiments, polynucleotides selected from DNA or RNA
fragments are delivered by the nanoparticles of the invention. In a more
particular embodiment,
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the polynucleotide is a small interfering RNA (siRNA), a double-stranded RNA
molecule of 20-
25 nucleotides. siRNAs play a variety of roles in biology. Most notably,
siRNAs are involved in
the RNA interference (RNAi) pathway, where they interfere with the expression
of a specific
gene. In addition to their role in the RNAi pathway, siRNAs also act in RNAi-
related pathways,
e.g., as an antiviral mechanism or in shaping the chromatin structure of a
genome. Some non
limiting examples for target genes, or biological pathways which can be
interfered by siRNA are
epidermal growth factor receptor variant III gene, which is expressed in 40-
50% of gliomas, and
the phosphoinositide 3-kinase (PI3K)/Akt pathway, which plays a crucial role
in
medulloblastoma biology. In other aspects of this embodiment, the
polynucleotide is a DNA-
RNA hybrid molecule.
[00195] In certain embodiments, the bolaamphiphile vesicle complexes
comprise one or
more bolaamphiphilic compounds and the biologically active compound is a siRNA
that is a
mixture of two or more siRNA, wherein at least one siRNA is directed to a
first target, and at
least one siRNA is directed to a second target.
[00196] In further embodiments, provided herein are novel siRNA and
bolamphiphilic
vesicle complex comprising siRNA and one or more bolaamphiphilic compounds.
[00197] In further embodiments, provided herein are novel formulations of
siRNA with
bolaamphiphilic compounds or with bolaamhphilic vesicles.
[00198] In another embodiment, provided here are methods of delivering
siRNA into
animal or human cells.
[00199] In an additional embodiment of the disclosure is directed to
delivery of siRNA-
bolaamphiphile vesicle complexes or siRNA-bolaamphiphilic vesicle complexes
into animals or
human wherein the bolaamphiphile vesicle complex comprises one or more
bolaamphiphilic
compounds and siRNA.
[00200] In another aspect, provided here are methods of delivering siRNA
into animal or
human cell comprising the step of administering to the animal or human a
pharmaceutical
composition comprising of a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA. In
one
embodiment, the cell is brain cell, liver cell, gall bladder, or a lung cell.
In other embodiments,
the cells are are cells of a lymph node, a CD4+ lymphocyte, or a cell of the
mononuclear
phagocyte system, including, without limitation, a monocyte, macrophage, a
resident brain
microglial cell and a dendritic cell. In a still further emobidment, the cell
is a cancer cell.
[00201] In another aspect, provided here are methods of delivering siRNA
into animal or
human organs comprising the step of administering to the animal or human a
pharmaceutical
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composition comprising of a bolaamphiphile vesicle complex; and wherein the
bolaamphiphile
vesicle complex comprises one or more bolaamphiphilic compounds and siRNA. In
one
embodiment, the organ is brain, liver, gall bladder, a lymph node or a lung.
In certain aspects of
this emobidment, the siRNA is delivered to a tumor.
[00202] In a further embodiment the active agent is an RNA-DNA heteroduplex
with
properties of siRNA molecules. In certain aspects of this embodiment, the
bolaamphiphile
vesicle complexes comprise one or more bolaamphiphilic compounds and the
biologically active
compound is a siRNA that is a mixture of two or more siRNA or a mixture
comprising at least
one siRNA and one RNA-DNA duplex, wherein at least one siRNA or RNA-DNA duplex
is
directed to a first target, and at least one siRNA or RNA-DNA duplex is
directed to a second
target.
[00203] In certain embodiments, the target is a promoter. In other
emodiments, the first
and second targets are sequences of separate and distinct genes.
[00204] In another specific aspect, provided herein are pharmaceutical
compositions,
comprising a nano-sized particle comprising one or more bolaamphiphilic
compounds and
siRNA; and a pharmaceutically acceptable carrier.
[00205] In another specific aspect, provided herein are methods for
treatment or diagnosis
of diseases or disorders selected from cancer such as breast cancer, prostate
cancer and brain
tumors using the nano-particles, pharmaceutical compositions or formulations
of the present
invention.
[00206] The Derivatives and Precursors disclosed can be prepared as
illustrated in the
Schemes provided herein. The syntheses can involve initial construction of,
for example,
vernonia oil or direct functionalization of natural derivatives by organic
synthesis manipulations
such as, but not limiting to, epoxide ring opening. In those processes
involving oxiranyl ring
opening, the epoxy group is opened by the addition of reagents such as
carboxylic acids or
organic or inorganic nucleophiles. Such ring opening results in a mixture of
two products in
which the new group is introduced at either of the two carbon atoms of the
epoxide moiety. This
provides beta substituted alcohols in which the substitution position most
remote from the CO
group of the main aliphatic chain of the vernonia oil derivative is
arbitrarily assigned as position
1, while the neighboring substituted carbon position is designated position 2.
For simplicity
purposes only, the Derivatives and Precursors shown herein may indicate
structures with the
hydroxy group always at position 2 but the Derivatives and Precursors wherein
the hydroxy is at
position 1 are also encompassed by the invention. Thus, a radical of the
formula --CH(OH)--
CH(R)-- refers to the substitution of --OH at either the carbon closer to the
CO group, designated
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position 2 or to the carbon at position 1. Moreover, with respect to the
preparation of
symmetrical bolaamphiphiles made via introducing the head groups through an
epoxy moiety
(e.g., as in vernolic acid) or a double bond (-C=C-) as in mono unsaturated
fatty acids (e.g., oleic
acid) a mixture of three different derivatives will be produced. In certain
embodiments, vesicles
are prepared using the mixture of unfractionated positional isomers. In one
aspect of this
embodiment, where one or more bolas are prepared from vernolic acid, and in
which a hydroxy
group as well as the head group introduced through an epoxy or a fatty acid
with the head group
introduced through a double bond (-C=C-), the bola used in vesicle preparation
can actually be a
mixture of three different positional isomers.
