U.S. patent application number 14/538447 was filed with the patent office on 2015-05-21 for parp inhibitor compounds, compositions and methods of use.
This patent application is currently assigned to EISAI INC.. The applicant listed for this patent is EISAI INC.. Invention is credited to Greg Delahanty, Ling Wei, Weizheng XU, Jie Zhang.
Application Number | 20150141429 14/538447 |
Document ID | / |
Family ID | 40526672 |
Filed Date | 2015-05-21 |
United States Patent
Application |
20150141429 |
Kind Code |
A1 |
XU; Weizheng ; et
al. |
May 21, 2015 |
PARP INHIBITOR COMPOUNDS, COMPOSITIONS AND METHODS OF USE
Abstract
The present invention relates to tetraaza phenalen-3-one
compounds which inhibit poly (ADP-ribose) polymerase (PARP) and are
useful in the chemosensitization of cancer therapeutics. The
induction of peripheral neuropathy is a common side-effect of many
of the conventional and newer chemotherapies. The present invention
further provides means to reliably prevent or cure
chemotherapy-induced neuropathy. The invention also relates to the
use of the disclosed PARP inhibitor compounds in enhancing the
efficacy of chemotherapeutic agents such as temozolomide. The
invention also relates to the use of the disclosed PARP inhibitor
compounds to radiosensitize tumor cells to ionizing radiation. The
invention also relates to the use of the disclosed PARP inhibitor
compounds for treatment of cancers with DNA repair defects.
Inventors: |
XU; Weizheng; (Ellicott
City, MD) ; Delahanty; Greg; (Nottingham, MD)
; Wei; Ling; (Lutherville, MD) ; Zhang; Jie;
(Ellicott City, MD) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
EISAI INC. |
Woodcliff Lake |
NJ |
US |
|
|
Assignee: |
EISAI INC.
Woodcliff Lake
NJ
|
Family ID: |
40526672 |
Appl. No.: |
14/538447 |
Filed: |
November 11, 2014 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
|
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13527158 |
Jun 19, 2012 |
8894989 |
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14538447 |
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12244399 |
Oct 2, 2008 |
8236802 |
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13527158 |
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60977115 |
Oct 3, 2007 |
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Current U.S.
Class: |
514/243 ;
514/248 |
Current CPC
Class: |
C07D 487/06 20130101;
A61P 35/00 20180101; A61P 43/00 20180101; C07D 519/00 20130101;
A61K 31/519 20130101; A61K 31/53 20130101; A61P 35/02 20180101 |
Class at
Publication: |
514/243 ;
514/248 |
International
Class: |
A61K 31/519 20060101
A61K031/519; A61K 31/53 20060101 A61K031/53 |
Claims
1-49. (canceled)
50. A method of treating a mammal having a cancer characterized by
having a defect in the homologous recombination (HR) pathway of
double-stranded DNA repair, comprising administering to said mammal
a compound selected from the compound 37: ##STR00094## and
pharmaceutically acceptable salts, esters, solvates or hydrates
thereof.
51. The method of claim 50, wherein the compound is a
pharmaceutically acceptable salt, ester, solvate or hydrate of
compound 37.
52. The method of claim 51, wherein the pharmaceutically acceptable
salt is the salt of an organic acid.
53. The method of claim 50, wherein the compound is:
##STR00095##
54. The method of claim 50, further comprising administering a
chemotherapeutic agent selected from the group consisting of
temozolomide, adriamycin, camptothecin, carboplatin, cisplatin,
daunorubicin, docetaxel, doxorubicin, interferon-alpha,
interferon-beta, interferon-gamma, interleukin 2, irinotecan,
paclitaxel, topotecan, a taxoid, dactinomycin, danorubicin,
4'-deoxydoxorubicin, bleomycin, pilcamycin, mitomycin, neomycin,
gentamycin, etoposide, 4-OH cyclophosphamide, a platinum
coordination complex, and mixtures thereof.
55. The method of claim 54, wherein said chemotherapeutic agent is
temozolomide, or a salt thereof.
56. The method of claim 50, wherein said mammal is a human.
57. The method of claim 50, wherein the cancer has a phenotype
selected from the group consisting of i) a BRCA-I defect, ii) a BRC
A-2 defect, iii) a BRCA-I and BRCA-2 defect, and iv) Fanconi
anemia.
58. The method of claim 50, wherein the cancer is selected from
breast cancer or ovarian cancer.
Description
[0001] This application claims the benefit of U.S. Provisional
Application No. 60/977,115, filed Oct. 3, 2007, the entire contents
of which are hereby incorporated by reference.
FIELD OF THE INVENTION
[0002] The present invention relates to tetraaza phenalen-3-one
compounds which inhibit poly (ADP-ribose) polymerase (PARP).
BACKGROUND
[0003] The present invention relates to inhibitors of the nuclear
enzyme poly(adenosine 5'-diphospho-ribose) polymerase
["poly(ADP-ribose) polymerase" or "PARP", which is also referred to
as ADPRT (NAD:protein (ADP-ribosyl transferase (polymerising)) and
PARS (poly(ADP-ribose) synthetase) and provides compounds and
compositions containing the disclosed compounds. Moreover, the
present invention provides methods of using the disclosed PARP
inhibitors to treat cancer.
[0004] There is considerable interest in the development of PARP
inhibitors as chemosensitizers for use in cancer therapy and to
limit cellular damage after ischemia or endotoxic stress. In
particular, potentiation of temozolomide cytotoxicity observed in
preclinical studies with potent PARP-1 inhibitors reflects
inhibition of base excision repair and subsequent cytotoxicity due
to incomplete processing of N.sup.7-methylguanine and
N.sup.3-methyladenine. There is now a body of preclinical data
demonstrating that the cytotoxicity of temozolomide is potentiated
by coadministration of a PARP inhibitor either in vitro or in vivo.
Plummer, et al., Clin. Cancer Res., 11(9), 3402 (2005).
[0005] Temozolomide, a DNA methylating agent, induces DNA damage,
which is repaired by O.sup.6-alkylguanine alkyltransferase (ATase)
and poly(ADP-ribose) polymerase-1 (PARP-1)-dependent base excision
repair. Temozolomide is an orally available monofunctional DNA
alkylating agent used to treat gliomas and malignant melanoma.
Temozolomide is rapidly absorbed and undergoes spontaneous
breakdown to form the active monomethyl triazene,
5-(3-methyl-1-triazeno)imidazole-4-carboxamide. Monomethyl triazene
forms several DNA methylation products, the predominate species
being N.sup.7-methylguanine (70%), N.sup.3-methyladenine (9%), and
O.sup.6-methylguanine (5%). Unless repaired by O.sup.6-alkylguanine
alkyltransferase, O.sup.6-methylguanine is cytotoxic due to
mispairing with thymine during DNA replication. This mispairing is
recognized on the daughter strand by mismatch repair proteins and
the thymine excised. However, unless the original
O.sup.6-methylguanine nucleotide in the parent strand is repaired
by ATase-mediated removal of the methyl adduct, thymine can be
reinserted. Repetitive futile rounds of thymine excision and
incorporation opposite an unrepaired O.sup.6-methylguanine
nucleotide causes a state of persistent strand breakage and the
MutS branch of mismatch repair system signals G2-M cell cycle
arrest and the initiation of apoptosis. The quantitatively more
important N.sup.7-methylguanine and N.sup.3-methyladenine
nucleotide alkylation products formed by temozolomide are rapidly
repaired by base excision repair. Plummer, et al., Clin. Cancer
Res., 11(9), 3402 (2005).
[0006] Chemosensitization by PARP inhibitors is not limited to
temozolomide. Cytotoxic drugs, generally, or radiation can induce
activation of PARP-1, and it has been demonstrated that inhibitors
of PARP-1 can potentiate the DNA damaging and cytotoxic effects of
chemotherapy and irradiation. Kock, et al., 45 J. Med. Chem. 4961
(2002). PARP-1 mediated DNA repair in response to DNA damaging
agents represents a mechanism for drug resistance in tumors, and
inhibition of this enzyme has been shown to enhance the activity of
ionizing radiation and several cytotoxic antitumor agents,
including temozolomide and topotecan. Suto et al., in U.S. Pat. No.
5,177,075, disclose several isoquinolines used for enhancing the
lethal effects of ionizing radiation or chemotherapeutic agents on
tumor cells. Weltin et al., "Effect of 6(5H)-Phenanthridinone, an
Inhibitor of Poly(ADP-ribose) Polymerase, on Cultured Tumor Cells",
Oncol. Res., 6:9, 399-403 (1994) disclose the inhibition of PARP
activity, reduced proliferation of tumor cells, and a marked
synergistic effect when tumor cells are co-treated with an
alkylating drug. PARP-1 is thus a potentially important therapeutic
target for enhancing DNA-damaging cancer therapies.
[0007] PARP inhibitors can also inhibit the growth of cells having
defects in the homologous recombination (HR) pathway of
double-stranded DNA repair. See Bryant et al., "Specific killing of
BRCA2-deficient tumours with inhibitors of poly(ADP-ribose)
polymerase," Nature 434, 913 (2005); Farmer et al., "Targeting the
DNA repair defect in BRCA mutant cells as a therapeutic strategy,"
Nature 434, 917 (2005). This effect operates without the presence
of chemosensitizers. Id. Known states associated with HR defects
include BRCA-1 defects, BRCA-2 defects, and Fanconi
anemia-associated cancers. McCabe et al., "Deficiency in the Repair
of DNA Damage by Homologous Recombination and Sensitivity to
Poly(ADP-Ribose) Polymerase Inhibition," Cancer Res. 66. 8109
(2006). Proteins identified as associated with a Fanconi anemia
include FANCA, FANCB, FANCC, FANCD2, FANCE, FANCF, FANCG, FANCL,
and FANCM. Id. For reviews, see Zaremba et al., "PARP Inhibitor
Development for Systemic Cancer Targeting," Anti-Cancer Agents in
Medicinal Chemistry 7, 515 (2007) and Lewis et al., "Clinical
poly(ADP-ribose) polymerase inhibitors for the treatment of
cancer," Curr. Opin. Investigational Drugs 8, 1061 (2007).
