U.S. patent application number 16/353047 was filed with the patent office on 2020-02-13 for cyclic nitro compounds, pharmaceutical compositions thereof and uses thereof.
The applicant listed for this patent is EpicentRx, Inc., Northrop Grumman Innovation Systems, Inc.. Invention is credited to Mark D. Bednarski, Louis Cannizzo, Susan Knox, Shoucheng Ning, Stephen Velarde, Robert Wardle, Kirstin Warner.
Application Number | 20200046682 16/353047 |
Document ID | / |
Family ID | 37758325 |
Filed Date | 2020-02-13 |
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United States Patent
Application |
20200046682 |
Kind Code |
A1 |
Bednarski; Mark D. ; et
al. |
February 13, 2020 |
CYCLIC NITRO COMPOUNDS, PHARMACEUTICAL COMPOSITIONS THEREOF AND
USES THEREOF
Abstract
The present invention provides cyclic nitro compound,
pharmaceutical compositions of cyclic nitro compounds and methods
of using cyclic nitro compounds and/or pharmaceutical compositions
thereof to treat or prevent diseases or disorders characterized by
abnormal cell proliferation, such as cancer, inflammation,
cardiovascular disease and autoimmune disease.
Inventors: |
Bednarski; Mark D.; (Los
Altos, CA) ; Knox; Susan; (Stanford, CA) ;
Cannizzo; Louis; (Ogden, UT) ; Warner; Kirstin;
(Ogden, UT) ; Wardle; Robert; (Logan, UT) ;
Velarde; Stephen; (North Ogden, UT) ; Ning;
Shoucheng; (Palo Alto, CA) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
EpicentRx, Inc.
Northrop Grumman Innovation Systems, Inc. |
La Jolla
Plymouth |
CA
MN |
US
US |
|
|
Family ID: |
37758325 |
Appl. No.: |
16/353047 |
Filed: |
March 14, 2019 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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15669403 |
Aug 4, 2017 |
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16353047 |
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14965062 |
Dec 10, 2015 |
10149832 |
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15669403 |
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14584177 |
Dec 29, 2014 |
9226915 |
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14965062 |
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13655618 |
Oct 19, 2012 |
8927527 |
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14584177 |
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12397651 |
Mar 4, 2009 |
8299053 |
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13655618 |
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11502810 |
Aug 11, 2006 |
7507842 |
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12397651 |
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60707851 |
Aug 12, 2005 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
A61N 5/10 20130101; A61P
29/00 20180101; A61K 31/397 20130101; A61P 35/02 20180101; A61P
43/00 20180101; A61P 37/06 20180101; A61P 9/00 20180101; C07D
205/04 20130101; A61P 35/00 20180101; A61K 9/0019 20130101 |
International
Class: |
A61K 31/397 20060101
A61K031/397; A61K 9/00 20060101 A61K009/00; A61N 5/10 20060101
A61N005/10; C07D 205/04 20060101 C07D205/04 |
Claims
1. A compound of structural Formula (I): ##STR00006## or salts,
solvates or hydrates thereof wherein: R.sup.1, R.sup.2, R.sup.3 and
R.sup.4 are each independently, hydrogen, alkyl, substituted alkyl,
aryl, substituted aryl, arylalkyl, substituted arylalkyl,
heteroaryl, substituted heteroaryl, heteroalkyl, substituted
heteroalkyl, heteroarylalkyl, substituted heteroarylalkyl, halo,
hydroxy or nitro; each R.sup.5 and R.sup.6 are each independently,
hydrogen, alkyl, substituted alkyl, aryl, substituted aryl,
arylalkyl, substituted arylalkyl, heteroaryl, substituted
heteroaryl, heteroalkyl, substituted heteroalkyl, heteroarylalkyl,
substituted heteroarylalkyl, halo, hydroxy or nitro; o is 0, 1, 2,
3 or 4; R.sup.7 is substituted alkyl, substituted arylalkyl,
substituted heteroalkyl, substituted heteroaryl, substituted
heteroarylalkyl, substituted acyl, substituted alkoxycarbonyl,
substituted phosphonyl or substituted sulfonyl; provided that at
least one of R.sup.1, R.sup.2, R.sup.3, R.sup.4, R.sup.5 and
R.sup.6 are nitro.
2.-16. (canceled)
17. A method for treating or preventing cancer in a patient
comprising administering to a patient in need of such treatment or
prevention a therapeutically effective amount of the compound of
claim 1.
18. (canceled)
19. A method for treating tumor cells with a reduced intracellular
environment in a patient, comprising administering to the patient
in need of treatment a therapeutically effective amount of the
compound of claim 1.
20. A method for treating or preventing solid tumors in a patient,
comprising administering to the patient in need of treatment or
prevention a therapeutically effective amount of the compound of
claim 1.
21. A method for treating or preventing leukemias and lymphomas in
a patient, comprising administering to the patient in need of
treatment or prevention a therapeutically effective amount of the
compound of claim 1.
22. A method for treating or preventing inflammation in a patient,
comprising administering to the patient in need of treatment or
prevention a therapeutically effective amount of the compound of
claim 1.
23. A method for treating or preventing autoimmune disease in a
patient, comprising administering to the patient in need of
treatment or prevention a therapeutically effective amount of the
compound of claim 1.
24. A method for treating or preventing cardiovascular disease in a
patient, comprising administering to the patient in need of
treatment or prevention a therapeutically effective amount of the
compound of claim 1.
Description
CROSS-REFERENCE TO RELATED APPLICATION
[0001] The present application claims priority to U.S. Provisional
Application No. 60/707,851 filed Aug. 12, 2005 which is hereby
incorporated by reference in its entirety.
FIELD OF THE INVENTION
[0002] The present invention relates generally to pharmaceutical
compositions of cyclic nitro compounds and methods of using cyclic
nitro compounds and pharmaceutical compositions thereof to treat or
prevent diseases characterized by abnormal cell proliferation such
as cancer.
BACKGROUND OF THE INVENTION
[0003] Abnormal cell proliferation is a characteristic symptom of
cancer. Further, abnormal cell proliferation has been implicated in
numerous other diseases (e.g., cardiovascular diseases,
inflammatory diseases such as rheumatoid arthritis, diabetic
retinopathy, etc.). Although many methods for treating or
preventing aberrant cell proliferation have been developed, a
significant problem with most existing therapies is selectively
distinguishing between normal and abnormal cell proliferation.
[0004] Radiotherapy is one promising approach to selectively
targeting abnormal cell proliferation. A number of different
radiosensitizers have been described in the art and include thiols,
nitroimidazoles and metal texaphyrin compounds (See e.g., Rosenthal
et al., Clin. Cancer. Res., 1999, 739). Significant problems with
existing radiosensitization approaches are (1) the formation of
toxic byproducts derived from the radiosensitizers, which has
limited their usefulness in cancer therapy; and (2) achieving
sufficiently high density of free radicals to be efficacious under
dose limiting toxicity.
[0005] Another popular approach to selectively targeting abnormal
cell proliferation, is treatment with bioreductive compounds, which
are selectively activated in a reducing environment. Since many
cancers typically contain regions of low oxygen tension (i.e.,
hypoxia), compounds with low redox potentials (i.e., bioreductive
compounds) may be selectively activated in the reducing environment
of tumor cells without external activation.
[0006] Accordingly, new compounds are required to fully explore
treating or preventing abnormal cell proliferation. These new
compounds may have radiotherapeutic activity or bioreductive
activity. Such compounds may be effective in treating or preventing
various diseases associated with abnormal cell proliferation such
as cancer without forming toxic byproducts.
SUMMARY OF THE INVENTION
[0007] The present invention satisfies this and other needs by
providing cyclic nitro compounds, pharmaceutical compositions of
cyclic nitro compounds and methods of using cyclic nitro compounds
or pharmaceutical compositions thereof to treat or prevent diseases
associated with abnormal cell proliferation.
[0008] In a first aspect, a compound of structural Formula (I):
##STR00001##
[0009] or salts, solvates or hydrates thereof is provided
wherein:
[0010] R.sup.1, R.sup.2, R.sup.3 and R.sup.4 are each
independently, hydrogen, alkyl, substituted alkyl, aryl,
substituted aryl, arylalkyl, substituted arylalkyl, heteroalkyl,
substituted heteroalkyl, heteroaryl, substituted heteroaryl,
heteroarylalkyl, substituted heteroarylalkyl, halo, hydroxy or
nitro;
[0011] each R.sup.5 and R.sup.6 are each independently, hydrogen,
alkyl, substituted alkyl, aryl, substituted aryl, arylalkyl,
substituted arylalkyl, heteroalkyl, substituted heteroalkyl,
heteroaryl, substituted heteroaryl, heteroarylalkyl, substituted
heteroarylalkyl, halo, hydroxy or nitro; [0012] o is 0, 1, 2, 3 or
4;
[0013] R.sup.7 is substituted alkyl, substituted arylalkyl,
substituted heteroalkyl, substituted heteroaryl, substituted
heteroarylalkyl, substituted acyl, substituted alkoxycarbonyl,
substituted phosphonyl or substituted sulfonyl;
[0014] provided that at least one of R.sup.1, R.sup.2, R.sup.3,
R.sup.4, R.sup.5 and R.sup.6 are nitro.
[0015] In a second aspect, methods for treating or preventing
diseases or disorders characterized by abnormal cell proliferation
are provided. The methods generally involve administering to a
patient in need of such treatment or prevention a therapeutically
effective amount of a cyclic nitro compound or a pharmaceutically
acceptable salt, hydrate, solvate or N-oxide thereof.
[0016] In a third aspect, pharmaceutical compositions of cyclic
nitro compounds are provided. The pharmaceutical compositions
generally comprise one or more cyclic nitro compounds,
pharmaceutically acceptable salts, hydrates, solvates or N-oxides
thereof and a pharmaceutically acceptable vehicle. The choice of
vehicle will depend upon, among other factors, the desired mode of
administration.
[0017] In a fourth aspect, pharmaceutical compositions for treating
or preventing diseases or disorders characterized by abnormal cell
proliferation are provided. The methods generally involve
administering to a patient in need of such treatment or prevention
a therapeutically effective amount of a pharmaceutical composition
comprising a cyclic nitro compound or a pharmaceutically acceptable
salt, hydrate, solvate or N-oxide thereof and a pharmaceutically
acceptable vehicle.