[00207] In other embodiments, the three different derivatives are isolated.
Accordingly,
the vesicles disclosed herein can be made from a mixture of the three isomers
of each derivative
or, in other embodiments, the individual isomers can be isolated and used for
preparation of
vesicles.
[00208] Symmetrical bolaamphiphiles can form relatively stable self
aggregate vesicle
structures by the use of additives such as cholesterol and cholesterol
derivatives (e.g., cholesterol
hemisuccinate, cholesterol oleyl ether, anionic and cationic derivatives of
cholesterol and the
like), or other additives including single headed amphiphiles with one, two or
multiple aliphatic
chains such as phospholipids, zwitterionic, acidic, or cationic lipids.
Examples of zwitterionic
lipids are phosphatidylcholines, phosphatidylethanol amines and
sphingomyelins. Examples of
acidic amphiphilic lipids are phosphatidylglycerols, phosphatidylserines,
phosphatidylinositols,
and phosphatidic acids. Examples of cationic amphipathic lipids are diacyl
trimethylammonium
propanes, diacyl dimethylammonium propanes, and stearylamines cationic
amphiphiles such as
spermine cholesterol carbamates, and the like, in optimum concentrations which
fill in the larger
spaces on the outer surfaces, and/or add additional hydrophilicity to the
particles. Such additives
may be added to the reaction mixture during formation of nanoparticles to
enhance stability of
the nanoparticles by filling in the void volumes of in the upper surface of
the vesicle membrane.
[00209] Stability of nano vesicles according to the present disclosure can
be demonstrated
by dynamic light scattering (DLS) and transmission electron microscopy (TEM).
For example,
suspensions of the vesicles can be left to stand for 1, 5, 10, and 30 days to
assess the stability of
the nanoparticle solution/suspension and then analyzed by DLS and TEM.
[00210] The vesicles disclosed herein may encapsulate within their core the
active agent,
which in particular embodiments is selected from peptides, proteins,
nucleotides and or non-
polymeric agents. In certain embodiments, the active agent is also associated
via one or more
non-covalent interactions to the vesicular membrane on the outer surface
and/or the inner surface,
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optionally as pendant decorating the outer or inner surface, and may further
be incorporated into
the membrane surrounding the core. In certain aspects, biologically active
peptides, proteins,
nucleotides or non-polymeric agents that have a net electric charge, may
associate ionically with
oppositely charged headgroups on the vesicle surface and/or form salt
complexes therewith.
[00211] In particular aspects of these embodiments, additives which may be
bolaamphiphiles or single headed amphiphiles, comprise one or more branching
alkyl chains
bearing polar or ionic pendants, wherein the aliphatic portions act as anchors
into the vesicle's
membrane and the pendants (e.g., chitosan derivatives or polyamines or certain
peptides)
decorate the surface of the vesicle to enhance penetration through various
biological barriers such
as the intestinal tract and the BBB, and in some instances are also
selectively hydrolyzed at a
given site or within a given organ. The concentration of these additives is
readily adjusted
according to experimental determination.
[00212] In certain embodiments, the oral formulations of the present
disclosure comprise
agents that enhance penetration through the membranes of the GI tract and
enable passage of
intact nanoparticles containing the drug. These agents may be any of the
additives mentioned
above and, in particular aspects of these embodiment, include chitosan and
derivatives thereof,
serving as vehicle surface ligands, as decorations or pendants on the
vesicles, or the agents may
be excipients added to the formulation.
[00213] In other embodiments, the nanoparticles and vesicles disclosed
herein may
comprise the fluorescent marker carboxyfluorescein (CF) encapsulated therein
while in particular
aspects, the nanoparticle and vesicles of the present disclosure may be
decorated with one or
more of PEG, e.g. PEG2000-vernonia derivatives as pendants. For example, two
kinds of PEG-
vernonia derivatives can be used: PEG-ether derivatives, wherein PEG is bound
via an ether
bond to the oxygen of the opened epoxy ring of, e.g., vernolic acid and PEG-
ester derivatives,
wherein PEG is bound via an ester bond to the carboxylic group of, e.g.,
vernolic acid.
[00214] In other embodiments, vesicles, made from synthetic amphiphiles, as
well as
liposomes, made from synthetic or natural phospholipids, substantially (or
totally) isolate the
therapeutic agent from the environment allowing each vesicle or liposome to
deliver many
molecules of the therapeutic agent. Moreover, the surface properties of the
vesicle or liposome
can be modified for biological stability, enhanced penetration through
biological barriers and
targeting, independent of the physico-chemical properties of the encapsulated
drug.
[00215] In still other embodiments, the headgroup is selected from: (i)
choline or
thiocholine, 0-alkyl, N-alkyl or ester derivatives thereof; (ii) non-aromatic
amino acids with
functional side chains such as glutamic acid, aspartic acid, lysine or
cysteine, or an aromatic
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amino acid such as tyrosine, tryptophan, phenylalanine and derivatives thereof
such as levodopa
(3,4-dihydroxy-phenylalanine) and p-aminophenylalanine; (iii) a peptide or a
peptide derivative
that is specifically cleaved by an enzyme at a diseased site selected from
enkephalin, N-acetyl-
ala-ala, a peptide that constitutes a domain recognized by beta and gamma
secretases, and a
peptide that is recognized by stromelysins; (iv) saccharides such as glucose,
mannose and
ascorbic acid; and (v) other compounds such as nicotine, cytosine, lobeline,
polyethylene glycol,
a cannabinoid, or folic acid.