[0008] Large numbers of known PARP inhibitors have been described
in Banasik et al., "Specific Inhibitors of Poly(ADP-Ribose)
Synthetase and Mono(ADP-Ribosyl)-Transferase", J. Biol. Chem.,
267:3, 1569-75 (1992), and in Banasik et al., "Inhibitors and
Activators of ADP-Ribosylation Reactions", Molec. Cell. Biochem.,
138, 185-97 (1994). However, effective use of these PARP
inhibitors, in the ways discussed above, has been limited by the
concurrent production of unwanted side-effects. See Milam et al.,
"Inhibitors of Poly(Adenosine Diphosphate-Ribose) Synthesis; Effect
on Other Metabolic Processes," Science, 223, 589-91 (1984).
[0009] In addition to the above, PARP inhibitors have been
disclosed and described in the following international patent
applications: WO 00/42040; WO 00/39070; WO 00/39104; WO 99/11623;
WO 99/11628; WO 99/11622; WO 99/59975; WO 99/11644; WO 99/11945; WO
99/11649; and WO 99/59973. A comprehensive review of the state of
the art has been published by Li and Zhang in IDrugs 2001, 4(7):
804-812 (PharmaPress Ltd ISSN 1369-7056).
[0010] The ability of PARP-inhibitors to potentiate the lethality
of cytotoxic agents by chemo sensitizing tumor cells to the
cytotoxic effects of chemotherapeutic agents has been reported in,
inter alia, US 2002/0028815; US 2003/0134843; US 2004/0067949;
White A W, et al., 14 Bioorg. and Med. Chem Letts. 2433 (2004);
Canon Koch S S, et al., 45 J. Med. Chem. 4961 (2002); Skalitsky D
J, et al., 46 J. Med. Chem. 210 (2003); Farmer H, et al., 434
Nature 917 (14 Apr. 2005); Plummer E R, et al., 11(9) Clin. Cancer
Res. 3402 (2005); Tikhe J G, et al., 47 J. Med. Chem. 5467 (2004);
Griffin R. J., et al., WO 98/33802; and Helleday T, et al., WO
2005/012305.
[0011] The induction of peripheral neuropathy is a common factor in
limiting therapy with chemotherapeutic drugs. Quasthoff and
Hartung, J. Neurology, 249, 9-17 (2002). Chemotherapy induced
neuropathy is a side-effect encountered following the use of many
of the conventional (e.g., Taxol, vincritine, cisplatin) and newer
chemotherapies (e.g. velcade, epothilone). Depending on the
substance used, a pure sensory and painful neuropathy (with
cisplatin, oxaliplatin, carboplatin) or a mixed sensorimotor
neuropathy with or without involvement of the autonomic nervous
system (with vincristine, taxol, suramin) can ensue. Neurotoxicity
depends on the total cumulative dose and the type of drug used. In
individual cases neuropathy can evolve even after a single drug
application. The recovery from symptoms is often incomplete and a
long period of regeneration is required to restore function. Up to
now, few drugs are available to reliably prevent or cure
chemotherapy-induced neuropathy.
[0012] There continues to be a need for effective and potent PARP
inhibitors which enhance the lethal effects of chemotherapeutic
agents on tumor cells while producing minimal side-effects.
[0013] In addition, PARP inhibitors have been reported to be
effective in radiosensitizing hypoxic tumor cells and effective in
preventing tumor cells from recovering from potentially lethal
damage of DNA after radiation therapy, presumably by their ability
to prevent DNA repair. U.S. Pat. Nos. 5,032,617; 5,215,738; and
5,041,653.
[0014] Recent publications suggest that PARP inhibitors kill breast
cancer cells that are deficient in breast cancer associated gene-1
and -2 (BRCA1/2). These studies suggest that PARP inhibitors may be
effective for treating BRCA1/2-associated breast cancers. [Farmer
et al., Nature 2005, 434, 917; DeSoto and Deng, Intl. J. Med. Sci.
2006, 3, 117; Bryant et al., Nature, 2005, 434, 913.]
[0015] There continues to be a need for effective and potent PARP
inhibitors which enhance the lethal effects of ionizing radiation
and/or chemotherapeutic agents on tumor cells, or inhibit the
growth of cells having defects in the homologous recombination (HR)
pathway of double-stranded DNA repair, while producing minimal
side-effects.
SUMMARY OF INVENTION
[0016] The present invention provides compounds described herein,
derivatives thereof and their uses to inhibit poly(ADP-ribose)
polymerase ("PARP"), compositions containing these compounds and
methods for making and using these PARP inhibitors to treat the
effects of the conditions described herein.
[0017] The present invention also provides a tetraaza
phenalen-3-one compound of Formula (I), or a pharmaceutically
acceptable salt thereof
##STR00001##
wherein R is (a) NR.sup.1R.sup.2, wherein R.sup.1 is selected from
the group consisting of hydrogen, C.sub.1-C.sub.6 straight or
branched chain alkyl, C.sub.2-C.sub.6 straight or branched chain
alkenyl, C.sub.3-C.sub.8 cycloalkyl, C.sub.1-C.sub.6 alkoxy,
C.sub.2-C.sub.6 alkenyloxy, phenyl, phenoxy, benzyloxy,
NR.sup.AR.sup.B (C.sub.1-C.sub.6 straight or branched chain alkyl),
NR.sup.AR.sup.B (C.sub.2-C.sub.6 straight or branched chain
alkenyl), (C.sub.1-C.sub.6 straight or branched chain
alkyl)carbonyl, (C.sub.2-C.sub.6 straight or branched chain
alkenyl)carbonyl, (C.sub.3-C.sub.8 cycloalkyl)carbonyl,
(C.sub.1-C.sub.6 straight or branched chain alkyl)oxycarbonyl,
(C.sub.2-C.sub.6 straight or branched chain alkenyl)oxycarbonyl,
(C.sub.3-C.sub.8 cycloalkyl)oxycarbonyl, arylcarbonyl, sulfonyl,
arylsulfonyl, aryl(C.sub.1-C.sub.6 straight or branched chain
alkyl), aryl(C.sub.2-C.sub.6 straight or branched chain alkenyl),
aryl(C.sub.3-C.sub.8 cycloalkyl), (C.sub.1-C.sub.6 straight or
branched chain alkyl)aryl, (C.sub.2-C.sub.6 straight or branched
chain alkenyl)aryl, (C.sub.3-C.sub.8 cycloalkyl)aryl, aryl,
heterocyclyl, heterocyclyl(C.sub.1-C.sub.6 straight or branched
chain alkyl), and heterocyclyl(C.sub.2-C.sub.6 straight or branched
chain alkenyl); wherein each heterocyclyl has between 1 and 7
heteroatoms independently selected from O, N, or S, and wherein
each of R.sup.A and R.sup.B are independently selected from the
group consisting of hydrogen, C.sub.1-C.sub.6 straight or branched
chain alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl,
and C.sub.3-C.sub.8 cycloalkyl; and R.sup.2 is selected from the
group consisting of C.sub.1-C.sub.6 straight or branched chain
alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl,
C.sub.3-C.sub.8 cycloalkyl, C.sub.1-C.sub.6 alkoxy, C.sub.2-C.sub.6
alkenyloxy, phenyl, phenoxy, benzyloxy, NR.sup.XR.sup.Y
(C.sub.1-C.sub.6 straight or branched chain alkyl), NR.sup.XR.sup.Y
(C.sub.2-C.sub.6 straight or branched chain alkenyl),
(C.sub.1-C.sub.6 straight or branched chain alkyl)carbonyl,
(C.sub.2-C.sub.6 straight or branched chain alkenyl)carbonyl,
(C.sub.3-C.sub.8 cycloalkyl)carbonyl, (C.sub.1-C.sub.6 straight or
branched chain alkyl)oxycarbonyl, (C.sub.2-C.sub.6 straight or
branched chain alkenyl)oxycarbonyl, (C.sub.3-C.sub.8
cycloalkyl)oxycarbonyl, arylcarbonyl, sulfonyl, arylsulfonyl,
aryl(C.sub.1-C.sub.6 straight or branched chain alkyl),
aryl(C.sub.2-C.sub.6 straight or branched chain alkenyl),
aryl(C.sub.3-C.sub.8 cycloalkyl), (C.sub.1-C.sub.6 straight or
branched chain alkyl)aryl, (C.sub.2-C.sub.6 straight or branched
chain alkenyl)aryl, (C.sub.3-C.sub.8 cycloalkyl)aryl, aryl,
heterocyclyl, heterocyclyl(C.sub.1-C.sub.6 straight or branched
chain alkyl), and heterocyclyl(C.sub.2-C.sub.6 straight or branched
chain alkenyl); wherein each heterocyclyl has between 1 and 7
heteroatoms independently selected from O, N, or S, and wherein
each of R.sup.X and R.sup.Y are independently selected from the
group consisting of hydrogen, C.sub.1-C.sub.6 straight or branched
chain alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl,
and C.sub.3-C.sub.8 cycloalkyl; wherein R.sup.1 and R.sup.2 are
independently substituted with between 0 and 4 substituents, each
independently selected from halo, C.sub.1-C.sub.6 straight or
branched chain alkyl, C.sub.2-C.sub.6 straight or branched chain
alkenyl, C.sub.1-C.sub.6 alkoxy, trifluoromethyl, trifluoroethyl,
and amino; and provided that R.sup.1 and R.sup.2 may not both be
methyl, and R.sup.2 may not be (phenyl)prop-1-yl when R.sup.1 is
hydrogen; or (b) aryloxy, substituted with between 0 and 4
substituents, each independently selected from the group consisting
of halo, C.sub.1-C.sub.6 alkoxy, trifluoromethyl, trifluoroethyl,
C.sub.1-C.sub.6 straight or branched chain alkyl, C.sub.2-C.sub.6
straight or branched chain alkenyl, C.sub.3-C.sub.8 cycloalkyl,
NR.sup.CR.sup.D, NR.sup.CR.sup.D(C.sub.1-C.sub.6 straight or
branched chain alkyl), and NR.sup.CR.sup.D(C.sub.2-C.sub.6 straight
or branched chain alkenyl), wherein each of R.sup.C and R.sup.D is
independently selected from the group consisting of hydrogen,
C.sub.1-C.sub.6 straight or branched chain alkyl, C.sub.2-C.sub.6
straight or branched chain alkenyl, and C.sub.3-C.sub.8 cycloalkyl;
and when more than one substituent is of the form NR.sup.CR.sup.D,
each occurrence of R.sup.C and R.sup.D is independently selected
from the group consisting of hydrogen, C.sub.1-C.sub.6 straight or
branched chain alkyl, C.sub.2-C.sub.6 straight or branched chain
alkenyl, and C.sub.3-C.sub.8 cycloalkyl; or (c) a heterocyclyl
having between 1 and 7 heteroatoms independently selected from O,
N, or S; and having between 0 and 4 substituents independently
selected from the group consisting of halo, haloalkyl, hydroxyl,
nitro, trifluoromethyl, trifluoroethyl, C.sub.1-C.sub.6 straight or
branched chain alkyl, C.sub.2-C.sub.6 straight or branched chain
alkenyl, C.sub.1-C.sub.6 alkoxy, C.sub.2-C.sub.6 alkenyloxy,
phenyl, phenoxy, benzyloxy, amino, thiocarbonyl, cyano, imino,