BRIEF DESCRIPTION OF THE DRAWINGS
[0018] FIG. 1 illustrates the dose and cell line dependency of ROS
production in tumor cells in the presence of ABDNAZ;
[0019] FIG. 2 illustrates ROS production in HT29 tumor cells in the
presence of irradiated ABDNAZ;
[0020] FIG. 3 illustrates ROS production in SCC VII tumor cells in
the presence of irradiated ABDNAZ;
[0021] FIG. 4 illustrates inhibition of proliferation of bcl-2 and
vector transfected HL60 cells by ABDNAZ;
[0022] FIG. 5 illustrates induction of apoptosis of bcl-2 and
vector transfected HL60 cells by ABDNAZ;
[0023] FIG. 6 illustrates the apoptosis and cell cycle profile of
HL60 neo cells after exposure to ABDNAZ;
[0024] FIG. 7 illustrates the apoptosis and cell cycle profile of
HL60 bcl-2 cells after exposure to ABDNAZ; and
[0025] FIG. 8 illustrates the inhibition of bcl-2 expression in HL
60 cells.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
Definitions
[0026] "Alkyl" by itself or as part of another substituent refers
to a saturated or unsaturated, branched, straight-chain or cyclic
monovalent hydrocarbon radical derived by the removal of one
hydrogen atom from a single carbon atom of a parent alkane, alkene
or alkyne. Typical alkyl groups include, but are not limited to,
methyl; ethyls such as ethanyl, ethenyl, ethynyl; propyls such as
propan-1-yl, propan-2-yl, cyclopropan-1-yl, prop-1-en-1-yl,
prop-1-en-2-yl, prop-2-en-1-yl (allyl), cycloprop-1-en-1-yl;
cycloprop-2-en-1-yl, prop-1-yn-1-yl, prop-2-yn-1-yl, etc.; butyls
such as butan-1-yl, butan-2-yl, 2-methyl-propan-1-yl,
2-methyl-propan-2-yl, cyclobutan-1-yl, but-1-en-1-yl,
but-1-en-2-yl, 2-methyl-prop-1-en-1-yl, but-2-en-1-yl,
but-2-en-2-yl, buta-1,3-dien-1-yl, buta-1,3-dien-2-yl,
cyclobut-1-en-1-yl, cyclobut-1-en-3-yl, cyclobuta-1,3-dien-1-yl,
but-1-yn-1-yl, but-1-yn-3-yl, but-3-yn-1-yl, etc.; and the
like.
[0027] The term "alkyl" is specifically intended to include groups
having any degree or level of saturation, i.e., groups having
exclusively single carbon-carbon bonds, groups having one or more
double carbon-carbon bonds, groups having one or more triple
carbon-carbon bonds and groups having mixtures of single, double
and triple carbon-carbon bonds. Where a specific level of
saturation is intended, the expressions "alkanyl," "alkenyl," and
"alkynyl" are used. In some embodiments, an alkyl group comprises
from 1 to 20 carbon atoms. In other embodiments, an alkyl group
comprises 1 to 10 carbon atoms. In still other embodiments an alkyl
group comprises from 1 to 6 carbon atoms.
[0028] "Alkanyl" by itself or as part of another substituent refers
to a saturated branched, straight-chain or cyclic alkyl radical
derived by the removal of one hydrogen atom from a single carbon
atom of a parent alkane. Typical alkanyl groups include, but are
not limited to, methanyl; ethanyl; propanyls such as propan-1-yl,
propan-2-yl (isopropyl), cyclopropan-1-yl, etc.; butanyls such as
butan-1-yl, butan-2-yl (sec-butyl), 2-methyl-propan-1-yl(isobutyl),
2-methyl-propan-2-yl (t-butyl), cyclobutan-1-yl, etc.; and the
like.
[0029] "Alkenyl" by itself or as part of another substituent refers
to an unsaturated branched, straight-chain or cyclic alkyl radical
having at least one carbon-carbon double bond derived by the
removal of one hydrogen atom from a single carbon atom of a parent
alkene. The group may be in either the cis or trans conformation
about the double bond(s). Typical alkenyl groups include, but are
not limited to, ethenyl; propenyls such as prop-1-en-1-yl,
prop-1-en-2-yl, prop-2-en-1-yl (allyl), prop-2-en-2-yl,
cycloprop-1-en-1-yl; cycloprop-2-en-1-yl; butenyls such as
but-1-en-1-yl, but-1-en-2-yl, 2-methyl-prop-1-en-1-yl,
but-2-en-1-yl, but-2-en-1-yl, but-2-en-2-yl, buta-1,3-dien-1-yl,
buta-1,3-dien-2-yl, cyclobut-1-en-1-yl, cyclobut-1-en-3-yl,
cyclobuta-1,3-dien-1-yl, etc.; and the like.
[0030] "Alkynyl" by itself or as part of another substituent refers
to an unsaturated branched, straight-chain or cyclic alkyl radical
having at least one carbon-carbon triple bond derived by the
removal of one hydrogen atom from a single carbon atom of a parent
alkyne. Typical alkynyl groups include, but are not limited to,
ethynyl; propynyls such as prop-1-yn-1-yl, prop-2-yn-1-yl, etc.;
butynyls such as but-1-yn-1-yl, but-1-yn-3-yl, but-3-yn-1-yl, etc.;
and the like.
[0031] "Acyl" by itself or as part of another substituent refers to
a radical --C(O)R.sup.30, where R.sup.30 is hydrogen, alkyl,
cycloalkyl, cycloheteroalkyl, aryl, arylalkyl, heteroalkyl,
heteroaryl, heteroarylalkyl as defined herein. Representative
examples include, but are not limited to formyl, acetyl,
cyclohexylcarbonyl, cyclohexylmethylcarbonyl, benzoyl,
benzylcarbonyl and the like.
[0032] "Alkoxycarbonyl" by itself or as part of another substituent
refers to a radical --C(O)OR.sup.31 where R.sup.31 represents an
alkyl or cycloalkyl group as defined herein. Representative
examples include, but are not limited to, methoxycarbonyl,
ethoxycarbonyl, propoxycarbonyl, butoxycarbonyl,
cyclohexyloxycarbonyl and the like.
[0033] "Aryl" by itself or as part of another substituent refers to
a monovalent aromatic hydrocarbon group derived by the removal of
one hydrogen atom from a single carbon atom of a parent aromatic
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,
trinaphthalene and the like. In some embodiments, an aryl group
comprises from 6 to 20 carbon atoms. In other embodiments, an aryl
group comprises from between 6 to 12 carbon atoms.
[0034] "Arylalkyl" by itself or as part of another substituent
refers to an acyclic alkyl group in which one of the hydrogen atoms
bonded to a carbon atom, typically a terminal or sp.sup.3 carbon
atom, is replaced with an aryl group. Typical arylalkyl groups
include, but are not limited to, benzyl, 2-phenylethan-1-yl,
2-phenylethen-1-yl, naphthylmethyl, 2-naphthylethan-1-yl,
2-naphthylethen-1-yl, naphthobenzyl, 2-naphthophenylethan-1-yl and
the like. Where specific alkyl moieties are intended, the
nomenclature arylalkanyl, arylalkenyl and/or arylalkynyl is used.
In some embodiments, an arylalkyl group is (C.sub.6-C.sub.30)
arylalkyl, e.g., the alkanyl, alkenyl or alkynyl moiety of the
arylalkyl group is (C.sub.1-C.sub.10) and the aryl moiety is
(C.sub.6-C.sub.20). In other embodiments, an arylalkyl group is
(C.sub.6-C.sub.20) arylalkyl, e.g., the alkanyl, alkenyl or alkynyl
moiety of the arylalkyl group is (C.sub.1-C.sub.9) and the aryl
moiety is (C.sub.6-C.sub.12).
[0035] "Heteroalkyl, Heteroalkanyl, Heteroalkenyl and
Heteroalkynyl" by themselves or as part of another substituent
refer to alkyl, alkanyl, alkenyl and alkynyl groups, respectively,
in which one or more of the carbon atoms (and any associated
hydrogen atoms) are independently replaced with the same or
different heteroatomic groups. Typical heteroatomic groups which
can be included in these groups include, but are not limited to,
--O--, --S--, --O--O--, --S--S--, --O--S--, --NR.sup.34R.sup.35,
--, .dbd.N--N.dbd., --N.dbd.N--, --N.dbd.N--NR.sup.36R.sup.37,
--PR.sup.38--, --P(O).sub.2--, --POR.sup.39--, --O--P(O).sub.2--,
--SO--, --SO.sub.2--, SnR.sup.40R.sup.41-- and the like, where
R.sup.34, R.sup.35, R.sup.36, R.sup.37, R.sup.38, R.sup.39,
R.sup.40 and R.sup.41 are independently hydrogen, alkyl,
substituted alkyl, aryl, substituted aryl, arylalkyl, substituted
arylalkyl, cycloalkyl, substituted cycloalkyl, cycloheteroalkyl,
substituted cycloheteroalkyl, heteroalkyl, substituted heteroalkyl,
heteroaryl, substituted heteroaryl, heteroarylalkyl or substituted
heteroarylalkyl.
[0036] "Heteroaryl" by itself or as part of another substituent
refers to a monovalent heteroaromatic radical derived by the
removal of one hydrogen atom from a single atom of a parent
heteroaromatic ring system. Typical heteroaryl groups include, but
are not limited to, groups derived from acridine, arsindole,
carbazole, .beta.-carboline, chromane, chromene, cinnoline, furan,
imidazole, indazole, indole, indoline, indolizine, isobenzofuran,
isochromene, isoindole, isoindoline, isoquinoline, isothiazole,
isoxazole, naphthyridine, oxadiazole, oxazole, perimidine,
phenanthridine, phenanthroline, phenazine, phthalazine, pteridine,
purine, pyran, pyrazine, pyrazole, pyridazine, pyridine,
pyrimidine, pyrrole, pyrrolizine, quinazoline, quinoline,
quinolizine, quinoxaline, tetrazole, thiadiazole, thiazole,
thiophene, triazole, xanthene, and the like. In some embodiments,
the heteroaryl group is between 5-20 membered heteroaryl. In other
embodiments, the heteroaryl group is between 5-10 membered
heteroaryl. In some embodiments, heteroaryl groups include those
derived from thiophene, pyrrole, benzothiophene, benzofuran,
indole, pyridine, quinoline, imidazole, oxazole and pyrazine.
[0037] "Heteroarylalkyl" by itself or as part of another
substituent refers to an acyclic alkyl group in which one of the
hydrogen atoms bonded to a carbon atom, typically a terminal or
sp.sup.3 carbon atom, is replaced with a heteroaryl group. Where
specific alkyl moieties are intended, the nomenclature
heteroarylalkanyl, heteroarylalkenyl and/or heterorylalkynyl is
used. In some embodiments, the heteroarylalkyl group is a 6-30
membered heteroarylalkyl, e.g., the alkanyl, alkenyl or alkynyl
moiety of the heteroarylalkyl is 1-10 membered and the heteroaryl
moiety is a 5-20-membered heteroaryl. In other embodiments, the
heteroarylalkyl group is a 6-20 membered heteroarylalkyl, e.g., the
alkanyl, alkenyl or alkynyl moiety of the heteroarylalkyl is 1-8
membered and the heteroaryl moiety is a 5-12-membered
heteroaryl.