[00216] In further embodiments, nano-sized particle and vesicles disclosed
herein further
comprise at least one additive for one or more of targeting purposes,
enhancing permeability and
increasing the stability the vesicle or particle. Such additives, in
particular aspects, may selected
from from: (i) a single headed amphiphilic derivative comprising one, two or
multiple aliphatic
chains, preferably two aliphatic chains linked to a midsection/spacer region
such as --NH--
(CH2)2--N--(CH2)2--N--, or --O--(CH2)2--N--(CH2)2--O--, and a sole headgroup,
which may be a
selectively cleavable headgroup or one containing a polar or ionic selectively
cleavable group or
moiety, attached to the N atom in the middle of said midsection. In other
asepcts, the additive
can be selected from among cholesterol and cholesterol derivatives such as
cholesteryl
hemmisuccinate; phospholipids, zwitterionic, acidic, or cationic lipids;
chitosan and chitosan
derivatives, such as vernolic acid-chitosan conjugate, quaternized chitosan,
chitosan-
polyethylene glycol (PEG) conjugates, chitosan-polypropylene glycol (PPG)
conjugates, chitosan
N-conjugated with different amino acids, carboxyalkylated chitosan, sulfonyl
chitosan,
carbohydrate-branched N-(carboxymethylidene) chitosan and N-(carboxymethyl)
chitosan;
polyamines such as protamine, polylysine or polyarginine; ligands of specific
receptors at a target
site of a biological environment such as nicotine, cytisine, lobeline, 1-
glutamic acid MK801,
morphine, enkephalins, benzodiazepines such as diazepam (valium) and librium,
dopamine
agonists, dopamine antagonists tricyclic antidepressants, muscarinic agonists,
muscarinic
antagonists, cannabinoids and arachidonyl ethanol amide; polycationic polymers
such as
polyethylene amine; peptides that enhance transport through the BBB such as OX
26,
transferfins, polybrene, histone, cationic dendrimer, synthetic peptides and
polymyxin B
nonapeptide (PMBN); monosaccharides such as glucose, mannose, ascorbic acid
and derivatives
thereof; modified proteins or antibodies that undergo absorptive-mediated or
receptor-mediated
transcytosis through the blood-brain barrier, such as bradykinin B2 agonist
RMP-7 or
monoclonal antibody to the transferrin receptor; mucoadhesive polymers such as
glycerides and
steroidal detergents; and Ca2+ chelators. The aforementioned head groups on
the additives
designed for one or more of targeting purposes and enhancing permeability may
also be a head
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group, preferably on an asymmetric bolaamphiphile wherein the other head group
is another
moiety, or the head group on both sides of a symmetrical bolaamphiphile. In a
further
embodiment the bolaamphiphile head groups that comprise the vesicles membranes
can interact
with the active agents to be encapsulated to be delivered in to the brain and
brain sites, and or
other targeted sites, by ionic interactions to enhance the % encapsulation via
complexation and
well as passive encapsulation within the vesicles core. Further the
formulation may contain other
additives within the vehicles membranes to further enhance the degree of
encapsulation of the
active agents by interactions other than ionic interactions such as polar or
hydrophobic
interactions
[00217] In other embodiments, nano-sized particle and vesicles discloser
herein may
comprises at least one biologically active agent is selected from: (i) a
natural or synthetic peptide
or protein such as analgesics peptides from the enkephalin class, insulin,
insulin analogs,
oxytocin, calcitonin, tyrotropin releasing hormone, follicle stimulating
hormone, luteinizing
hormone, yasopressin and yasopressin analogs, catalase, interleukin-II,
interferon, colony
stimulating factor, tumor necrosis factor (TNF), melanocyte-stimulating
hormone, superoxide
dismutase, glial cell derived neurotrophic factor (GDNF) or the Gly-Leu-Phe
(GLF) families; (ii)
nucleosides and polynucleotides selected from DNA or RNA molecules such as
small interfering
RNA (siRNA) or a DNA plasmid; (iii) antiviral and antibacterial; (iv)
antineoplastic and
chemotherapy agents such as cyclosporin, doxorubicin, epirubicin, bleomycin,
cisplatin,
carboplatin, yinca alkaloids, e.g. yincristine, Podophyllotoxin, taxanes, e.g.
Taxol and Docetaxel,
and topoisomerase inhibitors, e.g. irinotecan, topotecan.
[00218] Additional embodiments within the scope provided herein are set
forth in
non-limiting fashion elsewhere herein and in the examples. It should be
understood that these
examples are for illustrative purposes only and are not to be construed as
limiting in any manner.
PHARMACEUTICAL COMPOSITIONS
[00219] In another aspect, the invention provides a pharmaceutical
composition
comprising a pharmaceutically acceptable carrier and a pharmaceutically
effective amount of a
compound of Formula I or a complex thereof
[00220] When employed as pharmaceuticals, the compounds provided herein are
typically
administered in the form of a pharmaceutical composition. Such compositions
can be prepared
in a manner well known in the pharmaceutical art and comprise at least one
active compound.
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[00221] In certain embodiments, with respect to the pharmaceutical
composition, the
carrier is a parenteral carrier, oral or topical carrier.
[00222] The present invention also relates to a compound or pharmaceutical
composition
of compound according to Formula I; or a pharmaceutically acceptable salt or
solvate thereof for
use as a pharmaceutical or a medicament.
[00223] Generally, the compounds provided herein are administered in a
therapeutically
effective amount. The amount of the compound actually administered will
typically be
determined by a physician, in the light of the relevant circumstances,
including the condition to
be treated, the chosen route of administration, the actual compound
administered, the age,
weight, and response of the individual patient, the severity of the patient's
symptoms, and the
like.