NR.sup.ER.sup.F(C.sub.1-C.sub.6 straight or branched chain alkyl),
NR.sup.ER.sup.F(C.sub.2-C.sub.6 straight or branched chain alkenyl)
sulfhydryl, thioalkyl, dioxa-spiroethyl, (C.sub.1-C.sub.6 straight
or branched chain alkyl) carbonyl, (C.sub.2-C.sub.6 straight or
branched chain alkenyl)carbonyl, (C.sub.1-C.sub.6 straight or
branched chain alkyl)oxycarbonyl, (C.sub.2-C.sub.6 straight or
branched chain alkenyl)oxycarbonyl, arylcarbonyl, sulfonyl,
arylsulfonyl, aryl(C.sub.1-C.sub.6 straight or branched chain
alkyl), aryl(C.sub.2-C.sub.6 straight or branched chain alkenyl),
aryl(C.sub.3-C.sub.8 cycloalkyl), (C.sub.1-C.sub.6 straight or
branched chain alkyl)aryl, (C.sub.2-C.sub.6 straight or branched
chain alkenyl)aryl, (C.sub.3-C.sub.8 cycloalkyl)aryl, aryl,
heterocyclyl, heterocyclyl(C.sub.1-C.sub.6 straight or branched
chain alkyl), and heterocyclyl(C.sub.2-C.sub.6 straight or branched
chain alkenyl), wherein each heterocyclyl has between 1 and 7
heteroatoms independently selected from O, N, or S, wherein each of
R.sup.E and R.sup.F is independently selected from the group
consisting of hydrogen, C.sub.1-C.sub.6 straight or branched chain
alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl, and
C.sub.3-C.sub.8 cycloalkyl; and when more than one substituent is
of the form NR.sup.ER.sup.F each occurrence of R.sup.E and R.sup.F
is independently selected from the group consisting of hydrogen,
C.sub.1-C.sub.6 straight or branched chain alkyl, C.sub.2-C.sub.6
straight or branched chain alkenyl, and C.sub.3-C.sub.8 cycloalkyl;
wherein each of said 0-4 substituents is independently substituted
with between 0 and 4 further substituents, and each said further
substituent is independently selected from halo, C.sub.1-C.sub.6
straight or branched chain alkyl, C.sub.2-C.sub.6 straight or
branched chain alkenyl, C.sub.3-C.sub.8 cycloalkyl, C.sub.1-C.sub.6
alkoxy, trifluoromethyl, trifluoroethyl, and amino; provided that R
has at least one substituent when R is an N-piperidinyl,
N-pyrrolidinyl or an N-morpholinyl group.
[0018] In some embodiments each ring of each heterocyclyl of
Formula (I) is independently 5-7 atoms in size.
[0019] Some embodiments include one, two or three nitrogen atoms in
at least one ring of the heterocyclyl of Formula (I).
[0020] In some embodiments, the heterocyclyl of Formula(I)
comprises 1-3 rings. In some embodiments, the heterocyclyl has 1-7
heteroatoms independently selected from O, N, and S. In some
embodiments, the heterocyclyl comprises 1-2 rings. In some
embodiments, the heterocyclyl comprises one ring. In some
embodiments, the various occurrences of the heterocyclyl of Formula
(I) each independently comprise 1-3 rings. In some embodiments, the
various occurrences of the heterocyclyl of Formula (I) each
independently comprise 1-2 rings. In some embodiments, the various
occurrences of the heterocyclyl of Formula (I) each independently
comprise one ring.
[0021] In some embodiments, the heterocyclyl of Formula (I) is
selected from the group consisting of piperidinyl, piperazinyl,
pyridazinyl, dihydropyridyl, tetrahydropyridyl, pyridinyl,
pyrimidinyl, dihydropyrimidinyl, tetrahydrophyrimidinyl,
hexahydropyrimidinyl, dihydropyrazinyl, tetrahydropyrazinyl,
pyrrolidinyl, imidazolidinyl, pyrazolidinyl, pyrrolyl,
dihydropyrolyl, imidazolyl, dihydroimidazoyl, pyrazolyl,
dihydropyrazolyl, azepanyl, [1,2]diazepanyl, [1,3]diazepanyl,
[1,4]diazepanyl, indolyl, dihydroindolyl, isoindolyl,
dihydroisoindoly, dihydroquinolyl, tetrahydroquinolyl,
dihydroisoquinolyl, and tetrahydroisoquinolyl; or subsets
thereof.
[0022] The present invention also relates to a pharmaceutical
composition comprising (i) a therapeutically amount of a compound
of Formula (I) and (ii) a pharmaceutically acceptable carrier.
[0023] The present invention provides compounds which inhibit the
in vitro and/or in vivo polymerase activity of poly(ADP-ribose)
polymerase (PARP), and compositions containing the disclosed
compounds.
[0024] The present invention provides methods to inhibit, limit
and/or control the in vitro and/or in vivo polymerase activity of
poly(ADP-ribose) polymerase (PARP) in solutions cells, tissues,
organs or organ systems. In one embodiment, the present invention
provides methods of limiting or inhibiting PARP activity in a
mammal, such as a human, either locally or systemically.
[0025] In one embodiment, the invention provides a
chemosensitization method for treating cancer comprising contacting
the cancer cells with a cytotoxicity-potentiating tetraaza
phenalen-3-one compound of Formula (I) or a pharmaceutically
acceptable salt thereof and further contacting the tumor or cancer
cells with an anticancer agent.
[0026] An embodiment of the present invention provides a
chemosensitization method wherein a first dose of at least one
compound of Formula (I) or a pharmaceutically acceptable salt
thereof is administered singly or repeatedly to a patient in need
thereof, and wherein subsequently a second dose of at least one
chemotherapeutic agent is administered singly or repeatedly to said
patient after a time period to provide an effective amount of
chemosensitization.
[0027] An aspect of the present invention provides a pharmaceutical
formulation comprising the compound of Formula (I) in a form
selected from the group consisting of Non-limiting examples of such
chemotherapeutic agents are recited below. pharmaceutically
acceptable free bases, salts, hydrates, esters, solvates,
stereoisomers, and mixtures thereof. According to a further aspect,
the pharmaceutical formulation further comprises a pharmaceutically
acceptable carrier and, optionally, a chemotherapeutic agent. The
following embodiments are for illustrative purposes only and are
not intended to limit in any way the scope of the present
invention. In one embodiment, a pharmaceutical formulation of the
invention comprises a compound of the invention in a
pharmaceutically acceptable carrier. In another embodiment, a
pharmaceutical formulation of the invention comprises a
pharmaceutically acceptable salt of a compound of the invention in
a pharmaceutically acceptable carrier. In another embodiment, a
pharmaceutical formulation of the invention comprises a compound of
the invention and one or more chemotherapeutic agents in a
pharmaceutically acceptable carrier. In another embodiment, a
pharmaceutical formulation of the invention comprises a
pharmaceutically acceptable salt of a compound of the invention and
one or more chemotherapeutic agents in a pharmaceutically
acceptable carrier. Non-limiting examples of such chemotherapeutic
agents are recited below.
[0028] According to additional aspects of the invention, the
chemosensitizing compound and the chemotherapeutic agent are
administered essentially simultaneously.
[0029] According to an aspect of the invention, the
chemotherapeutic agent is selected from the group consisting of
temozolomide, adriamycin, camptothecin, carboplatin, cisplatin,
daunorubicin, docetaxel, doxorubicin, interferon-alpha,
interferon-beta, interferon-gamma, interleukin 2, irinotecan,
paclitaxel, a taxoid, dactinomycin, danorubicin,
4'-deoxydoxorubicin, bleomycin, pilcamycin, mitomycin, neomycin and
gentamycin, etoposide, 4-OH cyclophosphamide, a platinum
coordination complex, topotecan, therapeutically effective analogs
and derivatives of the same, and mixtures thereof. According to a
specific aspect, the chemotherapeutic agent is temozolomide.
[0030] In another embodiment, the present invention provides
methods of treating the effect of cancer and/or to radio sensitize
cancer cells to render the cancer cells more susceptible to
radiation therapy and thereby to prevent the tumor cells from
recovering from potentially lethal damage of DNA after radiation
therapy, comprising administering to a subject an effective amount
of a compound of Formula (I) or a pharmaceutically acceptable salt
thereof. A method of this embodiment is directed to specifically
and preferentially radiosensitizing cancer cells rendering the
cancer cells more susceptible to radiation therapy than non-tumor
cells.
[0031] The present invention also provides a method of treatment of
cancer in a subject in need thereof comprising administering to the
subject a therapeutically effective amount of a compound of Formula
(I) or a pharmaceutically acceptable salt thereof, wherein the
cancer cells have a defect in repair of double-stranded DNA
scission. In one embodiment, the defect in repair of
double-stranded DNA scission is a defect in homologous
recombination. In one embodiment, the cancer cells have a phenotype
selected from the group consisting of a BRCA-1 defect, a BRCA-2
defect, a BRCA-1 and BRCA-2 defect, and Fanconi anemia.
[0032] In another embodiment, the present invention provides
methods of treating BRCA1/2-associated breast cancer comprising
administering a compound of Formula (I) or a pharmaceutically
acceptable salt thereof.
[0033] According to one embodiment of the invention, the compound
for use in the chemosensitization method of the invention, the
radiosensitization method of the invention, or the treatment of
cancer wherein the cancer cells have a defect in repair of
double-stranded DNA scission method of the invention, is a compound
selected from Formula (I) or a pharmaceutically acceptable salt
thereof. In another aspect, the compound is selected from the group
consisting of
##STR00002## ##STR00003## ##STR00004## ##STR00005## ##STR00006##
##STR00007## ##STR00008## ##STR00009## ##STR00010##
and pharmaceutically acceptable salts thereof.