[0038] "Parent Aromatic Ring System" by itself or as part of
another substituent, refers to an unsaturated cyclic or polycyclic
ring system having a conjugated .pi. electron system. Specifically
included within the definition of "parent aromatic ring system" are
fused ring systems in which one or more of the rings are aromatic
and one or more of the rings are saturated or unsaturated, such as,
for example, fluorene, indane, indene, phenalene, etc. Typical
parent aromatic ring systems include, but are not limited to,
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, trinaphthalene and the like.
[0039] "Parent Heteroaromatic Ring System" by itself or as part of
another substituent, refers to a parent aromatic ring system in
which one or more carbon atoms (and any associated hydrogen atoms)
are independently replaced with the same or different heteroatom.
Typical heteroatoms to replace the carbon atoms include, but are
not limited to, N, P, O, S, Si, etc. Specifically included within
the definition of "parent heteroaromatic ring systems" are fused
ring systems in which one or more of the rings are aromatic and one
or more of the rings are saturated or unsaturated, such as, for
example, arsindole, benzodioxan, benzofuran, chromane, chromene,
indole, indoline, xanthene, etc. Typical parent heteroaromatic ring
systems include, but are not limited to, arsindole, carbazole,
.beta.-carboline, chromane, chromene, cinnoline, furan, imidazole,
indazole, indole, indoline, indolizine, isobenzofuran, isochromene,
isoindole, isoindoline, isoquinoline, isothiazole, isoxazole,
naphthyridine, oxadiazole, oxazole, perimidine, phenanthridine,
phenanthroline, phenazine, phthalazine, pteridine, purine, pyran,
pyrazine, pyrazole, pyridazine, pyridine, pyrimidine, pyrrole,
pyrrolizine, quinazoline, quinoline, quinolizine, quinoxaline,
tetrazole, thiadiazole, thiazole, thiophene, triazole, xanthene,
and the like.
[0040] "Pharmaceutically acceptable salt" refers to a salt of a
O-nitro compound, which is pharmaceutically acceptable and
possesses the desired pharmacological activity of the parent
compound. Such salts: (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, t-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 is replaced by an ammonium ion, a metal ion,
e.g., a alkali metal ion (e.g., sodium or potassium), an alkaline
earth ion (e.g., calcium or magnesium), or an aluminum ion; or
coordinates with an organic base such as ethanolamine,
diethanolamine, triethanolamine, N-methylglucamine, morpholine,
piperidine, dimethylamine, diethylamine and the like. Also included
are salts of amino acids such as arginates and the like, and salts
of organic acids like glucurmic or galactunoric acids and the
like.
[0041] "Pharmaceutically acceptable vehicle" refers to a diluent,
adjuvant, excipient or carrier with which a 0-nitro compound is
administered.
[0042] "Patient" includes humans and other mammals.
[0043] "Phosphonyl" by itself or as part of another substituent
refers to a radical --P(O)(OR.sup.32).sub.2, where each R.sup.32 is
independently hydrogen, alkyl, cycloalkyl, cycloheteroalkyl, aryl,
arylalkyl, heteroalkyl, heteroaryl, heteroarylalkyl as defined
herein.
[0044] "Preventing" or "prevention" refers to a reduction in risk
of acquiring a disease or disorder (i.e., causing at least one of
the clinical symptoms of the disease not to develop in a patient
that may be exposed to or predisposed to the disease but does not
yet experience or display symptoms of the disease).
[0045] "Substituted" refers to a group in which one or more
hydrogen atoms are independently replaced with the same or
different substituent(s). Typical substituents include, but are not
limited to, -M, --R.sup.60, --O.sup.-, .dbd.O, --OR.sup.60,
--SR.sup.60, S.sup.-, .dbd.S, --NR.sup.60R.sup.61, .dbd.NR.sup.60,
--CF.sub.3, --CN, --OCN, --SCN, --NO, --NO.sub.2, --ONO.sub.2,
.dbd.N.sub.2, --N.sub.3, --S(O).sub.2O.sup.-, --S(O).sub.2OH,
--S(O).sub.2R.sup.60, --OS(O.sub.2)O.sup.-, --OS(O).sub.2R.sup.60,
--P(O)(O.sup.-).sub.2, --P(O)(OR.sup.60)(O.sup.-),
--OP(O)(OR.sup.60)(OR.sup.61), --C(O)R.sup.60, --C(S)R.sup.60,
--C(O)OR.sup.60, --C(O)NR.sup.60, R.sup.61, --C(O)O.sup.-,
--C(S)OR.sup.60, --NR.sup.62C(O)NR.sup.60R.sup.61,
--NR.sup.62C(S)NR.sup.60R.sup.61,
--NR.sup.62C(NR.sup.63)NR.sup.60R.sup.61 and
--C(NR.sup.62)NR.sup.60R.sup.61 where M is independently a and
halogen; R.sup.60, R.sup.61, R.sup.62 and R.sup.63 are
independently hydrogen, alkyl, substituted alkyl, alkoxy,
substituted alkoxy, cycloalkyl, substituted cycloalkyl,
cycloheteroalkyl, substituted cycloheteroalkyl, aryl, substituted
aryl, heteroaryl or substituted heteroaryl, or optionally R.sup.60
and R.sup.61 together with the nitrogen atom to which they are
bonded form a cycloheteroalkyl or substituted cycloheteroalkyl
ring; and R.sup.64 and R.sup.65 are independently hydrogen, alkyl,
substituted alkyl, aryl, cycloalkyl, substituted cycloalkyl,
cycloheteroalkyl, substituted cycloheteroalkyl, aryl, substituted
aryl, heteroaryl or substituted heteroaryl, or optionally R.sup.64
and R.sup.65 together with the nitrogen atom to which they are
bonded form a cycloheteroalkyl or substituted cycloheteroalkyl
ring. In some embodiments, substituents include -M, --R.sup.60,
.dbd.O, --OR.sup.60, --SR.sup.60, --S.sup.-, .dbd.S,
--NR.sup.60R.sup.61, .dbd.NR.sup.60, --CF.sub.3, --CN, --OCN,
--SCN, --NO, --NO.sub.2, --ONO.sub.2, .dbd.N.sub.2, --N.sub.3,
--S(O).sub.2R.sup.60, --OS(O.sub.2)O.sup.-, --OS(O).sub.2R.sup.60,
--P(O)(O.sup.-).sub.2, --P(O)(OR.sup.60)(O.sup.-),
--OP(O)(OR.sup.60)(OR.sup.61), --C(O)R.sup.60, --C(S)R.sup.60,
--C(O)OR.sup.60, --C(O)NR.sup.60R.sup.61, --C(O)O.sup.-,
--NR.sup.62C(O)NR.sup.60R.sup.61, where R.sup.60, R.sup.61 and
R.sup.62 are as defined above. In other embodiments, substituents
include -M, --R.sup.60, .dbd.O, --OR.sup.60, --SR.sup.60,
--NR.sup.60R.sup.61, --CF.sub.3, --CN, --NO.sub.2, --ONO.sub.2,
--S(O).sub.2R.sup.60, --P(O)(OR.sup.60)(O.sup.-),
--OP(O)(OR.sup.60)(OR.sup.61), --C(O)R.sup.60, --C(O)OR.sup.60,
--C(O)NR.sup.60R.sup.61 and --C(O)O.sup.- where R.sup.60, R.sup.61
and R.sup.62 are as defined above. In still other embodiments,
substituents include -M, --R.sup.60, .dbd.O, --OR.sup.60,
--SR.sup.60, --NR.sup.60R.sup.61, --CF.sub.3, --CN, --NO.sub.2,
--ONO.sub.2, --S(O).sub.2R.sup.60, --OP(O)(OR.sup.60)(OR.sup.61),
--C(O)R.sup.60, --C(O)OR.sup.60 and --C(O)O.sup.-, where R.sup.60,
R.sup.61 and R.sup.62 are as defined above.
[0046] "Sulfonyl" by itself or as part of another substituent
refers to a radical --S(O).sub.2R.sub.33, where each R.sup.33 is
independently hydrogen, alkyl, cycloalkyl, cycloheteroalkyl, aryl,
arylalkyl, heteroalkyl, heteroaryl, heteroarylalkyl as defined
herein.
[0047] "Treating" or "treatment" of any disease or disorder refers,
in some embodiments, to ameliorating the disease or disorder (i.e.,
arresting or reducing the development of the disease or at least
one of the clinical symptoms thereof). In other embodiments
"treating" or "treatment" refers to ameliorating at least one
physical parameter, which may not be discernible by the patient. In
yet other embodiments, "treating" or "treatment" refers to
inhibiting the disease or disorder, either physically, (e.g.,
stabilization or eradication of a discernible symptom),
physiologically, (e.g., stabilization or eradication of a physical
parameter) or both. In still other embodiments, "treating" or
"treatment" refers to delaying the onset of the disease or
disorder.
[0048] "Therapeutically effective amount" means the amount of a
compound that, when administered to a patient for treating or
preventing a disease, is sufficient to effect such treatment or
prevention of the disease. The "therapeutically effective amount"
will vary depending on the compound, the disease and its severity
and the age, weight, etc., of the patient to be treated.
[0049] Reference will now be made in detail to embodiments of the
invention. While the invention will be described in conjunction
with these embodiments, it will be understood that it is not
intended to limit the invention to those preferred embodiments. To
the contrary, it is intended to cover alternatives, modifications,
and equivalents as may be included within the spirit and scope of
the invention as defined by the appended claims.
[0050] "Pharmaceutically acceptable salt" refers to a salt of a
cyclic nitro compound, which is pharmaceutically acceptable and
possesses the desired pharmacological activity of the parent
compound. Such salts: (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, t-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 is replaced by an ammonium ion, a metal ion,
e.g., a alkali metal ion (e.g., sodium or potassium), an alkaline
earth ion (e.g., calcium or magnesium), or an aluminum ion; or
coordinates with an organic base such as ethanolamine,
diethanolamine, triethanolamine, N-methylglucamine, morpholine,
piperidine, dimethylamine, diethylamine and the like. Also included
are salts of amino acids such as arginates and the like, and salts
of organic acids like glucurmic or galactunoric acids and the
like.
[0051] "Pharmaceutically acceptable vehicle" refers to a diluent,
adjuvant, excipient or carrier with which a cyclic nitro compound
is administered.
[0052] "Patient" includes humans and other mammals.
[0053] "Preventing" or "prevention" refers to a reduction in risk
of acquiring a disease or disorder (i.e., causing at least one of
the clinical symptoms of the disease not to develop in a patient
that may be exposed to or predisposed to the disease but does not
yet experience or display symptoms of the disease).
[0054] "Treating" or "treatment" of any disease or disorder refers,
in one embodiment, to ameliorating the disease or disorder (i.e.,
arresting or reducing the development of the disease or 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 patient. In
yet another embodiment, "treating" or "treatment" refers to
inhibiting the disease or disorder, either physically, (e.g.,
stabilization or eradication of a discernible symptom),
physiologically, (e.g., stabilization or eradication of a physical
parameter) or both. In yet another embodiment, "treating" or
"treatment" refers to delaying the onset of the disease or
disorder.