[00224] The pharmaceutical compositions provided herein can be administered
by a
variety of routes including oral, rectal, transdermal, subcutaneous,
intravenous, intramuscular,
and intranasal. Depending on the intended route of delivery, the compounds
provided herein are
preferably formulated as either injectable or oral compositions or as salves,
as lotions or as
patches all for transdermal administration.
[00225] The compositions for oral administration can take the form of bulk
liquid
solutions or suspensions, or bulk powders. More commonly, however, the
compositions are
presented in unit dosage forms to facilitate accurate dosing. The term "unit
dosage forms" refers
to physically discrete units suitable as unitary dosages for human subjects
and other mammals,
each unit containing a predetermined quantity of active material calculated to
produce the desired
therapeutic effect, in association with a suitable pharmaceutical excipient.
Typical unit dosage
forms include prefilled, premeasured ampules or syringes of the liquid
compositions or pills,
tablets, capsules or the like in the case of solid compositions. In such
compositions, the
compound is usually a minor component (from about 0.1 to about 50% by weight
or preferably
from about 1 to about 40% by weight) with the remainder being various vehicles
or carriers and
processing aids helpful for forming the desired dosing form.
[00226] Liquid forms suitable for oral administration may include a
suitable aqueous or
nonaqueous vehicle with buffers, suspending and dispensing agents, colorants,
flavors and the
like. Solid forms may include, for example, any of the following ingredients,
or compounds of a
similar nature: a binder such as microcrystalline cellulose, gum tragacanth or
gelatin; an
excipient such as starch or lactose, a disintegrating agent such as alginic
acid, Primogel, or corn
starch; a lubricant such as magnesium stearate; a glidant such as colloidal
silicon dioxide; a
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sweetening agent such as sucrose or saccharin; or a flavoring agent such as
peppermint, methyl
salicylate, or orange flavoring.
[00227] Injectable compositions are typically based upon injectable sterile
saline or
phosphate-buffered saline or other injectable carriers known in the art. As
before, the active
compound in such compositions is typically a minor component, often being from
about 0.05 to
10% by weight with the remainder being the injectable carrier and the like.
[00228] Transdermal compositions are typically formulated as a topical
ointment or cream
containing the active ingredient(s), generally in an amount ranging from about
0.01 to about 20%
by weight, preferably from about 0.1 to about 20% by weight, preferably from
about 0.1 to about
10% by weight, and more preferably from about 0.5 to about 15% by weight. When
formulated
as a ointment, the active ingredients will typically be combined with either a
paraffinic or a
water-miscible ointment base. Alternatively, the active ingredients may be
formulated in a cream
with, for example an oil-in-water cream base. Such transdermal formulations
are well-known in
the art and generally include additional ingredients to enhance the dermal
penetration of stability
of the active ingredients or the formulation. All such known transdermal
formulations and
ingredients are included within the scope provided herein.
[00229] The compounds provided herein can also be administered by a
transdermal device.
Accordingly, transdermal administration can be accomplished using a patch
either of the
reservoir or porous membrane type, or of a solid matrix variety.
[00230] The above-described components for orally administrable, injectable
or topically
administrable compositions are merely representative. Other materials as well
as processing
techniques and the like are set forth in Part 8 of Remington's Pharmaceutical
Sciences, 17th
edition, 1985, Mack Publishing Company, Easton, Pennsylvania, which is
incorporated herein by
reference.
[00231] The above-described components for orally administrable,
injectable, or topically
administrable compositions are merely representative. Other materials as well
as processing
techniques and the like are set forth in Part 8 of Remington's The Science and
Practice of
Pharmacy, 21st edition, 2005, Publisher: Lippincott Williams & Wilkins, which
is incorporated
herein by reference.
[00232] The compounds of this invention can also be administered in
sustained release
forms or from sustained release drug delivery systems. A description of
representative sustained
release materials can be found in Remington's Pharmaceutical Sciences.
[00233] The present invention also relates to the pharmaceutically
acceptable salts of
compounds of Formula I.
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[00234] The following formulation examples illustrate representative
pharmaceutical
compositions that may be prepared in accordance with this invention. The
present invention,
however, is not limited to the following pharmaceutical compositions.
Formulation 1 - Injection
[00235] A compound of the invention may be dissolved or suspended in a
buffered sterile
saline injectable aqueous medium to a concentration of approximately 5 mg/mL.
METHODS OF TREATMENT
[00236] Bolaamphiphilic vesicles (bolavesicles) may have certain advantages
over
conventional liposomes as potential vehicles for drug delivery. Bolavesicles
have thinner
membranes than comparable liposomal bilayer, and therefore possess bigger
inner volume and
hence higher encapsulation capacity than liposomes of the same diameter.
Moreover,
bolavesicles are more physically-stable than conventional liposomes, but can
be destabilized in a
triggered fashion (e.g., by hydrolysis of the headgroups using a specific
enzymatic reaction) thus
allowing controlled release of the encapsulated material at the site of action
(i.e., drug targeting).
[00237] Specific small interfering RNAs (siRNAs) designed to silence
different oncogenic
pathways can be used for cancer therapy. However, in the blood stream, non-
modified naked
non-modified siRNAs are unstable, thus having a short half-life in the blood
stream and
encounter difficulties in crossing biological membranes due to their negative
charge. Therefore,
siRNAs may not be used efficiently to silence genes. These obstacles can be
overcome by using
siRNAs complexed with bolaamphiphiles, consisting of two positively charged
head groups that
flank a hydrophobic chain. Bolaamphiphiles have relatively low toxicities,
long persistence in the
blood stream, and most importantly, can form poly-cationic micelles in aqueous
conditions thus,
becoming amenable to association with negatively charged siRNAs.