[0034] The present invention also provides means to treat
chemotherapy-induced peripheral neuropathy. According to an aspect
of the invention, the compounds of the present invention are
administered prior to, or together with, the administration of at
least one chemotherapy agent to prevent the development of
neuropathy symptoms or to mitigate the severity of such symptoms.
According to a further aspect, the compounds of the present
invention are administered after the administration of at least one
chemotherapeutic agent to treat a patient for the symptoms of
neuropathy or to mitigate the severity of such symptoms. In another
aspect, the present invention provides a method to retard, delay,
or arrest the growth of cancer cells in a mammal, comprising the
administration of a chemotherapeutic agent, and further comprising
the administration of a compound of Formula (I) or a
pharmaceutically acceptable salt thereof in an amount sufficient to
potentiate the anticancer activity of said chemotherapeutic
agent.
[0035] Still other aspects and advantages of the present invention
will become readily apparent by those skilled in the art from the
following detailed description, wherein it is shown and described
preferred embodiments of the invention, simply by way of
illustration of the best mode contemplated of carrying out the
invention. As will be realized the invention is capable of other
and different embodiments, and its several details are capable of
modifications in various obvious respects, without departing from
the invention. Accordingly, the description is to be regarded as
illustrative in nature and not as restrictive.
BRIEF DESCRIPTION OF THE DRAWINGS
[0036] FIG. 1.--The oral administration of PARP-1 inhibitor
Compound 13+ TMZ demonstrating the enhance survival of mice bearing
the B16 melanoma model.
[0037] FIG. 2.--The oral administration of PARP-1 inhibitor
Compound 13+ TMZ demonstrating the enhanced survival in the
intracranial SJGBM glioma model.
[0038] FIG. 3.--The oral administration of PARP-1 inhibitor
Compound 37+ TMZ demonstrating the enhance survival of mice bearing
the B16 melanoma model.
[0039] FIG. 4.--The oral administration of PARP-1 inhibitor
Compound 37+ TMZ demonstrating the enhanced survival in the
intracranial SJGBM glioma model.
[0040] FIG. 5.--The oral administration of PARP-1 inhibitor
Compound 37+ radiation demonstrating inhibition of tumor growth in
the model of head and neck cancer.
[0041] FIG. 6.--The oral administration of PARP-1 inhibitor
Compound 37 demonstrating inhibition of growth of BRCA1 mutant
tumors
DETAILED DESCRIPTION OF THE INVENTION
[0042] The present invention provides compounds described herein,
derivatives thereof and their uses to inhibit poly(ADP-ribose)
polymerase ("PARP"), compositions containing these compounds and
methods for making and using these compounds to treat, prevent
and/or ameliorate the effects of cancers by potentiating the
cytotoxic effects of ionizing radiation on tumor cells.
[0043] The present invention provides compounds described herein,
derivatives thereof and their uses to inhibit poly(ADP-ribose)
polymerase ("PARP"), compositions containing these compounds and
methods for making and using these compounds to treat the effects
of cancers by potentiating the cytotoxic effects of
chemotherapeutic agents on tumor cells.
[0044] The present invention provides a chemosensitization method
for treating tumor and/or cancer cells comprising contacting said
cancer cells with a compound of Formula (I) and further contacting
said cancer cells with an anticancer agent.
[0045] The present invention provides compounds described herein,
derivatives thereof and their uses to inhibit poly(ADP-ribose)
polymerase ("PARP"), compositions containing these compounds and
methods for making and using these compounds to inhibit the growth
of cells having defects in the homologous recombination (HR)
pathway of double-stranded DNA repair.
[0046] The compounds and compositions of the present invention can
be used in the presence or absence of radio- or chemo-sensitizers
for the treatment of cancer. The compounds and compositions are
preferably used in the absence of radio- or chemo-sensitizers where
the cancer has a defect in the homologous recombination (HR)
pathway of double-stranded DNA repair. Such defects are associated
with, and have the phenotypes of, BRCA-1 defects, BRCA-2 defects,
dual BRCA-1/BRCA-2 defects, and Fanconi anemia.
[0047] Fanconi anemia is a genetically heterogeneous disease and
patients with Fanconi anemia have a greatly increased risk of
cancer. Eleven proteins have been associated with Fanconi anemia.
FANCA, FANCB, FANCC, FANCE, FANCF, FANCG, and FANCM form a nuclear
core complex. The complex interacts with FANCL to incorporate
ubiquinone of FANCD2. Modified FANCD2 is need for repair of DNA
cross-links. FANCd2 accumulates at sites of DNA damage and
associates with BRCA-1 and BRCA-2.
[0048] Exemplary cancers that can be associated with HR defects
include breast cancer and ovarian cancer. Breast cancer for
treatment by the methods of the invention can include all types of
breast cancer and preferably includes invasive ductal carcinoma and
invasive lobular carcinoma. Ovarian cancer for treatment by the
methods of the invention include all types of ovarian cancer,
preferably epithelial ovarian tumors, germ cell ovarian tumors, and
sex cord stromal tumors.
[0049] The compounds of the present invention can be synthesized
using the starting materials and methods disclosed in U.S.
application Ser. No. 10/853,714, which is incorporated herein by
reference in its entirety.
[0050] Typically, the compounds, such as those of Formula (I), used
in the compositions of the invention will have an IC.sub.50 for
inhibiting poly(ADP-ribose) polymerase in vitro of about 20 .mu.M
or less, preferably less than about 10 .mu.M, more preferably less
than about 1 .mu.M, or preferably less than about 0.1 .mu.M, most
preferably less than about 0.01 .mu.M.
[0051] A convenient method to determine IC.sub.50 of a PARP
inhibitor compound is a PARP assay using purified recombinant human
PARP from Trevigan (Gaithersburg, Md.), as follows: The PARP enzyme
assay is set up on ice in a volume of 100 microliters consisting of
100 mM Tris-HCl (pH 8.0), 1 mM MgCl.sub.2, 28 mM KCl, 28 mM NaCl,
3.0 .mu.g/ml of DNase I-activated herring sperm DNA (Sigma, Mo.),
30 micromolar [.sup.3H]nicotinamide adenine dinucleotide (62.5
mCi/mmole), 15 micrograms/ml PARP enzyme, and various
concentrations of the compounds to be tested. The reaction is
initiated by adding enzyme and incubating the mixture at 25.degree.
C. After 2 minutes of incubation, the reaction is terminated by
adding 500 microliters of ice cold 30% (w/v) trichloroacetic acid.
The precipitate formed is transferred onto a glass fiber filter
(Packard Unifilter-GF/C) and washed three times with 70% ethanol.
After the filter is dried, the radioactivity is determined by
scintillation counting. The compounds of this invention were found
to have potent enzymatic activity in the range of a few nanomolar
to 20 micromolar in IC.sub.50 in this inhibition assay.
[0052] As used herein, "alkyl" means a branched or unbranched
saturated hydrocarbon chain comprising a designated number of
carbon atoms. For example, C.sub.1-C.sub.6 straight or branched
alkyl hydrocarbon chain contains 1 to 6 carbon atoms, and includes
but is not limited to substituents such as methyl, ethyl, propyl,
iso-propyl, butyl, iso-butyl, tert-butyl, n-pentyl, n-hexyl, and
the like, unless otherwise indicated. In some embodiments, the
alkyl chain is a C.sub.1 to C.sub.6 branched or unbranched carbon
chain. In some embodiments, the alkyl chain is a C.sub.2 to C.sub.5
branched or unbranched carbon chain. In some embodiments, the alkyl
chain is a C.sub.1 to C.sub.4 branched or unbranched carbon chain.
In some embodiments, the alkyl chain is a C.sub.2 to C.sub.4
branched or unbranched carbon chain. In some embodiments, the alkyl
chain is a C.sub.3 to C.sub.5 branched or unbranched carbon chain.
In some embodiments, the alkyl chain is a C.sub.1 to C.sub.2
branched or unbranched carbon chain. In some embodiments, the alkyl
chain is a C.sub.2 to C.sub.3 branched or unbranched carbon
chain.
[0053] "Alkenyl" means a branched or unbranched unsaturated
hydrocarbon chain comprising a designated number of carbon atoms.
For example, C.sub.2-C.sub.6 straight or branched alkenyl
hydrocarbon chain contains 2 to 6 carbon atoms having at least one
double bond, and includes but is not limited to substituents such
as ethenyl, propenyl, isopropenyl, butenyl, iso-butenyl,
tert-butenyl, n-pentenyl, n-hexenyl, and the like, unless otherwise
indicated. In some embodiments, the alkenyl chain is a C.sub.2 to
C.sub.6 branched or unbranched carbon chain. In some embodiments,
the alkenyl chain is a C.sub.2 to C.sub.5 branched or unbranched
carbon chain. In some embodiments, the alkenyl chain is a C.sub.2
to C.sub.4 branched or unbranched carbon chain. In some
embodiments, the alkenyl chain is a C.sub.3 to C.sub.5 branched or
unbranched carbon chain.
[0054] "Alkoxy", means the group --OZ wherein Z is alkyl as herein
defined. Z can also be a branched or unbranched saturated
hydrocarbon chain containing 1 to 6 carbon atoms.
[0055] "Cyclo", used herein as a prefix, refers to a structure
characterized by a closed ring.
[0056] "Halo" means at least one fluoro, chloro, bromo, or iodo
moiety, unless otherwise indicated.
[0057] Each of "NR.sup.AR.sup.B", "NR.sup.XR.sup.Y",
"NR.sup.CR.sup.D", and "NR.sup.ER.sup.F" as described herein
independently encompass amino (NH.sub.2) as well as substituted
amino. For example, NR.sup.AR.sup.B may be --NH(CH.sub.3),
--NH(cyclohexyl), and --N(CH.sub.2CH.sub.3)(CH.sub.3). When more
than one substituent is of the form "NR.sup.AR.sup.B",
"NR.sup.XR.sup.Y", "NR.sup.CR.sup.D", or "NR.sup.ER.sup.E", each
occurrence of R.sup.A, R.sup.B, R.sup.C, R.sup.D, R.sup.X, or
R.sup.Y is independently selected from the group consisting of
hydrogen, C.sub.1-C.sub.6 straight or branched chain alkyl,
C.sub.2-C.sub.6 straight or branched chain alkenyl, and
C.sub.3-C.sub.8 cycloalkyl. Such examples are for illustrative
purposes only and are not intended to be limiting in any way.