[0055] "Therapeutically effective amount" means the amount of a
compound that, when administered to a patient for treating or
preventing a disease, is sufficient to effect such treatment or
prevention of the disease. The "therapeutically effective amount"
will vary depending on the compound, the disease and its severity
and the age, weight, etc., of the patient to be treated.
[0056] Reference will now be made in detail to embodiments of the
invention. While the invention will be described in conjunction
with these embodiments, it will be understood that it is not
intended to limit the invention to those preferred embodiments. To
the contrary, it is intended to cover alternatives, modifications,
and equivalents as may be included within the spirit and scope of
the invention as defined by the appended claims.
Cyclic Nitro Compounds and Their Use to Treat or Prevent Abnormal
Cell Proliferation
[0057] The present invention provides cyclic nitro compounds,
pharmaceutical compositions of cyclic nitro compounds and methods
of using cyclic nitro compounds or pharmaceutical compositions
thereof to treat or prevent diseases associated with abnormal cell
proliferation.
[0058] The methods generally involve administering to a patient in
need of such treatment or prevention a therapeutically effective
amount of a cyclic nitro compound or a pharmaceutically acceptable
salt, hydrate, solvate or N-oxide thereof. In one embodiment, the
cyclic nitro compound is intracellularly activated by the reducing
environment of a tumor cell. In another embodiments, the patient is
irradiated to activate the cyclic nitro compound. Without wishing
to be bound by theory, irradiation or reduction of cyclic nitro
compounds may lead to formation of free radicals that subsequently
prevent cell replication and kill cells, presumably by interfering
with DNA replication and/or reacting with cell membranes. However,
other mechanisms, presently unknown, may account for the efficacy
of cyclic nitro compounds in treating or preventing abnormal cell
proliferation.
[0059] Accordingly, in some embodiments, the cyclic nitro compounds
of the present invention may be activated by both intracellular
reduction and external irradiation. In these embodiments, a
synergistic or additive effect may be observed.
[0060] Cyclic nitro compounds are generally organic compounds
substituted with one or more nitro groups (i.e., nitro compounds)
but also include nitrate salts (e.g., ammonium dinitride, aluminum
trinitride, etc.). Typically, cyclic nitro compounds have a high
enthalapy of formation (i.e., decomposition of cyclic nitro
compounds releases a high amount of energy). In some embodiments,
cyclic nitro compounds have an enthalapy of formation that varies
between about 5 kcal/mole and about 150 kcal/mole, more preferably,
between about 10 kcal/mole and about 110 kcal/mole. The enthalapy
of formation of nitro compounds may be readily calculated by
methods known to the skilled artisan. Accordingly, cyclic nitro
compounds include those nitro compounds that decompose with
explosive force upon activation. Such compounds may be readily
identified by those of skill in the art by calculation of the
enthalapy of formation.
[0061] Cyclic nitro compounds may also be reduced at low reduction
potentials. cyclic voltametry demonstrates that electron transfer
to cyclic nitro compounds occurs between about -0.1 volts and about
-1.0 volts using standard electrodes (e.g., mercury or carbon
cathode and platinum anode) and electrolyte solutions.
[0062] In some embodiments, cyclic nitro compounds contain a high
density of nitro groups (i.e., the nitro groups represent a
significant fraction of the overall mass of the compound). In other
embodiments, cyclic nitro compounds contain two nitro groups. In
still other embodiments, cyclic nitro compounds contain three nitro
groups. In still other embodiments, cyclic nitro compounds contain
three or more nitro groups. In still other embodiments, a cyclic
nitro compound contains six nitro groups.
[0063] In some embodiments, the cyclic nitro compound is a
nitrocarbon which has a ratio of nitro groups to carbon atoms of
1:1. In other embodiments, the cyclic nitro compound is a
nitrocarbon which has a ratio of nitro groups to carbon atoms of
1:2.
[0064] In some embodiments, a compound of structural Formula
(I):
##STR00002##
[0065] or salts, solvates or hydrates thereof is provided
wherein:
[0066] R.sup.1, R.sup.2, R.sup.3 and R.sup.4 are each
independently, hydrogen, alkyl, substituted alkyl, aryl,
substituted aryl, arylalkyl, substituted arylalkyl, heteroalkyl,
substituted heteroalkyl, heteroaryl, substituted heteroaryl,
heteroarylalkyl, substituted heteroarylalkyl, halo, hydroxy or
nitro;
[0067] each R.sup.5 and R.sup.6 are each independently, hydrogen,
alkyl, substituted alkyl, aryl, substituted aryl, arylalkyl,
substituted arylalkyl, heteroalkyl, substituted heteroalkyl,
heteroaryl, substituted heteroaryl, heteroarylalkyl, substituted
heteroarylalkyl, halo, hydroxy or nitro; [0068] o is 0, 1, 2, 3 or
4;
[0069] R.sup.7 is substituted alkyl, substituted arylalkyl,
substituted heteroalkyl, substituted heteroaryl, substituted
heteroarylalkyl, substituted acyl, substituted alkoxycarbonyl,
substituted phosphonyl or substituted sulfonyl;
[0070] provided that at least one of R.sup.1, R.sup.2, R.sup.3,
R.sup.4, R.sup.5 and R.sup.6 are nitro.
[0071] In some embodiments, at least two of R.sup.1, R.sup.2,
R.sup.3, R.sup.4, R.sup.5 or R.sup.6 are nitro. In other
embodiments, R.sup.1, R.sup.2, R.sup.3 and R.sup.4 are each
independently, hydrogen, alkyl or nitro and each R.sup.5 and
R.sup.6 are each independently, hydrogen, alkyl or nitro. In still
other embodiments, R.sup.7 is substituted alkyl, substituted acyl,
substituted alkoxycarbonyl, substituted phosphonyl or substituted
sulfonyl. In still other embodiments, R.sup.7 is alkyl, acyl,
alkoxycarbonyl, phosphonyl or sulfonyl substituted with one or more
halogen, --CF.sub.3 or --OS(O).sub.2R.sup.8 wherein R.sup.8 is
alkyl, substituted alkyl, aryl or substituted aryl. In still other
embodiments, R.sup.3 and R.sup.4 are nitro.
[0072] In some embodiments, R.sup.1, R.sup.2, R.sup.3 and R.sup.4
are each independently, hydrogen, alkyl or nitro, each R.sup.5 and
R.sup.6 are each independently, hydrogen, alkyl or nitro and
R.sup.7 is substituted alkyl, substituted acyl, substituted
alkoxycarbonyl, substituted phosphonyl or substituted sulfonyl. In
other embodiments, R.sup.1, R.sup.2, R.sup.3 and R.sup.4 are each
independently, hydrogen, alkyl or nitro, each R.sup.5 and R.sup.6
are each independently, hydrogen, alkyl or nitro and R.sup.7 is
alkyl, acyl, alkoxycarbonyl, phosphonyl or sulfonyl substituted
with one or more halogen, --CF.sub.3 or --OS(O).sub.2R.sup.8
wherein R.sup.8 is alkyl, substituted alkyl, aryl or substituted
aryl. In still other embodiments, R.sup.1 and R.sup.2 are each
independently, hydrogen, alkyl or nitro, R.sup.3 and R.sup.4 are
nitro, each R.sup.5 and R.sup.6 are each independently, hydrogen,
alkyl or nitro, R.sup.7 is substituted alkyl, substituted acyl,
substituted alkoxycarbonyl, substituted phosphonyl or substituted
sulfonyl. In still other embodiments, R.sup.1 and R.sup.2 are each
independently, hydrogen, alkyl or nitro, R.sup.3 and R.sup.4 are
nitro, each R.sup.5 and R.sup.6 are each independently, hydrogen,
alkyl or nitro, R.sup.7 is alkyl, acyl, alkoxycarbonyl, phosphonyl
or sulfonyl substituted with one or more halogen, --CF.sub.3 or
--OS(O).sub.2R.sup.8 wherein R.sup.8 is alkyl, substituted alkyl,
aryl or substituted aryl. In some of any of the above embodiments,
o is 1.
[0073] In some embodiments, R.sup.1 and R.sup.2 are each
independently, hydrogen, alkyl or nitro, R.sup.3 and R.sup.4 are
nitro, R.sup.5 and R.sup.6 are each independently, hydrogen, alkyl
or nitro, R.sup.7 is alkyl or acyl substituted with one or more
halogen or --CF.sub.3 and o is 1. In other embodiments, R.sup.1 and
R.sup.2 are hydrogen, R.sup.3 and R.sup.4 are nitro, R.sup.5 and
R.sup.6 are hydrogen, R.sup.7 is alkyl or acyl substituted with one
or more halogen or --CF.sub.3 and o is 1.
[0074] In some embodiments, the cyclic nitro compound has the
structure:
##STR00003##
[0075] wherein each X is independently --F, --Cl, --Br, --I or
--OS(O).sub.2R.sup.8 where R.sup.8 is methyl, CF.sub.3, phenyl or
tolyl and each p is independently 1, 2, 3, or 4.
[0076] In other embodiments, the cyclic nitro compound has the
structure:
##STR00004##
[0077] wherein each X is independently --F, --Cl, --Br, --I or
--OS(O).sub.2R.sup.8 where R.sup.8 is methyl, CF.sub.3, phenyl or
tolyl. In some specific embodiments, the cyclic nitro compound has
the structure:
##STR00005##
[0078] commonly referred to as ABDNAZ.
[0079] Cyclic nitro compounds may exist in several tautomeric forms
and mixtures thereof. Cyclic nitro compounds may also include
isotopically labeled compounds where one or more atoms have an
atomic mass different from the atomic mass conventionally found in
nature. Examples of isotopes that may be incorporated into cyclic
nitro compounds include, but are not limited to, .sup.2H, .sup.3H,
.sup.13C, .sup.14C, 15N, .sup.18O and .sup.17O. Cyclic nitro
compounds may exist in unsolvated forms as well as solvated forms,
including hydrated forms or a N-oxides. In general, hydrated and
solvated forms are within the scope of the present invention.
Certain cyclic nitro compounds may exist in multiple crystalline or
amorphous forms. In general, all physical forms are equivalent for
the uses contemplated by the present invention and are intended to
be within the scope of the present invention.
[0080] Cyclic nitro compounds may be activated by intracellular
reduction. In some embodiments, cyclic nitro compounds are
activated by intracellular reduction in hypoxic tumor cells,
secondary to elevated glutathione levels (high GSH:GSSG (i.e.,
glutathione to glutathione disulfide ratios)) and possibly high
levels of other antioxidant enzymes in many tumor cells and/or a
median tumor cell pO.sub.2 of less than about 10 mm Hg.