[00238] Experiments confirmed the formation of stable complexes the
bolaamphiphiles of
the present invention those can protect nucleic acids from their degradation
and thus effectively
deliver siRNAs into the cells causing the silencing of target genes.
GENERAL SYNTHETIC PROCEDURES
[00239] The compounds provided herein can be purchased or prepared from
readily
available starting materials using the following general methods and
procedures. See, e.g.,
Synthetic Schemes below. It will be appreciated that where typical or
preferred process
conditions (i.e., reaction temperatures, times, mole ratios of reactants,
solvents, pressures, etc.)
are given, other process conditions can also be used unless otherwise stated.
Optimum reaction
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conditions may vary with the particular reactants or solvent used, but such
conditions can be
determined by one skilled in the art by routine optimization procedures.
[00240] Additionally, as will be apparent to those skilled in the art,
conventional
protecting groups may be necessary to prevent certain functional groups from
undergoing
undesired reactions. The choice of a suitable protecting group for a
particular functional group as
well as suitable conditions for protection and deprotection are well known in
the art. For
example, numerous protecting groups, and their introduction and removal, are
described in T. W.
Greene and P. G. M. Wuts, Protecting Groups in Organic Synthesis, Second
Edition, Wiley,
New York, 1991, and references cited therein.
[00241] The compounds provided herein may be isolated and purified by known
standard
procedures. Such procedures include (but are not limited to)
recrystallization, column
chromatography or HPLC. The following schemes are presented with details as to
the
preparation of representative substituted biarylamides that have been listed
herein. The
compounds provided herein may be prepared from known or commercially available
starting
materials and reagents by one skilled in the art of organic synthesis.
[00242] The enantiomerically pure compounds provided herein may be prepared
according
to any techniques known to those of skill in the art. For instance, they may
be prepared by chiral
or asymmetric synthesis from a suitable optically pure precursor or obtained
from a racemate by
any conventional technique, for example, by chromatographic resolution using a
chiral column,
TLC or by the preparation of diastereoisomers, separation thereof and
regeneration of the desired
enantiomer. See, e.g., "Enantiomers, Racemates and Resolutions," by J.
Jacques, A. Collet, and
S.H. Wilen, (Wiley-Interscience, New York, 1981); S.H. Wilen, A. Collet, and
J. Jacques,
Tetrahedron, 2725 (1977); E.L. Eliel Stereochemistry of Carbon Compounds
(McGraw-Hill, NY,
1962); and S.H. Wilen Tables of Resolving Agents and Optical Resolutions 268
(E.L. Eliel ed.,
Univ. of Notre Dame Press, Notre Dame, IN, 1972, Stereochemistry of Organic
Compounds,
Ernest L. Eliel, Samuel H. Wilen and Lewis N. Manda (1994 John Wiley & Sons,
Inc.), and
Stereoselective Synthesis A Practical Approach, Mihaly Nagradi (1995 VCH
Publishers, Inc.,
NY, NY).
[00243] In certain embodiments, an enantiomerically pure compound of
formula (1) may
be obtained by reaction of the racemate with a suitable optically active acid
or base. Suitable
acids or bases include those described in Bighley et al., 1995, Salt Forms of
Drugs and
Adsorption, in Encyclopedia of Pharmaceutical Technology, vol. 13, Swarbrick &
Boylan, eds.,
Marcel Dekker, New York; ten Hoeve & H. Wynberg, 1985, Journal of Organic
Chemistry
50:4508-4514; Dale & Mosher, 1973, J. Am. Chem. Soc. 95:512; and CRC Handbook
of Optical
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Resolution via Diastereomeric Salt Formation, the contents of which are hereby
incorporated by
reference in their entireties.
[00244] Enantiomerically pure compounds can also be recovered either from
the
crystallized diastereomer or from the mother liquor, depending on the
solubility properties of the
particular acid resolving agent employed and the particular acid enantiomer
used. The identity
and optical purity of the particular compound so recovered can be determined
by polarimetry or
other analytical methods known in the art. The diasteroisomers can then be
separated, for
example, by chromatography or fractional crystallization, and the desired
enantiomer regenerated
by treatment with an appropriate base or acid. The other enantiomer may be
obtained from the
racemate in a similar manner or worked up from the liquors of the first
separation.
[00245] In certain embodiments, enantiomerically pure compound can be
separated from
racemic compound by chiral chromatography. Various chiral columns and eluents
for use in the
separation of the enantiomers are available and suitable conditions for the
separation can be
empirically determined by methods known to one of skill in the art. Exemplary
chiral columns
available for use in the separation of the enantiomers provided herein
include, but are not limited
to CHIRALCELO OB, CHIRALCELO OB-H, CHIRALCELO OD, CHIRALCELO OD-H,
CHIRALCELO OF, CHIRALCELO OG, CHIRALCELO OJ and CHIRALCELO OK.
[00246] ABBREVIATIONS
BBB, blood brain barrier
BCECs, brain capillary endothelial cells
CF, carboxyfluorescein
CHEMS, cholesteryl hemisuccinate
CHOL, cholesterol
Cryo-TEM, Cryo-transmission electron microscope
DAPI, 4',6- diamidino-2-phenylindole
DDS, drug delivery system
DLS, dynamic light scattering
DMPC, 1,2-dimyristoyl-sn-glycero-3-phosphocholine
DMPE, 1,2-dimyristoyl-sn-glycero-3-phosphoethanolamine
DMPG,1,2-dimyristoyl-sn-glycero-3-phospho-(1'-rac-glycerol)
EPR, electron paramagnetic resonance
FACS, fluorescence-activated cell sorting
FCR, fluorescence colorimetric response
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GUVs, giant unilamellar vesicles
HPLC, high performance liquid chromatography
IR, infrared
MNPs, Magnetic Nanoparticles
MRI, magnetic resonance imaging
NMR, nuclear magnetic resonance
NPs, nanoparticles
PBS, phosphate buffered saline
PC, phosphatidylcholine
PDA, polydiacetylene.