[0058] "Arylcarbonyl" refers to a carbonyl radical substituted with
aryl as described herein. Non-limiting examples include
phenylcarbonyl and naphthylcarbonyl.
[0059] "Alkylcarbonyl" refers to a carbonyl radical substituted
with alkyl as described herein. Non-limiting examples include acyl
and propylcarbonyl.
[0060] "Alkoxycarbonyl" refers to a carbonyl radical substituted
with alkoxy as described herein. Non-limiting examples include
methoxycarbonyl and tert-butyloxycarbonyl.
[0061] "Ar" or "aryl" refer to an aromatic carbocyclic moiety
having one or more closed rings. Examples include, without
limitation, phenyl, naphthyl, anthracenyl, phenanthracenyl,
biphenyl, and pyrenyl.
[0062] "Heterocyclyl" refers to a cyclic moiety having one or more
closed rings, with one or more heteroatoms (for example, oxygen,
nitrogen or sulfur) in at least one of the rings, and wherein the
ring or rings may independently be aromatic, nonaromatic, fused,
and/or bridged, Examples include without limitation piperidinyl,
piperazinyl, pyridazinyl, dihydropyridyl, tetrahydropyridyl,
pyridinyl, pyrimidinyl, dihydropyrimidinyl, tetrahydrophyrimidinyl,
hexahydropyrimidinyl, dihydropyrazinyl, tetrahydropyrazinyl,
pyrrolidinyl, imidazolidinyl, pyrazolidinyl, pyrrolyl,
dihydropyrolyl, imidazolyl, dihydroimidazoyl, pyrazolyl,
dihydropyrazolyl, azepanyl, [1,2]diazepanyl, [1,3]diazepanyl,
[1,4]diazepanyl, indolyl, dihydroindolyl, isoindolyl,
dihydroisoindoly, dihydroquinolyl, tetrahydroquinolyl,
dihydroisoquinolyl, and tetrahydroisoquinolyl.
[0063] "Arylalkyl" refers to an alkyl radical substituted with
aryl. Non-limiting examples include benzyl, phenylethyl, and
phenylpropyl.
[0064] "Alkylaryl" refers to an aryl radical substituted with
alkyl. Non-limiting examples include tolyl and dimethylphenyl.
[0065] "Cycloalkyl" refers to a hydrocarbon cyclic moiety that is
nonaromatic. Examples include without limitation cyclopropane,
cyclobutane, cyclopentane, cyclohexane, cyclopheptane, cyclooctane,
cyclopentene, cyclohexene, cycloheptene, and cyclooctene.
[0066] The term "nervous insult" refers to any damage to nervous
tissue and any disability or death resulting therefrom. The cause
of nervous insult may be metabolic, toxic, neurotoxic, iatrogenic,
thermal or chemical, and includes without limitation, ischemia,
hypoxia, cerebrovascular accident, trauma, surgery, pressure, mass
effect, hemorrhage, radiation, vasospasm, neurodegenerative
disease, infection, Parkinson's disease, amyotrophic lateral
sclerosis (ALS), myelination/demyelination process, epilepsy,
cognitive disorder, glutamate abnormality and secondary effects
thereof.
[0067] The term "neuroprotective" refers to the effect of reducing,
arresting or ameliorating nervous insult, and protecting,
resuscitating, or reviving nervous tissue that has suffered nervous
insult.
[0068] The term "preventing neurodegeneration" includes the ability
to prevent a neurodegenerative disease or preventing further
neurodegeneration in patients who are already suffering from or
have symptoms of a neurodegenerative disease.
[0069] The term "treating" refers to:
[0070] (i) preventing a disease, disorder or condition from
occurring in an animal that may be predisposed to the disease,
disorder and/or condition, but has not yet been diagnosed as having
it; and/or
[0071] (ii) inhibiting the disease, disorder or condition, i.e.,
arresting its development; and/or
[0072] (iii) relieving the disease, disorder or condition, i.e.,
causing regression of the disease, disorder and/or condition.
[0073] The term "chemosensitizer", as used herein, is defined as a
molecule, such as a low molecular weight molecule, administered to
animals in therapeutically effective amounts to potentiate the
antitumoral activity of chemotherapeutic agents. Such
chemosensitizers are useful, for example, to increase the tumor
growth-retarding or--arresting effect of a given dose of a
chemotherapeutic agent, or to improve the side-effect profile of a
chemotherapeutic agent by allowing for reductions in its dose while
maintaining its antitumoral efficacy.
[0074] The term "radiosensitizer", as used herein is defined as a
molecule, such as a low molecular weight molecule, administered to
animals in therapeutically effective amounts to increase the
sensitivity of the cells to be radiosensitized to electromagnetic
radiation and/or to promote the treatment of diseases which are
treatable with electromagnetic radiation. Diseases which are
treatable with electromagnetic radiation include neoplastic
diseases, benign and malignant tumors, and cancerous cells.
Electromagnetic radiation treatment of other diseases not listed
herein is also contemplated.
[0075] "Effective amount" refers to the amount required to produce
the desired effect.
[0076] "Substituted" means that at least one hydrogen on a
designated group is replaced with another radical, provided that
the designated group's normal valence is not exceeded. With respect
to any group containing one or more substituents, such groups are
not intended to introduce any substitution that is sterically
impractical, synthetically non-feasible and/or inherently unstable.
In some embodiments of the invention as described herein, a
substituent may substitute a radical, which said radical is itself
a substituent. For example, in the compound shown below for
illustrative purposes only, the piperazinyl yl ring is a
heterocyclyl, which may be substituted with 0-4 substituents as
described herein. In the example compound, the piperazinyl ring is
substituted with arylsulfonyl wherein aryl is phenyl, and wherein
the arylsulfonyl may be further substituted 0-4 times as described
herein. In the example compound, the phenylsulfonyl moiety is
further substituted with tert-butyl. Such example is given for
illustrative purposes only and is not intended to be limiting in
any way.
##STR00011##
[0077] "Subject" refers to a cell or tissue, in vitro or in vivo,
an animal or a human. An animal or human subject may also be
referred to as a "patient."
[0078] "Animal" refers to a living organism having sensation and
the power of voluntary movement, and which requires for its
existence oxygen and organic food. Examples include, without
limitation, members of the human, mammalian and primate
species.
[0079] Broadly, the compounds and compositions of the present
invention can be used to treat or prevent cell damage or death due
to necrosis or apoptosis, cerebral ischemia and reperfusion injury
or neurodegenerative diseases in an animal, such as a human. The
compounds and compositions of the present invention can be used to
extend the lifespan and proliferative capacity of cells and thus
can be used to treat or prevent diseases associated therewith; they
alter gene expression of senescent cells; and they radiosensitize
hypoxic tumor cells. Preferably, the compounds and compositions of
the invention can be used to treat or prevent tissue damage
resulting from cell damage or death due to necrosis or apoptosis,
and/or effect neuronal activity, either mediated or not mediated by
NMDA toxicity. The compounds of the present invention are not
limited to being useful in treating glutamate mediated
neurotoxicity and/or NO-mediated biological pathways. Further, the
compounds of the invention can be used to treat or prevent other
tissue damage related to PARP activation, as described herein.
[0080] The present invention provides compounds which inhibit the
in vitro and/or in vivo polymerase activity of poly(ADP-ribose)
polymerase (PARP), and compositions containing the disclosed
compounds.
[0081] The present invention provides methods to inhibit, limit
and/or control the in vitro and/or in vivo polymerase activity of
poly(ADP-ribose) polymerase (PARP) in any of solutions, cells,
tissues, organs or organ systems. In one embodiment, the present
invention provides methods of limiting or inhibiting PARP activity
in a mammal, such as a human, either locally or systemically.
[0082] The compounds of the invention act as PARP inhibitors to
treat or prevent cancers by chemopotentiating the cytotoxic effects
of the chemotherapeutic agents. The compounds of the invention act
as PARP inhibitors to treat or prevent cancers by sensitizing cells
to the cytotoxic effects of radiation. The compounds of the
invention act as PARP inhibitors to treat or prevent
BRCA1/2-associated breast cancer.
[0083] The compounds of the present invention may possess one or
more asymmetric center(s) and thus can be produced as mixtures
(racemic and non-racemic) of stereoisomers, or as individual
enantiomers or diastereomers. The individual stereoisomers may be
obtained by using an optically active staring material, by
resolving a racemic or non-racemic mixture of an intermediate at
some appropriate stage of the synthesis, or by resolution of the
compound of Formula (I). It is understood that the individual
stereoisomers as well as mixtures (racemic and non-racemic) of
stereoisomers are encompassed by the scope of the present
invention.
[0084] The compounds of the invention are useful in a free base
form, in the form of pharmaceutically acceptable salts,
pharmaceutically acceptable hydrates, pharmaceutically acceptable
esters, pharmaceutically acceptable solvates, pharmaceutically
acceptable prodrugs, pharmaceutically acceptable metabolites, and
in the form of pharmaceutically acceptable stereoisomers. These
forms are all within the scope of the disclosure.
[0085] "Pharmaceutically acceptable salt", "hydrate", "ester" or
"solvate" refers to a salt, hydrate, ester, or solvate of the
inventive compounds which possesses the desired pharmacological
activity and which is neither biologically nor otherwise
undesirable. Organic acids can be used to produce salts, hydrates,
esters, or solvates such as acetate, adipate, alginate, aspartate,
benzoate, benzenesulfonate, p-toluenesulfonate, bisulfate,
sulfamate, sulfate, naphthylate, butyrate, citrate, camphorate,
camphorsulfonate, cyclopentane-propionate, digluconate,
dodecylsulfate, ethanesulfonate, fumarate, glucoheptanoate,
glycerophosphate, hemisulfate heptanoate, hexanoate,
2-hydroxyethanesulfonate, lactate, maleate, methanesulfonate,
2-naphthalenesulfonate, nicotinate, oxalate, tosylate and
undecanoate. Inorganic acids can be used to produce salts,
hydrates, esters, or solvates such as hydrochloride, hydrobromide,
hydroiodide, and thiocyanate.
[0086] Examples of suitable base salts, hydrates, esters, or
solvates include hydroxides, carbonates, and bicarbonates of
ammonia, alkali metal salts such as sodium, lithium and potassium
salts, alkaline earth metal salts such as calcium and magnesium
salts, aluminum salts, and zinc salts.