[0081] Cyclic nitro compounds may also be activated by application
of external energy. Methods useful for decomposing cyclic nitro
compounds include, but are not limited to, irradiation (e.g., with
x-rays, visible light, infrared irradiation) ultrasound (e.g.
focused ultrasound), electrochemical reduction, heating,
co-administration of free radical initiators (e.g., thiols), etc.
In some embodiments, a cyclic nitro compound is activated by photon
irradiation of the patient. In some embodiments, the patient's
tumor is irradiated using a linear accelerator at a dose rate of
about 100 cGy/min. The patient may also be treated with electron
beam therapy, interoperative radiation therapy, stereostatic
radiosurgery and high or low dose brachytherapy.
[0082] In some situations the entire patient may be irradiated. In
some embodiments, a portion of the patient is irradiated so that
only cyclic nitro compound localized in the irradiated portion
(e.g., tumor region) of the patient is activated. Preferably, the
portion of the patient which is irradiated is the site of abnormal
cell proliferation.
[0083] Cyclic nitro compounds may be obtained via conventional
synthetic methods described in the art or are commercially
available, e.g., from ATK Thiokol, Salt Lake City, Utah Starting
materials useful for preparing cyclic nitro compounds and
intermediates thereof are commercially available or can be prepared
by well-known synthetic methods. Other methods for synthesis of the
cyclic nitro compounds described herein and/or starting materials
are either described in the art or will be readily apparent to the
skilled artisan.
[0084] In accordance with the invention, a cyclic nitro compound or
a pharmaceutical composition thereof is administered to a patient,
preferably a human, suffering from a disease characterized by
abnormal cell proliferation. The cyclic nitro compound and
pharmaceutical compositions thereof may be used to treat or prevent
diseases characterized by abnormal cell proliferation.
[0085] Diseases characterized by abnormal cell proliferation
include, but are not limited to, cancer (e.g., any vascularized
tumor, preferably, a solid tumor, including but not limited to,
carcinomas of the lung, breast, ovary, stomach, pancreas, larynx,
esophagus, testes, liver, parotid, bilary tract, colon, rectum,
cervix, uterus, endometrium, kidney, bladder, prostrate, thyroid,
squamous cell carcinomas, adenocarcinomas, small cell carcinomas,
melanomas, gliomas, neuroblastomas, sarcomas (e.g., angiosarcomas,
chondrosarcomas), diabetes, cardiovascular diseases (e.g.,
arteriosclerosis), inflammatory diseases (e.g., arthritis, diabetic
retinopathy, rheumatoid arthritis, neovascular glaucoma and
psoriasis) and autoimmune diseases.
[0086] In other embodiments, cyclic nitro compounds may be used for
in-vitro sterilization. Biological solutions may be treated with
cyclic nitro compounds, which are toxic to pathogenic bacteria,
viruses and cells. This process can also be catalyzed by the
application of external energy such as light and heat.
[0087] Further, in certain embodiments, a cyclic nitro compound
and/or pharmaceutical compositions thereof are administered to a
patient, preferably a human, as a preventative measure against
various diseases or disorders characterized by abnormal cell
proliferation. Thus, cyclic nitro compounds and/or pharmaceutical
compositions thereof may be administered as a preventative measure
to a patient having a predisposition for a disease characterized by
abnormal cell proliferation. Accordingly, cyclic nitro compounds
and/or pharmaceutical compositions thereof may be used for the
prevention of one disease or disorder and concurrently treating
another (e.g., preventing arthritis while treating cancer).
[0088] Therapeutic/Prophylactic Administration
[0089] Cyclic nitro compounds and/or pharmaceutical compositions
thereof may be advantageously used in human medicine. As previously
described in Section 5.2 supra, cyclic nitro compounds and/or
pharmaceutical compositions thereof are useful for the treatment or
prevention of various diseases or disorders such as those listed
above.
[0090] When used to treat or prevent the above disease or
disorders, cyclic nitro compounds and/or pharmaceutical
compositions thereof may be administered or applied singly, or in
combination with other agents. Cyclic nitro compounds and/or
pharmaceutical compositions thereof may also be administered or
applied singly, or in combination with other pharmaceutically
active agents (e.g., other anti-cancer agents, other arthritis
agents, etc.), including other cyclic nitro compounds and/or
pharmaceutical compositions thereof.
[0091] The current invention provides methods of treatment and
prophylaxis by administration to a patient of a therapeutically
effective amount of a cyclic nitro compound and/or pharmaceutical
composition thereof. The patient is preferably, a mammal and most
preferably, is a human.
[0092] Cyclic nitro compounds and/or pharmaceutical compositions
thereof may be administered orally. Cyclic nitro compounds and/or
pharmaceutical compositions thereof may also be administered by any
other convenient route, for example, by infusion or bolus
injection, by absorption through epithelial or mucocutaneous
linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.).
Administration can be systemic or local. Various delivery systems
are known, (e.g., encapsulation in liposomes, microparticles,
microcapsules, capsules, etc.) that can be used to administer a
cyclic nitro compound and/or pharmaceutical composition thereof.
Methods of administration include, but are not limited to,
intradermal, intramuscular, intraperitoneal, intravenous,
subcutaneous, intranasal, epidural, oral, sublingual, intranasal,
intracerebral, intravaginal, transdermal, rectally, by inhalation,
or topically, particularly to the ears, nose, eyes or skin. The
mode of administration is left to the discretion of the
practitioner, and will depend in-part upon the site of the medical
condition. In most instances, administration will result in the
release of cyclic nitro compounds and/or pharmaceutical
compositions thereof into the bloodstream.
[0093] In specific embodiments, it may be desirable to administer
one or more cyclic nitro compounds and/or pharmaceutical
compositions thereof locally to the area in need of treatment. This
may be achieved, for example, and not by way of limitation, by
local infusion during surgery, topical application, e.g., in
conjunction with a wound dressing after surgery, by injection, by
means of a catheter, by means of a suppository, or by means of an
implant, said implant being of a porous, non-porous, or gelatinous
material, including membranes, such as sialastic membranes or
fibers. In one embodiment, administration can be by direct
injection at the site (or former site) of the disease or
disorder.
[0094] In certain embodiments, it may be desirable to introduce one
or more cyclic nitro compounds and/or pharmaceutical compositions
thereof into the central nervous system by any suitable route,
including intraventricular, intrathecal and epidural injection.
Intraventricular injection may be facilitated by an
intraventricular catheter, for example, attached to a reservoir,
such as an Ommaya reservoir.
[0095] Cyclic nitro compounds and/or pharmaceutical compositions
thereof may also be administered directly to the lung by
inhalation. For administration by inhalation, cyclic nitro
compounds and/or pharmaceutical composition thereof may be
conveniently delivered to the lung by a number of different
devices. For example, a Metered Dose Inhaler ("MDI"), which
utilizes canisters that contain a suitable low boiling propellant,
(e.g., dichlorodifluoromethane, trichlorofluoromethane,
dichlorotetrafluoroethane, carbon dioxide or any other suitable
gas) may be used to deliver cyclic nitro compounds and/or
pharmaceutical compositions thereof directly to the lung.
[0096] Alternatively, a Dry Powder Inhaler ("DPI") device may be
used to administer a cyclic nitro compound and/or pharmaceutical
composition thereof to the lung. DPI devices typically use a
mechanism such as a burst of gas to create a cloud of dry powder
inside a container, which may then be inhaled by the patient and
are well known in the art. A popular variation is the multiple dose
DPI ("MDDPI") system, which allows for the delivery of more than
one therapeutic dose. MDDPI devices are commercially available from
a number of pharmaceutical companies e.g., Schering Plough,
Madison, N.J.). For example, capsules and cartridges of gelatin for
use in an inhaler or insufflator may be formulated containing a
powder mix of a cyclic nitro compound and/or pharmaceutical
composition thereof and a suitable powder base such as lactose or
starch for these systems.
[0097] Another type of device that may be used to deliver a cyclic
nitro compound and/or pharmaceutical composition thereof to the
lung is a liquid spray device supplied, for example, by Aradigm
Corporation, Hayward, Calif. Liquid spray systems use extremely
small nozzle holes to aerosolize liquid drug formulations that may
then be directly inhaled into the lung.
[0098] In some embodiments, a nebulizer is used to deliver a cyclic
nitro compound and/or pharmaceutical composition thereof to the
lung. Nebulizers create aerosols from liquid drug formulations by
using, for example, ultrasonic energy to form fine particles that
may be readily inhaled (see e.g., Verschoyle et al., British J.
Cancer, 1999, 80, Suppl. 2, 96). Examples of nebulizers include
devices supplied by Sheffield Pharmaceuticals, St. Louis, Mo.
(Armer et al., U.S. Pat. No. 5,954,047; van der Linden et al., U.S.
Pat. No. 5,950,619; van der Linden et al., U.S. Pat. No. 5,970,974)
and Batelle Pulmonary Therapeutics, Columbus, Ohio).
[0099] In other embodiments, an electrohydrodynamic ("EHD") aerosol
device is used to deliver a cyclic nitro compound and/or
pharmaceutical composition thereof to the lung of a patient. EHD
aerosol devices use electrical energy to aerosolize liquid drug
solutions or suspensions (see e.g., Noakes et al., U.S. Pat. No.
4,765,539). The electrochemical properties of the formulation may
be important parameters to optimize when delivering a cyclic nitro
compound and/or pharmaceutical composition thereof to the lung with
an EHD aerosol device and such optimization is routinely performed
by one of skill in the art. EHD aerosol devices may more
efficiently deliver drugs to the lung than existing pulmonary
delivery technologies.
[0100] In some embodiments, a cyclic nitro compound and/or
pharmaceutical compositions thereof can be delivered in a vesicle,
in particular a liposome (e.g., Langer, 1990, Science,
249:1527-1533; Treat et al., in "Liposomes in the Therapy of
Infectious Disease and Cancer," Lopez-Berestein and Fidler (eds.),
Liss, New York, pp. 353-365 (1989)).
[0101] In some embodiments, a cyclic nitro compound and/or
pharmaceutical compositions thereof can be delivered via sustained
release systems, preferably oral sustained release systems. In
other embodiments, a pump may be used (e.g., Langer, supra, Sefton,
1987, CRC Crit. Ref Biomed. Eng. 14:201; Saudek et al., 1989, N.
Engl. J Med. 321:574).
[0102] In some embodiments, polymeric materials can be used (e.g.,
"Medical Applications of Controlled Release," Langer and Wise
(eds.), CRC Press, Boca Raton, Fla. (1974); "Controlled Drug
Bioavailability," Drug Product Design and Performance, Smolen and
Ball (eds.), Wiley, New York (1984); Ranger et al., 1983, J
Macromol. Sci. Rev. Macromol Chem. 23:61; Levy et al., 1985,
Science 228: 190; During et al., 1989, Ann. Neurol. 25:351; Howard
et al., 1989, J. Neurosurg. 71:105).