TMA-DPH, 1-(4 trimethylammoniumpheny1)-6-phenyl-1,3,5-hexatriene
Example 1
Bolaamphiphile synthesis
[00247] The boloamphiphles or bolaamphiphilic compounds of the invention
can be
synthesized following the procedures described previously (see below).
[00248] Briefly, the carboxylic group of methyl vernolate or vernolic acid
was interacted
with aliphatic diols to obtain bisvernolesters. Then the epoxy group of the
vernolate moiety,
located on C12 and C13 of the aliphatic chain of vernolic acid, was used to
introduce two ACh
headgroups on the two vicinal carbons obtained after the opening of the
oxirane ring. For GLH-
20 (Table 1), the ACh head group was attached to the vernolate skeleton
through the nitrogen
atom of the choline moiety. The bolaamphiphile was prepared in a two-stage
synthesis: First,
opening of the epoxy ring with a haloacetic acid and, second, quaternization
with the N,N-
dimethylamino ethyl acetate. For GLH-19 (Table 1) that contains an ACh head
group attached to
the vernolate skeleton through the acetyl group, the bolaamphiphile was
prepared in a three-stage
synthesis, including opening of the epoxy ring with glutaric acid, then
esterification of the free
carboxylic group with N,N-dimethyl amino ethanol and the final product was
obtained by
quaternization of the head group, using methyl iodide followed by exchange of
the iodide ion by
chloride using an ion exchange resin.
[00249] Each bolaamphiphile was characterized by mass spectrometry, NMR and
IR
spectroscopy. The purity of the two bolaamphiphiles was >97% as determined by
HPLC.
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[00250] Materials: Diphenyl ether, 1,2-hexadecanediol, oleic acid,
oleylamine, and
carboxyfluorescein (CF) were purchased from Sigma Aldrich (Rehovot, Israel).
Chloroform and
ethanol were purchased from Bio-Lab Ltd. Jerusalem, Israel. 1,2-dimyristoyl-sn-
glycero-3-
phospho-(1'-rac-glycerol) (DMPG), 1,2-dimyristoyl-sn-glycero-3-
phosphoethanolamine
(DMPE), 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC), cholesterol
(CHOL),
cholesteryl hemisuccinate (CHEMS) were purchased from Avanti Lipids
(Alabaster, AL, USA),
The diacetylenic monomer 10,12- tricosadiynoic acid was purchased from Alfa
Aesar (Karlsruhe,
Germany), and purified by dissolving the powder in chloroform, filtering the
resulting solution
through a 0.45 i.tm nylon filter (Whatman Inc., Clifton, NJ, USA), and
evaporation of the solvent.
1-(4 trimethylammoniumpheny1)-6-phenyl-1,3,5-hexatriene (TMA-DPH) was
purchased from
Molecular Probes Inc. (Eugene, OR, USA).
SYNTHESIS OF REPRESENTATIVE BOLAAMPHIPHILIC COMPOUNDS
[00251] The synthesis of bolaamphiphilic compounds of this invention can be
carried out
in accordance with the methods described previously (Chemistry and Physics of
Lipids 2008,
153, 85-97; Journal of Liposome Research 2010, 20, 147-59; W02002/055011;
W02003/047499; or W02010/128504) and using the appropriate reagents, starting
materials,
and purification methods known to those skilled in the art. Several
representative
bolaamphiphilic compounds of the invention, which are prepared in according
the methods
described herein or can be prepared following the methods described in the
literature or
following the methods known to those skilled in the art, are given in Table 1.
[00252] Table 1: Representative Bolaamphiphiles
Structure
0 0 e (
GLH-3
õ C r,Lir i
" 2 / 1 1 *** A (CH2)16
AØ0(CH2)1 01=:*polro-
%
Cr Cr
GLH-4
-
jcLe=
GLH-5 0N%
HO ¨ N,(CH2)24%N OH
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# Structure
GLH-6 a 0 0
HO - N N OH
H H
N
GLH-7 , 0 0 No(0-12)2,. 9 =/\"(0.110'n
%
HO ¨ N OH
H H
N NJci.= I ¨ V4
GLH-8*
HO - OH
H H
o 0
GLH-9
H N
GLH-10
(iL.= riNIfor ,N \
H H
'
HO C,.C.X.)-CH ,- CR ,- .H-C.000H
h) I 1
NH- C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
GLH-11 /
(CH 2)2 ; -
HO )..CO-Cli ,-(.:ki - CH -Ct-,yooi
\ l' I I
NH- C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
iv
:-.= = e-,= = , \\,.
GLH-12 a I-2N.'-j1.A-m2.1\ / 9 yH
---- , ,
cH3_(cH2)4-CH - CH- CH2- CH=CH-(CH2)7- COOCH3
on 0,
V/
GLH-13 a ('FirC0-0-0-12-C-112-N- CH2-CO-HN OH
I I
CH3-(CH2)4-CH - CH- CH2- CH=CH-(CH2)7- COOCH3
CH3 cilõ
X" C"3 CH3 V , X
GLH-13 a V/
cH3-00-0-CH2-CH.--N-CH2-CO-HN 0-CO-CH2-N-C1-12-CH2-0-CO-CH3
I I
CH3-(CH2)4-CH - CH- CH2- CH=CH-(CH2)2- COOCH3
GLH-14 cr
=Ny04,...."K,,,stro 0
a e lt, A
I.J 0.,(CH2)2 0 an
õ.,
.., Cie.