[0087] Salts, hydrates, esters, or solvates may also be formed with
organic bases. Organic bases suitable for the formation of
pharmaceutically acceptable base addition salts, hydrates, esters,
or solvates of the compounds of the present invention include those
that are non-toxic and strong enough to form such salts, hydrates,
esters, or solvates. For purposes of illustration, the class of
such organic bases may include mono-, di-, and trialkylamines, such
as methylamine, dimethylamine, triethylamine and dicyclohexylamine;
mono-, di- or trihydroxyalkylamines, such as mono-, di-, and
triethanolamine; amino acids, such as arginine and lysine;
guanidine; N-methyl-glucosamine; N-methyl-glucamine; L-glutamine;
N-methyl-piperazine; morpholine; ethylenediamine;
N-benzyl-phenethylamine; (trihydroxy-methyl)aminoethane; and the
like. See, for example, "Pharmaceutical Salts," J. Pharm. Sci.,
66:1, 1-19 (1977). Accordingly, basic nitrogen-containing groups
can be quaternized with agents including: lower alkyl halides such
as methyl, ethyl, propyl, and butyl chlorides, bromides and
iodides; dialkyl sulfates such as dimethyl, diethyl, dibutyl and
diamyl sulfates; long chain halides such as decyl, lauryl, myristyl
and stearyl chlorides, bromides and iodides; and aralkyl halides
such as benzyl and phenethyl bromides.
[0088] The acid addition salts, hydrates, esters, or solvates of
the basic compounds may be prepared either by dissolving the free
base of a compound of the present invention in an aqueous or an
aqueous alcohol solution or other suitable solvent containing the
appropriate acid or base, and isolating the salt by evaporating the
solution. Alternatively, the free base of a compound of the present
invention can be reacted with an acid, as well as reacting a
compound of the present invention having an acid group thereon with
a base, such that the reactions are in an organic solvent, in which
case the salt separates directly or can be obtained by
concentrating the solution.
[0089] "Pharmaceutically acceptable prodrug" refers to a derivative
of the inventive compounds which undergoes biotransformation prior
to exhibiting its pharmacological effect(s). The prodrug is
formulated with the objective(s) of improved chemical stability,
improved patient acceptance and compliance, improved
bioavailability, prolonged duration of action, improved organ
selectivity, improved formulation (e.g., increased
hydrosolubility), and/or decreased side effects (e.g., toxicity).
The prodrug can be readily prepared from the inventive compounds
using methods known in the art, such as those described by Burgers
Medicinal Chemistry and Drug Chemistry, Fifth Ed, Vol. 1, pp.
172-178, 949-982 (1995). For example, the inventive compounds can
be transformed into prodrugs by converting one or more of the
hydroxy or carboxy groups into esters.
[0090] "Pharmaceutically acceptable metabolite" refers to drugs
that have undergone a metabolic transformation. After entry into
the body, most drugs are substrates for chemical reactions that may
change their physical properties and biologic effects. These
metabolic conversions, which usually affect the polarity of the
compound, alter the way in which drugs are distributed in and
excreted from the body. However, in some cases, metabolism of a
drug is required for therapeutic effect. For example, anticancer
drugs of the antimetabolite class must be converted to their active
forms after they have been transported into a cancer cell. Since
most drugs undergo metabolic transformation of some kind, the
biochemical reactions that play a role in drug metabolism may be
numerous and diverse. The main site of drug metabolism is the
liver, although other tissues may also participate.
[0091] Further still, the methods of the invention can be used to
treat cancer and to chemo sensitize and radiosensitize cancer
and/or tumor cells. The term "cancer," as used herein, is defined
broadly. The compounds of the present invention can potentiate the
effects of "anti-cancer agents," which term also encompasses
"anti-tumor cell growth agents," "chemotherapeutic agents,"
"cytostatic agents," "cytotoxic agents," and "anti-neoplastic
agents". The term "BRCA1/2-associated breast cancer" encompasses
breast cancer in which the breast cancer cells are deficient in the
breast cancer tumor suppressor genes BRCA1 and/or BRCA2.
[0092] For example, the methods of the invention are useful for
treating cancers such as ACTH-producing tumors, acute lymphocytic
leukemia, acute nonlymphocytic leukemia, cancer of the adrenal
cortex, bladder cancer, brain cancer, breast cancer, cervical
cancer, chronic lymphocytic leukemia, chronic myelocytic leukemia,
colorectal cancer, cutaneous T-cell lymphoma, endometrial cancer,
esophageal cancer, Ewing's sarcoma, gallbladder cancer, hairy cell
leukemia, head and neck cancer, Hodgkin's lymphoma, Kaposi's
sarcoma, kidney cancer, liver cancer, lung cancer (small and/or
non-small cell), malignant peritoneal effusion, malignant pleural
effusion, melanoma, mesothelioma, multiple myeloma, neuroblastoma,
non-Hodgkin's lymphoma, osteosarcoma, ovarian cancer, ovary (germ
cell) cancer, prostate cancer, pancreatic cancer, penile cancer,
retinoblastoma, skin cancer, soft-tissue sarcoma, squamous cell
carcinomas, stomach cancer, testicular cancer, thyroid cancer,
trophoblastic neoplasms, uterine cancer, vaginal cancer, cancer of
the vulva and Wilm's tumor.
[0093] In some non-limiting embodiments, the cancer and/or tumor
cells are selected from the group consisting of brain cancer,
melanoma, head and neck cancer, non small cell lung cancer,
testicular cancer, ovarian cancer, colon cancer and rectal
cancer.
[0094] The present invention also relates to a pharmaceutical
composition comprising (i) a therapeutically effective amount of a
compound of a compound of Formula (I) and (ii) a pharmaceutically
acceptable carrier.
[0095] The above discussion relating to the preferred embodiments'
utility and administration of the compounds of the present
invention also applies to the pharmaceutical composition of the
present invention.
[0096] The term "pharmaceutically acceptable carrier" as used
herein refers to any carrier, diluent, excipient, suspending agent,
lubricating agent, adjuvant, vehicle, delivery system, emulsifier,
disintegrant, absorbent, preservative, surfactant, colorant,
flavorant, or sweetener.
[0097] For these purposes, the composition of the invention may be
administered orally, parenterally, by inhalation spray, adsorption,
absorption, topically, rectally, nasally, bucally, vaginally,
intraventricularly, via an implanted reservoir in dosage
formulations containing conventional non-toxic
pharmaceutically-acceptable carriers, or by any other convenient
dosage form. The term parenteral as used herein includes
subcutaneous, intravenous, intramuscular, intraperitoneal,
intrathecal intraventricular, intrasternal, and intracranial
injection or infusion techniques.
[0098] When administered parenterally, the composition will
normally be in a unit dosage, sterile injectable form (solution,
suspension or emulsion) which is preferably isotonic with the blood
of the recipient with a pharmaceutically acceptable carrier.
Examples of such sterile injectable forms are sterile injectable
aqueous or oleaginous suspensions. These suspensions may be
formulated according to techniques known in the art using suitable
dispersing or wetting agents and suspending agents. The sterile
injectable forms may also be sterile injectable solutions or
suspensions in non-toxic parenterally-acceptable diluents or
solvents, for example, as solutions in 1,3-butanediol. Among the
acceptable vehicles and solvents that may be employed are water,
saline, Ringer's solution, dextrose solution, isotonic sodium
chloride solution, and Hanks' solution. In addition, sterile, fixed
oils are conventionally employed as solvents or suspending mediums.
For this purpose, any bland fixed oil may be employed including
synthetic mono- or di-glycerides, corn, cottonseed, peanut, and
sesame oil. Fatty acids such as ethyl oleate, isopropyl myristate,
and oleic acid and its glyceride derivatives, including olive oil
and castor oil, especially in their polyoxyethylated versions, are
useful in the preparation of injectables. These oil solutions or
suspensions may also contain long-chain alcohol diluents or
dispersants.
[0099] Sterile saline is a preferred carrier, and the compounds are
often sufficiently water soluble to be made up as a solution. The
carrier may contain minor amounts of additives, such as substances
that enhance solubility, isotonicity, and chemical stability, e.g.,
anti-oxidants, buffers and preservatives.
[0100] Formulations suitable for nasal or buccal administration
(such as self-propelling powder dispensing formulations) may
comprise about 0.1% to about 5% w/w, for example 1% w/w of active
ingredient. The formulations for human medical use of the present
invention comprise an active ingredient in association with a
pharmaceutically acceptable carrier therefore and optionally other
therapeutic ingredient(s).
[0101] When administered orally, the composition will usually be
formulated into unit dosage forms such as tablets, cachets, powder,
granules, beads, chewable lozenges, capsules, liquids, aqueous
suspensions or solutions, or similar dosage forms, using
conventional equipment and techniques known in the art. Such
formulations typically include a solid, semisolid, or liquid
carrier. Exemplary carriers include lactose, dextrose, sucrose,
sorbitol, mannitol, starches, gum acacia, calcium phosphate,
mineral oil, cocoa butter, oil of theobroma, alginates, tragacanth,
gelatin, syrup, methyl cellulose, polyoxyethylene sorbitan
monolaurate, methyl hydroxybenzoate, propyl hydroxybenzoate, talc,
magnesium stearate, and the like.
[0102] The composition of the invention is preferably administered
as a capsule or tablet containing a single or divided dose of the
compound of Formula (I) or pharmaceutically acceptable salt
thereof. The composition may be administered as a sterile solution,
suspension, or emulsion, in a single or divided dose. Tablets may
contain carriers such as lactose and corn starch, and/or
lubricating agents such as magnesium stearate. Capsules may contain
diluents including lactose and dried corn starch.
[0103] A tablet may be made by compressing or molding the active
ingredient optionally with one or more accessory ingredients.
Compressed tablets may be prepared by compressing, in a suitable
machine, the active ingredient in a free-flowing form such as a
powder or granules, optionally mixed with a binder, lubricant,
inert diluent, surface active, or dispersing agent Molded tablets
may be made by molding in a suitable machine, a mixture of the
powdered active ingredient and a suitable carrier moistened with an
inert liquid diluent.
[0104] The compounds of this invention may also be administered
rectally in the form of suppositories. These compositions can be
prepared by mixing the drug with a suitable non-irritating
excipient which is solid at room temperature, but liquid at rectal
temperature, and, therefore, will melt in the rectum to release the
drug. Such materials include cocoa butter, beeswax, and
polyethylene glycols.