[0103] In other embodiments, polymeric materials are used for oral
sustained release delivery. Polymers include, but are not limited
to, sodium carboxymethylcellulose, hydroxypropylcellulose,
hydroxypropylmethylcellulose and hydroxyethylcellulose (most
preferred, hydroxypropyl methylcellulose). Other cellulose ethers
have been described (Alderman, Int. J. Pharm. Tech. &Prod. Mfr.
1984, 5(3) 1-9). Factors affecting drug release are well known to
the skilled artisan and have been described in the art (Bamba et
al., Int. J. Pharm. 1979, 2, 307).
[0104] In other embodiments, enteric-coated preparations can be
used for oral sustained release administration. Coating materials
include polymers with a pH-dependent solubility (i.e.,
pH-controlled release), polymers with a slow or pH-dependent rate
of swelling, dissolution or erosion (i.e., time-controlled
release), polymers that are degraded by enzymes (i.e.,
enzyme-controlled release) and polymers that form firm layers that
are destroyed by an increase in pressure (i.e., pressure-controlled
release).
[0105] In other embodiments, osmotic delivery systems are used for
oral sustained release administration (Verma et al., Drug Dev. Ind.
Pharm., 2000, 26:695-708). In some embodiments, OROS.TM. osmotic
devices are used for oral sustained release delivery devices
(Theeuwes et al., U.S. Pat. No. 3,845,770; Theeuwes et al., U.S.
Pat. No. 3,916,899).
[0106] In yet other embodiments, a controlled-release system can be
placed in proximity of the target of the cyclic nitro compound
and/or pharmaceutical composition, thus requiring only a fraction
of the systemic dose (e.g., Goodson, in "Medical Applications of
Controlled Release," supra, vol. 2, pp. 115-138 (1984)). Other
controlled-release systems previously may also be used (Langer,
1990, Science 249:1527-1533).
[0107] Pharmaceutical Compositions
[0108] The present pharmaceutical compositions typically contain a
therapeutically effective amount of one or cyclic nitro compounds,
preferably, in purified form, together with a suitable amount of a
pharmaceutically acceptable vehicle, so as to provide the form for
proper administration to a patient. When administered to a patient,
the cyclic nitro compound and pharmaceutically acceptable vehicles
are preferably sterile. Water is a preferred vehicle when the
cyclic nitro compound is administered intravenously. Saline
solutions and aqueous dextrose and glycerol solutions can also be
employed as liquid vehicles, particularly for injectable solutions.
Suitable pharmaceutical vehicles also include excipients such as
starch, glucose, lactose, sucrose, gelatin, malt, rice, flour,
chalk, silica gel, sodium stearate, glycerol monostearate, talc,
sodium chloride, dried skim milk, glycerol, propylene, glycol,
water, ethanol and the like. The present pharmaceutical
compositions, if desired, can also contain minor amounts of wetting
or emulsifying agents, or pH buffering agents. In addition,
auxiliary, stabilizing, thickening, lubricating and coloring agents
may be used.
[0109] Pharmaceutical compositions comprising a cyclic nitro
compound may be manufactured by means of conventional mixing,
dissolving, granulating, dragee-making, levigating, emulsifying,
encapsulating, entrapping or lyophilizing processes. Pharmaceutical
compositions may be formulated in conventional manner using one or
more physiologically acceptable carriers, diluents, excipients or
auxiliaries, which facilitate processing of compounds into
preparations which can be used pharmaceutically. Proper formulation
is dependent upon the route of administration chosen.
[0110] The present pharmaceutical compositions can take the form of
solutions, suspensions, emulsion, tablets, pills, pellets,
capsules, capsules containing liquids, powders, sustained-release
formulations, suppositories, emulsions, aerosols, sprays,
suspensions, or any other form suitable for use. In one embodiment,
the pharmaceutically acceptable vehicle is a capsule (e.g.,
Grosswald et al., U.S. Pat. No. 5,698,155). A general discussion of
the preparation of pharmaceutical compositions may be found in
Remington, "The Science and Practice of Pharmacy," 19.sup.th
Edition.
[0111] For topical administration a cyclic nitro compound may be
formulated as solutions, gels, ointments, creams, suspensions, etc.
as is well-known in the art.
[0112] Systemic formulations include those designed for
administration by injection, e.g., subcutaneous, intravenous,
intramuscular, intrathecal or intraperitoneal injection, as well as
those designed for transdermal, transmucosal, oral or pulmonary
administration. Systemic formulations may be made in combination
with a further active agent that improves mucociliary clearance of
airway mucus or reduces mucous viscosity. These active agents
include, but are not limited to, sodium channel blockers,
antibiotics, N-acetyl cysteine, homocysteine and phospholipids.
[0113] In a preferred embodiment, cyclic nitro compounds are
formulated in accordance with routine procedures as a
pharmaceutical composition adapted for intravenous administration
to human beings. Typically, cyclic nitro compounds are solutions in
sterile isotonic aqueous buffer for intravenous administration. For
injection, cyclic nitro compounds may be formulated in aqueous
solutions, preferably in physiologically compatible buffers such as
Hanks' solution, Ringer's solution, or physiological saline buffer.
The solution may contain formulatory agents such as suspending,
stabilizing and/or dispersing agents. When necessary, the
pharmaceutical compositions may also include a solubilizing agent.
Pharmaceutical compositions for intravenous administration may
optionally include a local anesthetic such as lignocaine to ease
pain at the site of the injection. Generally, the ingredients are
supplied either separately or mixed together in unit dosage form,
for example, as a lyophilized powder or water free concentrate in a
hermetically sealed container such as an ampoule or sachette
indicating the quantity of active agent. When the cyclic nitro
compounds are administered by infusion, it can be dispensed, for
example, with an infusion bottle containing sterile pharmaceutical
grade water or saline. When the cyclic nitro compound is
administered by injection, an ampoule of sterile water for
injection or saline can be provided so that the ingredients may be
mixed prior to administration.
[0114] For transmucosal administration, penetrants appropriate to
the barrier to be permeated are used in the formulation. Such
penetrants are generally known in the art.
[0115] Pharmaceutical compositions for oral delivery may be in the
form of tablets, lozenges, aqueous or oily suspensions, granules,
powders, emulsions, capsules, syrups, or elixirs, for example.
Orally administered pharmaceutical compositions may contain one or
more optional agents, for example, sweetening agents such as
fructose, aspartame or saccharin; flavoring agents such as
peppermint, oil of wintergreen, or cherry coloring agents and
preserving agents, to provide a pharmaceutically palatable
preparation. Moreover, when in tablet or pill form, the
pharmaceutical compositions may be coated to delay disintegration
and absorption in the gastrointestinal tract, thereby providing a
sustained action over an extended period of time. Selectively
permeable membranes surrounding an osmotically active driving
compound are also suitable for orally administered compounds. In
these later platforms, fluid from the environment surrounding the
capsule is imbibed by the driving compound, which swells to
displace the agent or agent composition through an aperture. These
delivery platforms can provide an essentially zero order delivery
profile as opposed to the spiked profiles of immediate release
formulations. A time delay material such as glycerol monostearate
or glycerol stearate may also be used. Oral compositions can
include standard vehicles such as mannitol, lactose, starch,
magnesium stearate, sodium saccharine, cellulose, magnesium
carbonate, etc. Such vehicles are preferably of pharmaceutical
grade.
[0116] For oral liquid preparations such as, for example,
suspensions, elixirs and solutions, suitable carriers, excipients
or diluents include water, saline, alkyleneglycols (e.g., propylene
glycol), polyalkylene glycols (e.g., polyethylene glycol) oils,
alcohols, slightly acidic buffers between pH 4 and pH 6 (e.g.,
acetate, citrate, ascorbate at between about 5.0 mM to about 50.0
mM), etc. Additionally, flavoring agents, preservatives, coloring
agents, bile salts, acylcarnitines and the like may be added.
[0117] For buccal administration, the pharmaceutical compositions
may take the form of tablets, lozenges, etc. formulated in
conventional manner.
[0118] Liquid drug formulations suitable for use with nebulizers
and liquid spray devices and EHD aerosol devices typically include
a cyclic nitro compound with a pharmaceutically acceptable vehicle.
Preferably, the pharmaceutically acceptable vehicle is a liquid
such as alcohol, water, polyethylene glycol or a perfluorocarbon.
Optionally, another material may be added to alter the aerosol
properties of the solution or suspension of compounds. Preferably,
this material is liquid such as an alcohol, glycol, polyglycol or a
fatty acid. Other methods of formulating liquid drug solutions or
suspension suitable for use in aerosol devices are known to those
of skill in the art (see, e.g., Biesalski, U.S. Pat. No. 5,112,598;
Biesalski, U.S. Pat. No. 5,556,611).
[0119] A cyclic nitro compound may also be formulated in rectal or
vaginal pharmaceutical compositions such as suppositories or
retention enemas, e.g., containing conventional suppository bases
such as cocoa butter or other glycerides.
[0120] In addition to the formulations described previously, a
cyclic nitro compound may also be formulated as a depot
preparation. Such long acting formulations may be administered by
implantation (for example, subcutaneously or intramuscularly) or by
intramuscular injection. Thus, for example, a cyclic nitro compound
may be formulated with suitable polymeric or hydrophobic materials
(e.g., as an emulsion in an acceptable oil) or ion exchange resins,
or as sparingly soluble derivatives, such as a sparingly soluble
salt.
[0121] When a cyclic nitro compound is acidic or basic, it may be
included in any of the above-described formulations as the free
acid or free base, a pharmaceutically acceptable salt, a solvate or
hydrate. Pharmaceutically acceptable salts substantially retain the
activity of the free acid or base, may be prepared by reaction with
bases or acids and tend to be more soluble in aqueous and other
protic solvents than the corresponding free acid or base form.
[0122] Doses
[0123] A cyclic nitro compound and/or pharmaceutical composition
thereof, will generally be used in an amount effective to achieve
the intended purpose. For use to treat or prevent the above
diseases or disorders the cyclic nitro compound and/or
pharmaceutical compositions thereof, are administered or applied in
a therapeutically effective amount.
[0124] The amount of a cyclic nitro compound and/or pharmaceutical
composition thereof that will be effective in the treatment of a
particular disorder or condition disclosed herein will depend on
the nature of the disorder or condition, and can be determined by
standard clinical techniques known in the art. In addition, in
vitro or in vivo assays may optionally be employed to help identify
optimal dosage ranges. The amount of a cyclic nitro compound and/or
pharmaceutical composition thereof administered will, of course, be
dependent on, among other factors, the subject being treated, the
weight of the subject, the severity of the affliction, the manner
of administration and the judgment of the prescribing
physician.
[0125] For example, the dosage may be delivered in a pharmaceutical
composition by a single administration, by multiple applications or
controlled release. Dosing may be repeated intermittently, may be
provided alone or in combination with other drugs and may continue
as long as required for effective treatment of the disease state or
disorder.