GLH-15 '\y-..." N 'The.' 0
0 1 % 6 (c H2), 0 o qat
.,,. ,,, N _e,,o.,xe
-i1 ,', ; ,
H H
0
(CH2 0 ) 2 alr-1
GLH-16 o ,..
,1,,, N
H H 6
GLH-17
H H
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# Structure
GLH-18
NO."......0y"...."ir 0 (b
0 0 \Nie
/N.%
0 (.9 0,...Y1 re
0
= µ--m2,1
-12 HO = N =NJA0.".=*4'0%
H H
,,õe".õ,oy,...,,,,yot
0.11õ",0õ,,,yo.....=,..00
GLH-19 /Ns 0 0 =it#,=
.(c1-12)10 c9
(V Ho ¨ o so OH
GLH-20
o = o oo ,,, e % 0
,k N.,,,,, r=zy. A)11 ,rZY
0 Hi 1)-\= 1-,.,:z=-,_,L,..-,4.,,2
il ....4
/ C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4I3
GLH-21 NH
i
COOH
(CH 2)2
I 111.?
NH i
\ C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH3
CI-I = CH
li HO 0-CO-CH 2- N -CH 2-0-1 2-0-C.0-C1 3
I I
,-- C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
GLH-22 NH
I CH - Cfi 3
r2)12
0 HO 0-CO-CH r N -CH 2-0-1 2-0-CO-CH 3
Nu......... II I I
C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
GLH-23 0 rrs/N/NeN le--10
0 0
HOK-\
"A1 O"= Olk /
ne0H
NH2 OH 0 NH2
0 Cit , .43 Ci:
GLH-24
oI. "=o " 4' %
ri 0
GLH-25
(LT HO ..(CH2)11)
0 '0 OH UV
CT-I, Cal) ci-
-i'
1? li)-CO-CH2- N -CH2-CH2-0-CO-CH3
......-- C-(CH 2)7 -CH=CH-CH 2-CH-(CH 2)3-CH3
GLH-26 y
I-3 ci- if ..
- ..5 ' .3 u
(CH 2),
I go p-CO-CH2- N -CH2-Cii2-0-CO-CH3
0 ii
----...._
C-(CH 2)7 -CH=CH-CH 2-CH-(CH 2)5-CH 3
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Structure
ca-13
\
H? rCO-CH2- N -CH.2-0-12-0-CO-C113
GLH-27 C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
cll, cH3
(CH 2)8 I-
I ID HQ 0-CO-CH2- N -CH?-042-0-00-013
oil I I
C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
cil,
H? rCO-CH2- N -C112-CHTO-CO-Cas
c4c14 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
GLH-28
CH; CHs
(VI 2)6
I H9 1)-CO-CH2- N -CH2-012-0-CO-CH3
II
c4c14 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-C113
Cli^6
i+
HO 0-CO-CH2- N -CH2-CH2-0-CG-CH3
I
GLH-29 C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
(H2)10 CH3 C143
cr
-CO-CH2-N -CIL2-012-0-CO-CM,
0 CH., CH.
IT
C-(CH 2)11 -N CH2-CH2-0420-013
GLH-30 (H6
H CH7
1 r
0 U 3
- C-(CH 2)11 -N -CH2-CH2-0-CO-CHI
o CHI CH3 BII r-
= C-(CH 2)11 -N -CH2 -CH2 -0-CO-CH3
GLH-302/ ')
16
CH 3 CH.3
0I t + Cr
C-(CH 2)11 -0-CO-CH-1
CH3 Cli3 cr.
HO 0-CO-CH2- N -CH2-CH2-0-CO-CH3
GLH-31 I
C-(CH 2)7 -CH=CH-CH 2-CH-CH-(CH 2)4-CH 3
CH,
1E12)12 C17...1,.:
GL H-3 2
HO 0 OH
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# Structure
no
GLH-33 o e %
o 0
o(c1-12)10 II^ %,, o
(A 6
or o cr .
GLH-34
0 e k 0 0 Ire\ to,
0
Cr Cr
GLH-35
HO 0 "0 OH
CI Cr
GLH-36
o#(01-12)2
GLH-37
cHolo
GLH-38
\õ....
\
,.
o o
GLH-39a H,Ns\l() 0 (C1-12)100.1t(C1-1242141"OH
Cr , C-
GLH-40
o o
0-
,
GLH-41 Y/N\ 0
0-
+
_
0 0 0
n H 2, AOH
H2ise\61(7) 1-12),4. 0A , ¨12
GLH-42 a 0 (0 C.)
0 H 0 0 0
H2N4C4.14(') 0(C1-12)120).1(CH2)1j2sLOH
GLH-43 a a
GLH-44 )4o-
s\o'';',',01400# tir'f,,!Ni'"""\-c'n"
0 0 44 0
:WI
0 t ,, 0 0
stM0"...44 .04 00%,...W."... 0 tr.W.=======14 0 "..."=================== 01/0.
4.,... 0 pe
GLH-45 ecl 0 n * .6
GLH-46
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# Structure
0 ,, 4, 0
C) , , 0 o
)404.8.z.Alto,~%,N,NA0=~%,,olewl=,01-.ir,1-%.,011
00 0 0 .* 0
2:: e
o * $ o
2cia
2. , ..? o Br E-3
0
GLH-47 0 4 :0
Br G
0 Br G
GLH-48
$ !,
0
,.., $ 0
)...1
GLH-49 a
(le 0
GLH-50a ,s'µc).' \'0.14'0W==''==''=== .pr.,................../.-%oi_,
1.,= A'oH
Ao"..0,61%,,Ao
GLH-51 a Cl. e 0
-
)11
GLH-52a H,0H0 o
OH'04,)11OH
0....1),IH _
0
HO 0-(CH2CH20),-H
0
GLH-53 a
¨
H3C 0¨ C H3
0 0
GLH-54 a _
H3C 0¨H
, -
HO ilf..,21._
i,C;....==.0 L'
GLH-55. 0 0 / - \OH -c=0 .