[0105] Compositions and methods of the invention also may utilize
controlled release technology. Thus, for example, the disclosed
compounds may be incorporated into a hydrophobic polymer matrix for
controlled release over a period of days. The composition of the
invention may then be molded into a solid implant, or externally
applied patch, suitable for providing efficacious concentrations of
the PARP inhibitors over a prolonged period of time without the
need for frequent re-dosing. Such controlled release films are well
known to the art. Particularly preferred are transdermal delivery
systems. Other examples of polymers commonly employed for this
purpose that may be used in the present invention include
nondegradable ethylene-vinyl acetate copolymer a degradable lactic
acid-glycolic acid copolymers which may be used externally or
internally. Certain hydrogels such as
poly(hydroxyethylmethacrylate) or poly(vinylalcohol) also may be
useful, but for shorter release cycles than the other polymer
release systems, such as those mentioned above.
[0106] In an embodiment, the carrier is a solid biodegradable
polymer or mixture of biodegradable polymers with appropriate time
release characteristics and release kinetics. The composition of
the invention may then be molded into a solid implant suitable for
providing efficacious concentrations of the compounds of the
invention over a prolonged period of time without the need for
frequent re-dosing. The composition of the present invention can be
incorporated into the biodegradable polymer or polymer mixture in
any suitable manner known to one of ordinary skill in the art and
may form a homogeneous matrix with the biodegradable polymer, or
may be encapsulated in some way within the polymer, or may be
molded into a solid implant.
[0107] In one embodiment, the biodegradable polymer or polymer
mixture is used to form a soft "depot" containing the
pharmaceutical composition of the present invention that can be
administered as a flowable liquid, for example, by injection, but
which remains sufficiently viscous to maintain the pharmaceutical
composition within the localized area around the injection site.
The degradation time of the depot so formed can be varied from
several days to a few years, depending upon the polymer selected
and its molecular weight. By using a polymer composition in
injectable form, even the need to make an incision may be
eliminated. In any event, a flexible or flowable delivery "depot"
will adjust to the shape of the space it occupies with the body
with a minimum of trauma to surrounding tissues. The pharmaceutical
composition of the present invention is used in amounts that are
therapeutically effective, and may depend upon the desired release
profile, the concentration of the pharmaceutical composition
required for the sensitizing effect, and the length of time that
the pharmaceutical composition has to be released for
treatment.
[0108] The compounds of the invention are used in the composition
in amounts that are therapeutically effective. The compositions may
be sterilized and/or contain adjuvants, such as preserving,
stabilizing, welling, or emulsifying agents, solution promoters,
salts for regulating the osmotic pressure, and/or buffers. In
addition, they may also contain other therapeutically valuable
substances, such as, without limitation, the specific
chemotherapeutic agents recited herein. The compositions are
prepared according to conventional mixing, granulating, or coating
methods, and contain about 0.1 to 75% by weight, preferably about 1
to 50% by weight, of the compound of the invention.
[0109] To be effective therapeutically as central nervous system
targets, the compounds of the present invention should readily
penetrate the blood-brain barrier when peripherally administered.
Compounds which cannot penetrate the blood-brain barrier can be
effectively administered by an intraventricular route or other
appropriate delivery system suitable for administration to the
brain.
[0110] For medical use, the amount required of the active
ingredient to achieve a therapeutic effect will vary with the
particular compound, the route of administration, the mammal under
treatment, and the particular disorder or disease being treated. A
suitable systematic dose of a compound of the present invention or
a pharmacologically acceptable salt thereof for a mammal suffering
from, or likely to suffer from, any of condition as described
hereinbefore is in the range of about 0.1 mg/kg to about 100 mg/kg
of the active ingredient compound, the typical dosage being about 1
to about 10 mg/kg.
[0111] It is understood, however, that a specific dose level for
any particular patient will depend upon a variety of factors
including the activity of the specific compound employed, the age,
body weight, general health, sex, diet, time of administration,
rate of excretion, drug combination, and the severity of the
particular disease being treated and form of administration.
[0112] It is understood that the ordinarily skilled physician or
veterinarian will readily determine and prescribe the effective
amount of the compound for prophylactic or therapeutic treatment of
the condition for which treatment is administered. In so
proceeding, the physician or veterinarian can, for example, employ
an intravenous bolus followed by an intravenous infusion and
repeated administrations, parenterally or orally, as considered
appropriate. While it is possible for an active ingredient to be
administered alone, it is preferable to present it as a
formulation.
[0113] When preparing dosage form incorporating the compositions of
the invention, the compounds may also be blended with conventional
excipients such as binders, including gelatin, pregelatinized
starch, and the like; lubricants, such as hydrogenated vegetable
oil, stearic acid, and the like; diluents, such as lactose,
mannose, and sucrose; disintegrants, such as carboxymethylcellulose
and sodium starch glycolate; suspending agents, such as povidone,
polyvinyl alcohol, and the like; absorbants, such as silicon
dioxide; preservatives, such as methylparaben, propylparaben, and
sodium benzoate; surfactants, such as sodium lauryl sulfate,
polysorbate 80, and the like; colorants; flavorants; and
sweeteners. Pharmaceutically acceptable excipients are well known
in the pharmaceutical arts and are described, for example, in
Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton,
Pa. (e.g., 20.sup.th Ed., 2000), and Handbook of Pharmaceutical
Excipients, American Pharmaceutical Association, Washington, D.C.,
(e.g., 1.sup.st, 2.sup.nd and 3.sup.rd Eds., 1986, 1994, and 2000,
respectively).
[0114] The present invention relates to the use of a compound of
Formula (I) in the preparation of a medicament for the treatment of
any disease or disorder in an animal described herein. In an
embodiment, the compounds of the present invention are used to
treat cancer. In a preferred embodiment, the compounds of the
present invention are used to potentiate the cytotoxic effects of
ionizing radiation. In such an embodiment, the compounds of the
present invention act as a radiosensitizer. In an alternative
preferred embodiment, the compounds of the present invention are
used to potentiate the cytotoxic effects of chemotherapeutic
agents. In such an embodiment, the compounds of the present
invention act as a chemo sensitizer. In another preferred
embodiment, the compounds of the present invention are used to
inhibit the growth of cells having defects in the homologous
recombination (HR) pathway of double-stranded DNA repair.
[0115] Any pharmacologically-acceptable chemotherapeutic agent that
acts by damaging DNA is suitable as the chemotherapeutic agent of
the present invention. In particular, the present invention
contemplates the use of a chemotherapeutically effective amount of
at least one chemotherapeutic agent including, but not limited to:
temozolomide, adriamycin, camptothecin, carboplatin, cisplatin,
daunorubicin, docetaxel, doxorubicin, interferon-alpha,
interferon-beta, interferon-gamma, interleukin 2, irinotecan,
paclitaxel, topotecan, a taxoid, dactinomycin, danorubicin,
4'-deoxydoxorubicin, bleomycin, pilcamycin, mitomycin, neomycin,
gentamycin, etoposide 4-OH cyclophosphamide, a platinum
coordination complex, topotecan, and mixtures thereof. According to
a preferred aspect, the chemotherapeutic agent is temozolomide.
[0116] The invention contained herein demonstrates the usefulness
of the compounds and compositions of the present invention in
treating and/or preventing cancer, such as by radiosensitizing
and/or chemosensitizing tumor and/or cancer cells to
chemotherapeutic agents, and to inhibit the growth of cells having
defects in the homologous recombination (HR) pathway of
double-stranded DNA repair.
[0117] The following examples are for illustrative purposes only
and are not intended to limit the scope of the application.