[0126] Suitable dosage ranges for oral administration are dependent
on the efficiency of radiosensitization, but are generally about
0.001 mg to about 100 mg of the cyclic nitro compound per kg body
weight. Dosage ranges may be readily determined by methods known to
the artisan of ordinary skill.
[0127] Suitable dosage ranges for intravenous (i.v.) administration
are about 0.01 mg to about 100 mg per kg/body weight. Suitable
dosage ranges for intranasal administration are generally about
0.01 mg/kg body weight to about 1 mg/kg body weight. Suppositories
generally contain about 0.01 milligram to about 50 milligrams of a
cyclic nitro compound per kg/body weight and comprise active
ingredient in the range of about 0.5% to about 10% by weight.
Recommended dosages for intradermal, intramuscular,
intraperitoneal, subcutaneous, epidural, sublingual or
intracerebral administration are in the range of about 0.001 mg to
about 200 mg per kg/body weight. Effective doses may be
extrapolated from dose-response curves derived from in vitro or
animal model test systems. Such animal models and systems are
well-known in the art.
[0128] The cyclic nitro compounds are preferably assayed in vitro
and in vivo, for the desired therapeutic or prophylactic activity,
prior to use in humans. For example, in vitro assays can be used to
determine whether administration of a specific cyclic nitro
compound or a combination of cyclic nitro compounds is preferred.
The cyclic nitro compound may also be demonstrated to be effective
and safe using animal model systems.
[0129] Preferably, a therapeutically effective dose of a cyclic
nitro compound and/or pharmaceutical composition thereof described
herein will provide therapeutic benefit without causing substantial
toxicity. Toxicity of cyclic nitro compounds and/or pharmaceutical
compositions thereof may be determined using standard
pharmaceutical procedures and may be readily ascertained by the
skilled artisan. The dose ratio between toxic and therapeutic
effect is the therapeutic index. A cyclic nitro compound and/or
pharmaceutical composition thereof will preferably exhibit
particularly high therapeutic indices in treating disease and
disorders characterized by aberrant abnormal cell proliferation.
The dosage of a cyclic nitro compound and/or pharmaceutical
composition thereof described herein will preferably be within a
range of circulating concentrations that include an effective dose
with little or no toxicity.
[0130] Combination Therapy
[0131] In certain embodiments of the present invention, cyclic
nitro compounds and/or pharmaceutical compositions thereof can be
used in combination therapy with at least one other therapeutic
agent. The cyclic nitro compound and/or pharmaceutical composition
thereof and the therapeutic agent can act additively or, more
preferably, synergistically. In one embodiment, a cyclic nitro
compound and/or a pharmaceutical composition thereof is
administered concurrently with the administration of another
therapeutic agent. In another embodiment, a cyclic nitro compound
and/or pharmaceutical composition thereof is administered prior or
subsequent to administration of another therapeutic agent.
[0132] In particular, in one embodiment, cyclic nitro compounds
and/or pharmaceutical compositions thereof can be used in
combination therapy with other chemotherapeutic agents (e.g.,
alkylating agents (e.g., nitrogen mustards (e.g., cyclophosphamide,
ifosfamide, mechlorethamine, melphalen, chlorambucil,
hexamethylmelamine, thiotepa), alkyl sulfonates (e.g., busulfan),
nitrosoureas, triazines)), antimetabolites (e.g., folic acid
analogs, pyrimidine analogs (e.g., fluorouracil, floxuridine,
cytosine arabinoside, etc.), purine analogs (e.g., mercaptopurine,
thiogunaine, pentostatin, etc.), natural products (e.g.,
vinblastine, vincristine, etoposide, tertiposide, dactinomycin,
daunorubicin, doxurubicin, bleomycin, mithrmycin, mitomycin C,
L-asparaginase, interferon alpha), platinum coordination complexes
(e.g., cis-platinum, carboplatin, etc.), apoptosis inducing agents,
glutathione depleting agents or other agents that can alter the
redox status of the cell. Those of skill in the art will appreciate
that cyclic nitro compounds may also be used in concurrent
combination therapy with both the chemotherapeutic agents listed
above and radiotherapy.
[0133] Therapeutic Kits
[0134] The current invention also provides therapeutic kits
comprising cyclic nitro compounds and/or pharmaceutical
compositions thereof. The therapeutic kits may also contain other
compounds (e.g., chemotherapeutic agents, natural products,
apoptosis-inducing agents, etc.) or pharmaceutical compositions
thereof.
[0135] Therapeutic kits may have a single container which contains
a cyclic nitro compound and/or pharmaceutical compositions thereof
with or without other components (e.g., other compounds or
pharmaceutical compositions of these other compounds) or may have
distinct container for each component. Preferably, therapeutic kits
include a cyclic nitro compound and/or a pharmaceutical composition
thereof packaged for use in combination with the co-administration
of a second compound (preferably, a chemotherapeutic agent, a
natural product, an apoptosis-inducing agent, etc.) or a
pharmaceutical composition thereof. The components of the kit may
be pre-complexed or each component may be in a separate distinct
container prior to administration to a patient.
[0136] The components of the kit may be provided in one or more
liquid solutions, preferably, an aqueous solution, more preferably,
a sterile aqueous solution. The components of the kit may also be
provided as solids, which may be converted into liquids by addition
of suitable solvents, which are preferably provided in another
distinct container.
[0137] The container of a therapeutic kit may be a vial, test tube,
flask, bottle, syringe, or any other means of enclosing a solid or
liquid. Usually, when there is more than one component, the kit
will contain a second vial or other container, which allows for
separate dosing. The kit may also contain another container for a
pharmaceutically acceptable liquid.
[0138] Preferably, a therapeutic kit will contain apparatus (e.g.,
one or more needles, syringes, eye droppers, pipette, etc.), which
enables administration of the components of the kit.
EXAMPLES
[0139] The invention is further defined by reference to the
following examples, which describe in detail, preparation of
compounds and methods for assaying for biological activity. It will
be apparent to those skilled in the art that many modifications,
both to materials and methods, may be practiced without departing
from the scope.
Example 1: Production of ROS in Tumor
[0140] Cells by ABDNAZ and Irradiation
[0141] Human colon cancer cell line HT29 cells and murine squamous
cell carcinoma cell line SCC VII cells were grown in 96-well plate
overnight at 37.degree. C. and then a fluorescent probe
2'7'-dichlorofluorescin diacetate (DCFH-DA) was added at a
concentration of 20 .mu.M for 1 hour and then washed out. ABDNAZ
was added in the growth media at concentrations of 1 .mu.M, 10
.mu.M or 100 .mu.M. The green fluorescence was observed under a
fluorescence microscope and measured using a microplate
spectrofluorometer with an excitation at 488 nm and an emission at
525 nm. For cells that were treated with both ABDNAZ and
irradiation, the plates were irradiated immediately after addition
of ABDNAZ using a .sup.137Cs source.
[0142] FIG. 1 shows the production of reactive oxygen species (ROS)
in HT29 cells and SCC VII cells after exposure to ABDNAZ. The
production of ROS was dose, time and cell line dependent. The ROS
production in HT29 cells was gradually increased over time and
peaked at 24 hours. For SCC VII cells, the production of ROS was
peaked 2 hours after addition of ABDNAZ, and the levels of ROS were
significantly higher than that induced in HT29 cells.
[0143] FIGS. 2 and 3 illustrate the ROS production in HT29 cells
and SCC VII cells treated with ABDNAZ and radiation. Combined
treatment of ABDNAZ and radiation synergistically induced
intracellular ROS generation in HT29 cells and SCC VII cells, as
compared with each modality alone.
Example 2: Inhibition of Proliferation of HL60 Cells by ABDNAZ
[0144] HL60 cell line which is an acute promyelocytic leukemia cell
line was stably transfected with bcl-2 oncogene (HL60 bcl-2 cells).
The HL60 neo cells were used as a control (HL60 neo). Cells were
grown in RPMI1640 media in the presence of ABDNAZ at a
concentration of 1 uM, 2 uM or 5 uM. The number of viable cells was
counted daily for 10 days. The cell growth curves were shown in
FIG. 4 which demonstrates that ABDNAZ inhibited cell growth in a
dose-dependent manner. A dose of 5 uM of ABDNAZ inhibited cell
growth by >95% and HL60 bcl-2 cells were as sensitive as neo
cells to ABDNAZ.
Example 3: Induction of Apoptosis of HL 60 Cells by ABDNAZ
[0145] Cells, prepared and grown as described in Example 2, supra,
were collected at 8, 24, 48, and 72 hours after addition of ABDNAZ,
and analyzed using FACS. FIG. 5 illustrates the percent of
apoptosis vs. time in the presence of ABDNAZ. As can be seen in
FIGS. 5, 6 and 7, ABDNAZ induced a very high level of apoptotic
cell death in both HL60 neo and bcl-2 cells in a dose-dependent
manner. ABDNAZ at 5 uM induced 95% and 78% apoptosis at 48 hours
for neo and bcl-2 cells, respectively. At 2 uM, ABDNAZ produced
apoptotic cell death that was very similar in HL60 neo and bcl-2
cells with peaks of -40% at 8 hours. FIGS. 6 and 7 illustrates the
detailed histograms of FAGS analysis for HL60 neo cells and HL60
bcl-2 cells, respectively.
Example 4: Inhibition of bcl-2 Oncogene Expression by ABDNAZ
[0146] HL60 cells were treated as described in Example 2,supra.
Cells were collected at 6 and 24 hours for Western blot analysis.
As shown in FIG. 8, ABDNAZ at 2 and 5 uM inhibited bcl-2 protein
expression in both neo and bcl-2 cells in a dose-dependent manner.
The bcl-2 protein in HL60 bcl-2 transfected cells may be cleaved in
the presence of 2 uM ABDNAZ as indicated by the presence of the
lower molecular weight bands after both 6 and 24 hours.
Example 5: Synthesis of ABDNAZ
[0147] A 25 ml, three-neck, round bottom flask was charged with 7
ml of methylene chloride and 2.50 g (12.3 mmol) of t-BuDNAZ
prepared as described in Archibald et al., Journal of Organic
Chemistry, 1990, 2920. Under nitrogen, 0.16 ml (1.23 mmol) of boron
trifluoride etherate was added. After stirring 5 min. at ambient
temperature, 0.54 ml (6.15 mol) of bromoacetyl bromide was added.
The solution was heated between 50-60.degree. C. for 2 h. The
darkened reaction mixture was cooled to ambient temperature,
diluted with 50 ml methylene chloride, and filtered. The solid was
identified as the HBr salt oft-BuDNAZ. The methylene chloride
filtrate was washed with two 20 ml portions of water, dried over
sodium sulfate, filtered, and evaporated under reduced pressure.