0
P
,MoNo!s,4,,,A10==========================Orr=OWN"/"0"."==========ONIN0CV H
0 0
1-10y 8 0
GLH-56 0 . 0
' , , e '-=
= v ,...(CH2)44.040
N /
mgGLH-
r.,..-*/!.. (,,, ., 0*-- ¨ 0H 1"),,,e
N=kk,...,Y,),<.:¨ '
a ¨ an intermediate
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Example 2
Formation of bolaamphiphiles/siRNA complex
[00253] Vesicles are prepared by dissolving 10 mg/ml bolaamphiphile in
chloroform
together with 2.1 mg/ml cholesteryl hemisuccinate and 1.6 mg/ml cholesterol.
The organic
solvent is evaporated under nitrogen and then is kept under vacuum overnight.
The thin film that
was formed is hydrated by RNAs-free water to a concentration of 10 mg/ml of
the
bolaamphiphile and the suspension which was formed after mixing is sonicated
to form 100 nm
vesicles at a concentration of 10 mg/ml of bolaamphiphile. These vesicles are
used to form the
complex with the siRNA duplex as described below.
[00254] The siRNA duplex is mixed with pre-prepared vesicles (concentration
of the
siRNA ranges between 100 nM and 10 pM, and the concentration of the
bolaamphiphile ranges
between 200 p g/ml and 1 mg/ml). The vesicles that are prepared at a
concentration of the
bolaamphiphile of 10 mg/ml are diluted before mixing to a concentration that
ranges between
200 p g/ml and 1 mg/ml. The mixture is allowed to stand on ice for 30 min-24
hours.
Example 3
Transfection of cell cultures with bolaamphiphile/siRNA complexes
[00255] All transfections are performed using bolaamphiphiles of the
invention. The
concentration of siRNA (200 nM- 10p M) to bolaamphiphiles (200 jig/ml-1 mg/ml)
can be 10X-
1000X. The transfection is done with either eGFP siRNA that can silence the
eGFP gene, which
is expressed by the transfected cells. Alternatively, to determine if the
siRNA penetrated into the
cells, the inventors used siRNA-fluorescent probe conjugate. The fluorescent
probe could be
FITC or Alexa Flour such as AF-555. Prior to each transfection, the cell media
is swapped with
growth medium without serum and the prepared siRNA/ bolaamphiphile vesicles
complexes (as
described in Example 2) are diluted to the final concentration of 1X. The
cells are incubated for
5-12 hours followed by changing the media to the growth medium.
Example 4
Fluorescent Light Microscopy
[00256] To assess the silencing efficiency, or the number of the
fluorescent cells in the
case where non-fluorescent cells were transfected by siRNA-fluorescent probe
conjugate, cells
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are imaged 72 hours after the transfection or 5-24 hours after the
transfection, respectively, with
a fluorescent microscope.
Example 5
Imaging isolated organs after administration of siRNA/bolaamphiphile vesicle
complex to mice
[00257] siRNA/bolaamphiphile vesicles complex, prepared by mixing siRNA-
AF555
conjugate with bolaamphiphilic vesicles (as described in example 2 above), was
injected
intravenously to mice via the tail vein. The injected dose ranged between 20
mg/kg to 40 mg/kg
of the bolaamphiphile. Mice were sacrificed 30 min and 2 hours after the
injection and organs
were washed by saline and fluorescence imaging of the isolated organs was
performed.
[00258] Based on the results, it can be determined that bolaamphiles have
the potential to
be used as the carriers for siRNA delivery. Bola/siRNA vesicle complexes
significantly increase
the stability of siRNAs, provided resistence against nucleases activity and
provide excellent
intracellular uptake followed by a specific gene silencing. Moreover,
depending on application,
the extent of protection of siRNA can be altered by simply changing the
carrier.
[00259] From the foregoing description, various modifications and changes
in the
compositions and methods provided herein will occur to those skilled in the
art. All such
modifications coming within the scope of the appended claims are intended to
be included
therein.
[00260] All publications, including but not limited to patents and patent
applications, cited
in this specification are herein incorporated by reference as if each
individual publication were
specifically and individually indicated to be incorporated by reference herein
as though fully set
forth.
[00261] At least some of the chemical names of compounds of the invention
as given and
set forth in this application, may have been generated on an automated basis
by use of a
commercially available chemical naming software program, and have not been
independently
verified. Representative programs performing this function include the
Lexichem naming tool
sold by Open Eye Software, Inc. and the Autonom Software tool sold by MDL,
Inc. In the
instance where the indicated chemical name and the depicted structure differ,
the depicted
structure will control.
[00262] Chemical structures shown herein were prepared using ISIS /DRAW.
Any open
valency appearing on a carbon, oxygen or nitrogen atom in the structures
herein indicates the
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presence of a hydrogen atom. Where a chiral center exists in a structure but
no specific
stereochemistry is shown for the chiral center, both enantiomers associated
with the chiral
structure are encompassed by the structure.
[00263] References
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cyclodextrin polymer-based nanoparticle: from concept to clinic. Mol Pharm 6,
659-668
(2009).
3. Davis, M.E. et al. Evidence of RNAi in humans from systemically
administered siRNA
via targeted nanoparticles. Nature 464, 1067-1070 (2010).
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interference and
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14. Elbashir, S.M., Martinez, J., Patkaniowska, A., Lendeckel, W. & Tuschl,
T. Functional
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