The resultant solid was washed with three 20 ml portions of ethyl
ether and dried under vacuum to yield 1.24 g (75.2% based on
bromoacetyl bromide) of BrADNAZ as a white solid
(mp=124-125.degree. C.). .sup.1H NMR (CDCl.sub.3): .delta. 3.76 (s,
2H), 4.88 (br s, 2H), 5.14 (br s, 2H); .sup.13C NMR (CDCl.sub.3):
.delta. 165.2, 105.0, 59.72, 57.79, 23.90. Calc. for
C.sub.5H.sub.6BrN.sub.3O.sub.5: % C, 22.41, % H, 2.26, % N, 15.68;
Found: % C, 22.61, % H, 2.36, % N, 15.58. HPLC/MS C-8 reverse phase
column with acetonitrile/water mobile phase--m/e 266.95 (100%),
268.95 (98.3%). FT-IR 3014.24 (weak), 1677.66, 1586.30, 1567.65,
1445.55 (NO2), 1367.80, 1338.00, 1251.27 cm.sup.1.
Example 6: Synthesis of N-(chloroacetyl)-3,3-dinitroazetidine
(ClADNAZ)
[0148] A 25 ml, three-neck, round bottom flask was charged with 7
ml of methylene chloride and 2.50 g (12.3 mmol) of t-BuDNAZ. Under
nitrogen, 0.16 ml (1.23 mmol) of boron trifluoride etherate was
added. After stirring 5 min. at ambient temperature, 0.54 ml (6.15
mol) of chloroacetyl chloride was added. The solution was heated
between 50-60.degree. C. for 2 h. The darkened reaction mixture was
cooled to ambient temperature, diluted with 50 ml methylene
chloride and filtered. The solid was identified as the HBr salt
oft-BuDNAZ. The methylene chloride filtrate was washed with two 20
ml portions of water, dried over sodium sulfate, filtered, and
evaporated under reduced pressure. The resultant solid was washed
with three 20 ml portions of ethyl ether and dried under vacuum to
afford a white solid (mp: 130-132.degree. C.) in 60% yield. CHN for
C.sub.5H.sub.6ClN.sub.3O.sub.5: Found C: 26.94%, H: 2.53%,
N:17.77%; Calculated C: 26.86%, H: 2.71% N: 18.79%. FTIR: 2979
(weak), 1690.01, 1577.57, 1438.91 (NO.sub.2), 1368.21, 1338.99,
1286.21 cm.sup.-1. .sup.1H NMR: (DMSO-d.sub.6), .delta. 5.09 (2H),
4.81 (2H), 3.77 (2H). .sup.13C NMR: (DMSO-d.sub.6), .delta. 168.58,
106.98, 60.39, 50.38. HPLC: >98% pure. Safety Data: ABL Impact:
80 cm; ABL Friction: 800@ 8 ft/sec; TC ESD Unconfined at 50%: 1.10
Joules (mass ignition on bulk test). DSC Onset: 259.56.degree.
C.
Example 7: Synthesis of
N-Iodoacetyl-3,3-dinitroazetidine(IADNAZ)
[0149] A 100 ml, three neck round bottom flask was charged with 40
mL of anhydrous acetone and 2.01 g of BrADNAZ under nitrogen. 1.4 g
K.sub.2CO.sub.3 was added followed by the addition of 1.2 g sodium
iodide. The reaction mixture was allowed to reflux overnight and
monitored by proton NMR. The darkened solution was diluted with
methylene chloride, the solid was filtered and the filtrate was
extracted with 2.times.30 mL portions of methylene chloride and
water. The organic layer was dried over sodium sulfate and
concentrated under vacuum. The solid was purified by flash column
chromatography (10% ethyl acetate/hexanes) to yield a white solid
(mp 97-100.degree. C.) in 80% yield. Analysis for
C.sub.5H.sub.6IN.sub.3O.sub.5: Found C: 19.67%, H: 1.80%, N:
12.70%. Calculated C: 19.06%, H: 1.92%, N: 13.24%. FTIR: 2980
(weak), 1667.44, 1568.49, 1439.74 (NO.sub.2), 1373.69, 1335.60,
1305.35 cm.sup.-. .sup.1H NMR: (DMSO-d.sub.6), .delta. 5.09 (2H),
4.81 (2H), 3.77 (2H). .sup.13C NMR: (DMSO-d.sub.6), .delta. 168.58,
106.98, 60.39, 50.38. HPLC: >98% pure. Safety Data: ABL Impact:
80 cm; ABL Friction: 800@ 8 ft/sec; TC ESD, Unconfined 50% 7.30
Joules (no mass ignition on bulk test); SBAT Onset: 286.degree. F.
DSC Onset: 253.52.degree. C.
Example 8: Synthesis of
N-Azidoacetyl-3,3-dinitroazetidine(AzADNAZ)
[0150] A 100 ml, three neck flask was charged with 40 mL of
anhydrous acetone and 2.01 g of BrADNAZ under nitrogen. 1.05 g
K.sub.2CO.sub.3 was added followed by the addition of 0.4 g sodium
azide. The reaction mixture was allowed to reflux overnight and
monitored by proton NMR. The darkened solution was diluted with
methylene chloride and the solid was filtered. The filtrate was
extracted with 2 30 mL portions of methylene chloride and water.
The organic layer was dried over sodium sulfate and concentrated
under vacuum. The solid was purified by flash column chromatography
(10% Ethyl acetate/Hexanes) to yield a white solid (mp
103-104.degree. C.) in 80% yield. Analysis for
C.sub.5H.sub.6N.sub.6O.sub.5: Found C: 26.84%, H: 2.70%, N: 35.49%.
Calculated C: 26.09%, H: 2.63%, N: 34.76%. FTIR: 2981.60 (weak),
2109.15 (strong), 1678.88, 1598.80, 1571.47, 1463.18 (NO.sub.2),
1446.89, 1332.20, 1275.28 cm.sup.-1. .sup.1H NMR: (DMSO-d.sub.6),
.delta. 5.08 (2H), 4.83 (2H), 4.02 (2H). .sup.13C NMR:
(DMSO-d.sub.6), .delta. 169.098, 107.74, 59.84, 58.16. HPLC:
>99.7% pure. Safety Data: ABL Impact: 64 cm; ABL Friction: 800@
8 ft/sec; TC ESD, Unconfined 50% <0.5 Joules (no mass ignition
on bulk test); SBAT Onset: 314.degree. F.
Example 9: Synthesis of N-Succinyl-3,3-Dinitroazetidine
[0151] A 100 ml, three neck round bottom flask was charged with 30
mL of anhydrous dichloromethane and 5.0 grams of
tert-butyl-3,3-dinitroazetidine(t-BDNAZ) under nitrogen. 4.5 grams
of succinyl chloride was added followed by the addition of 0.5 mL
of boron trifluoride etherate. The reaction mixture was heated to
50.degree. C. and monitored by NMR. The reaction mixture was poured
slowly into ice and then filtered. The brown solid was washed with
3.times.20 mL portions of dichloromethane, dried with sodium
sulfate and concentrated under vacuum. The solid was purified by
flash column chromatography (10% ethyl acetate/hexanes) to yield a
pale white solid in 20% yield (mp: 190-192.degree. C.). Analysis
for C.sub.7H.sub.9N.sub.3O.sub.7: Found C: 33.93%, H: 3.63%, N:
19%. Calculated C: 34.02%; H: 3.67%; N: 17.00. FTIR: 3004.44
(weak), 1644.78 (strong), 1558.45, 1472.60, 1450.06, 1423.01,
1369.90, 1338.05, 1310.05, 1260.99 cm.sup.-1. .sup.1H NMR:
(DMSO-d.sub.6), .delta. 5.27 (2H), 4.85 (2H), 2.03 (4H). HPLC:
>97%. Safety Data: ABL Impact: 64 cm; ABL Friction: 800@ 8
ft/sec; TC ESD, Unconfined at 50% <0.26 Joules (no mass ignition
at 8 Joules). DSC Onset: 253.86.degree. C.
Example 10: Synthesis of N-Fumaryl-3,3-Dinitroazetidine
[0152] A 100 ml, three neck round bottom flask was charged with
8.69 grams of tert-butyl-3,3-dinitroazetidine(t-BDNAZ) under
nitrogen and 5 mL of furmaryl chloride was added followed by the
addition of 0.5 mL of boron trifluoride etherate at 0.degree. C.
for 2 hours. The reaction mixture was monitored by NMR. The thick
paste was washed with methanol and then poured into ice-water. The
solid was filtered and washed with 200 mL of water and dried under
vacuum which afforded a pale yellow solid in 20% yield (mp:
240.degree. C.). Analysis for C.sub.7H.sub.7N.sub.3O.sub.7: Found
C: 34.9%, H: 3.2%, N: 19.6%. Calculated C: 34.3%; H: 2.9%; N: 17.1.
FTIR: 3082.73 (weak), 1664.79 (strong), 1577.69, 1430.19
(NO.sub.2), 1366.92, 1274.30, 1231.24, 1213.45 cm.sup.-1. .sup.1H
NMR: (DMSO-d.sub.6), .delta. 5.88 (2H), 5.29 (2H), 4.90 (2H).
HPLC:>96%. Safety Data: ABL Friction: 800@ 8 ft/sec, TC ESD
Unconfined at 50%: 1.05 Joules (mass ignition on bulk test).
Example 11: Synthesis of N-Trifluromethyl-3,3-Dinitroazetidine
[0153] A 100 ml, three neck round bottom flask was charged with
2.28 grams of tert-butyl-3,3-dinitroazetidine(t-BDNAZ) under
nitrogen. 10 mL of trifluoroacetic anhydride was added followed by
the addition of 0.3 mL of boron trifluoride etherate. The reaction
mixture was heated to 50.degree. C. and monitored by NMR. The
reaction was concentrated under vacuum. The residual oil was washed
with water. The residual oil was added to hot hexanes and to afford
460 mg of white needles (mp: 70-71.degree. C.). Analysis for
C.sub.5H.sub.4F.sub.3N.sub.3O.sub.5: Found C: H: N: Calculated C:
24.70%, H: 1.66%, N: 17.29%. FTIR: 2991 (weak), 1716, 1683.96,
1591.37, 1576.43 (NO.sub.2), 1165.92, 1134.12 cm.sup.-1. .sup.1H
NMR: (DMSO-d.sub.6), .delta. 5.39 (2H), 5.04 (2H). HPLC: >96%
pure. Safety Data: ABL Friction: 800@ 8 ft/sec; TC ESD Unconfined
at 50%: >8 Joules. DSC Onset: 240.75.degree. C.s
[0154] Finally, it should be noted that there are alternative ways
of implementing the present invention. Accordingly, the present
embodiments are to be considered as illustrative and not
restrictive, and the invention is not to be limited to the details
given herein, but may be modified within the scope and equivalents
of the appended claims. All publications and patents cited herein
are incorporated by reference.
[0155] All references and publications cited herein are
incorporated by reference in their entirety.
* * * * *