[0118] In one embodiment, the present invention provides a tetraaza
phenalen-3-one compound of Formula (I), or a pharmaceutically
acceptable salt thereof
##STR00012##
wherein R is (a) NR.sup.1R.sup.2, wherein R.sup.1 is selected from
the group consisting of hydrogen, C.sub.1-C.sub.6 straight or
branched chain alkyl, C.sub.2-C.sub.6 straight or branched chain
alkenyl, C.sub.3-C.sub.8 cycloalkyl, C.sub.1-C.sub.6 alkoxy,
C.sub.2-C.sub.6 alkenyloxy, phenyl, phenoxy, benzyloxy,
NR.sup.AR.sup.B (C.sub.1-C.sub.6 straight or branched chain alkyl),
NR.sup.AR.sup.B (C.sub.2-C.sub.6 straight or branched chain
alkenyl), (C.sub.1-C.sub.6 straight or branched chain
alkyl)carbonyl, (C.sub.2-C.sub.6 straight or branched chain
alkenyl)carbonyl, (C.sub.3-C.sub.8 cycloalkyl)carbonyl,
(C.sub.1-C.sub.6 straight or branched chain alkyl)oxycarbonyl,
(C.sub.2-C.sub.6 straight or branched chain alkenyl)oxycarbonyl,
(C.sub.3-C.sub.8 cycloalkyl)oxycarbonyl, arylcarbonyl, sulfonyl,
arylsulfonyl, aryl(C.sub.1-C.sub.6 straight or branched chain
alkyl), aryl(C.sub.2-C.sub.6 straight or branched chain alkenyl),
aryl(C.sub.3-C.sub.8 cycloalkyl), (C.sub.1-C.sub.6 straight or
branched chain alkyl)aryl, (C.sub.2-C.sub.6 straight or branched
chain alkenyl)aryl, (C.sub.3-C.sub.8 cycloalkyl)aryl, aryl,
heterocyclyl, heterocyclyl(C.sub.1-C.sub.6 straight or branched
chain alkyl), and heterocyclyl(C.sub.2-C.sub.6 straight or branched
chain alkenyl); wherein each heterocyclyl has between 1 and 7
heteroatoms independently selected from O, N, or S, and wherein
each of R.sup.A and R.sup.B are independently selected from the
group consisting of hydrogen, C.sub.1-C.sub.6 straight or branched
chain alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl,
and C.sub.3-C.sub.8 cycloalkyl; and R.sup.2 is selected from the
group consisting of C.sub.1-C.sub.6 straight or branched chain
alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl,
C.sub.3-C.sub.8 cycloalkyl, C.sub.1-C.sub.6 alkoxy, C.sub.2-C.sub.6
alkenyloxy, phenyl, phenoxy, benzyloxy, NR.sup.XR.sup.Y
(C.sub.1-C.sub.6 straight or branched chain alkyl), NR.sup.XR.sup.Y
(C.sub.2-C.sub.6 straight or branched chain alkenyl),
(C.sub.1-C.sub.6 straight or branched chain alkyl)carbonyl,
(C.sub.2-C.sub.6 straight or branched chain alkenyl)carbonyl,
(C.sub.3-C.sub.8 cycloalkyl)carbonyl, (C.sub.1-C.sub.6 straight or
branched chain alkyl)oxycarbonyl, (C.sub.2-C.sub.6 straight or
branched chain alkenyl)oxycarbonyl, (C.sub.3-C.sub.8
cycloalkyl)oxycarbonyl, arylcarbonyl, sulfonyl, arylsulfonyl,
aryl(C.sub.1-C.sub.6 straight or branched chain alkyl),
aryl(C.sub.2-C.sub.6 straight or branched chain alkenyl),
aryl(C.sub.3-C.sub.8 cycloalkyl), (C.sub.1-C.sub.6 straight or
branched chain alkyl)aryl, (C.sub.2-C.sub.6 straight or branched
chain alkenyl)aryl, (C.sub.3-C.sub.8 cycloalkyl)aryl, aryl,
heterocyclyl, heterocyclyl(C.sub.1-C.sub.6 straight or branched
chain alkyl), and heterocyclyl(C.sub.2-C.sub.6 straight or branched
chain alkenyl); wherein each heterocyclyl has between 1 and 7
heteroatoms independently selected from O, N, or S, and wherein
each of R.sup.X and R.sup.Y are independently selected from the
group consisting of hydrogen, C.sub.1-C.sub.6 straight or branched
chain alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl,
and C.sub.3-C.sub.8 cycloalkyl; wherein R.sup.1 and R.sup.2 are
independently substituted with between 0 and 4 substituents, each
independently selected from halo, C.sub.1-C.sub.6 straight or
branched chain alkyl, C.sub.2-C.sub.6 straight or branched chain
alkenyl, C.sub.1-C.sub.6 alkoxy, trifluoromethyl, trifluoroethyl,
and amino; and provided that R.sup.1 and R.sup.2 may not both be
methyl, and R.sup.2 may not be (phenyl)prop-1-yl when R.sup.1 is
hydrogen; or (b) aryloxy, substituted with between 0 and 4
substituents, each independently selected from the group consisting
of halo, C.sub.1-C.sub.6 alkoxy, trifluoromethyl, trifluoroethyl,
C.sub.1-C.sub.6 straight or branched chain alkyl, C.sub.2-C.sub.6
straight or branched chain alkenyl, C.sub.3-C.sub.8 cycloalkyl,
NR.sup.CR.sup.D, NR.sup.CR.sup.D(C.sub.1-C.sub.6 straight or
branched chain alkyl), and NR.sup.CR.sup.D(C.sub.2-C.sub.6 straight
or branched chain alkenyl), wherein each of R.sup.C and R.sup.D is
independently selected from the group consisting of hydrogen,
C.sub.1-C.sub.6 straight or branched chain alkyl, C.sub.2-C.sub.6
straight or branched chain alkenyl, and C.sub.3-C.sub.8 cycloalkyl;
and when more than one substituent is of the form NR.sup.CR.sup.D,
each occurrence of R.sup.C and R.sup.D is independently selected
from the group consisting of hydrogen, C.sub.1-C.sub.6 straight or
branched chain alkyl, C.sub.2-C.sub.6 straight or branched chain
alkenyl, and C.sub.3-C.sub.8 cycloalkyl; or (c) a heterocyclyl
having between 1 and 7 heteroatoms independently selected from O,
N, or S; and having between 0 and 4 substituents independently
selected from the group consisting of halo, haloalkyl, hydroxyl,
nitro, trifluoromethyl, trifluoroethyl, C.sub.1-C.sub.6 straight or
branched chain alkyl, C.sub.2-C.sub.6 straight or branched chain
alkenyl, C.sub.1-C.sub.6 alkoxy, C.sub.2-C.sub.6 alkenyloxy,
phenyl, phenoxy, benzyloxy, amino, thiocarbonyl, cyano, imino,
NR.sup.ER.sup.F(C.sub.1-C.sub.6 straight or branched chain alkyl),
NR.sup.ER.sup.F (C.sub.1-C.sub.6 straight or branched chain
alkenyl) sulfhydryl, thioalkyl, dioxa-spiroethyl, (C.sub.1-C.sub.6
straight or branched chain alkyl) carbonyl, (C.sub.2-C.sub.6
straight or branched chain alkenyl)carbonyl, (C.sub.1-C.sub.6
straight or branched chain alkyl)oxycarbonyl, (C.sub.2-C.sub.6
straight or branched chain alkenyl)oxycarbonyl, arylcarbonyl,
sulfonyl, arylsulfonyl, aryl(C.sub.1-C.sub.6 straight or branched
chain alkyl), aryl(C.sub.2-C.sub.6 straight or branched chain
alkenyl), aryl(C.sub.3-C.sub.8 cycloalkyl), (C.sub.1-C.sub.6
straight or branched chain alkyl)aryl, (C.sub.2-C.sub.6 straight or
branched chain alkenyl)aryl, (C.sub.3-C.sub.8 cycloalkyl)aryl,
aryl, heterocyclyl, heterocyclyl(C.sub.1-C.sub.6 straight or
branched chain alkyl), and heterocyclyl(C.sub.2-C.sub.6 straight or
branched chain alkenyl), wherein each heterocyclyl has between 1
and 7 heteroatoms independently selected from O, N, or S, wherein
each of R.sup.E and R.sup.E is independently selected from the
group consisting of hydrogen, C.sub.1-C.sub.6 straight or branched
chain alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl,
and C.sub.3-C.sub.8 cycloalkyl; and when more than one substituent
is of the form NR.sup.ER.sup.F each occurrence of R.sup.E and
R.sup.F is independently selected from the group consisting of
hydrogen, C.sub.1-C.sub.6 straight or branched chain alkyl,
C.sub.2-C.sub.6 straight or branched chain alkenyl, and
C.sub.3-C.sub.8 cycloalkyl; wherein each of said 0-4 substituents
is independently substituted with between 0 and 4 further
substituents, and each said further substituent is independently
selected from halo, C.sub.1-C.sub.6 straight or branched chain
alkyl, C.sub.2-C.sub.6 straight or branched chain alkenyl,
C.sub.3-C.sub.8 cycloalkyl, C.sub.1-C.sub.6 alkoxy,
trifluoromethyl, trifluoroethyl, and amino; provided that R has at
least one substituent when R is an N-piperidinyl, N-pyrrolidinyl or
an N-morpholinyl group.
[0119] In some embodiments each ring of each heterocycle of Formula
(I) is independently 5-7 atoms in size.
[0120] Some embodiments include one, two or three nitrogen atoms in
at least one ring of the heterocycle of Formula (I).
[0121] In some embodiments, the heterocyclyl of Formula(I)
comprises 1-3 rings. In some embodiments, the heterocyclyl has 1-7
heteroatoms independently selected from 0, N, and S.
[0122] In some embodiments, the heterocyclyl of Formula (I) is
selected from the group consisting of piperidinyl, piperazinyl,
pyridazinyl, dihydropyridyl, tetrahydropyridyl, pyridinyl,
pyrimidinyl, dihydropyrimidinyl, tetrahydrophyrimidinyl,
hexahydropyrimidinyl, dihydropyrazinyl, tetrahydropyrazinyl,
pyrrolidinyl, imidazolidinyl, pyrazolidinyl, pyrrolyl,
dihydropyrolyl, imidazolyl, dihydroimidazoyl, pyrazolyl,
dihydropyrazolyl, azepanyl, [1,2]diazepanyl, [1,3]diazepanyl,
[1,4]diazepanyl, indolyl, dihydroindolyl, isoindolyl,
dihydroisoindoly, dihydroquinolyl, tetrahydroquinolyl,
dihydroisoquinolyl, and tetrahydroisoquinolyl.
[0123] In another embodiment, the present invention provides a
compound selected from the group consisting of
##STR00013## ##STR00014## ##STR00015## ##STR00016## ##STR00017##
##STR00018## ##STR00019## ##STR00020## ##STR00021##
and pharmaceutically acceptable salts thereof.
[0124] In some embodiments the invention provides the compound
which is
##STR00022##
or a pharmaceutically acceptable salt thereof.
[0125] In some embodiments the invention provides the compound
which is
##STR00023##
or a pharmaceutically acceptable salt thereof.
[0126] In some embodiments the present invention provides a method
of chemo sensitizing cancer cells in a mammal in need of
chemotherapy, comprising administering to said mammal a compound of
Formula (I) as described herein, or a pharmaceutically acceptable
salt thereof. In some embodiments, said mammal is a human. In some
embodiments, said administration is administration of a
pharmaceutical composition comprising said compound and a
pharmaceutically acceptable carrier. In some embodiments, the chemo
sensitization method further comprises administering to said mammal
a chemotherapeutic agent. In some embodiments, said chemo
sensitizing compound and said chemotherapeutic agent are
administered essentially simultaneously.
[0127] In some embodiments the present invention provides a method
of chemo sensitizing cancer cells in a mammal in need of
chemotherapy, comprising administering to said mammal a compound
selected from the group consisting of compounds 7-28, 30-46, 50-66,
69, 72, 74-76, and pharmaceutically acceptable salts thereof, as
described herein. In some embodiments, said mammal is a human. In
some embodiments, said administration is administration of a
pharmaceutical composition comprising said compound and a
pharmaceutically acceptable carrier. In some embodiments, the chemo
sensitization method further comprises administering to said mammal
a chemotherapeutic agent. In some embodiments, said chemo
sensitizing compound and said chemotherapeutic agent are
administered essentially simultaneously.
[0128] In some embodiments, the chemotherapeutic agent of the
invention is selected is selected from the group consisting of
temozolomide, adriamycin, camptothecin, carboplatin, cisplatin,
daunorubicin, docetaxel, doxorubicin, interferon-alpha,
interferon-beta, interferon-gamma, interleukin 2, irinotecan,
paclitaxel, topotecan, a taxoid, dactinomycin, danorubicin,
4'-deoxydoxorubicindeoxydoxorubicin, bleomycin, pilcamycin,
mitomycin, neomycin, gentamycin, etoposide, 4-OH cyclophosphamide,
a platinum coordination complex, and mixtures thereof. In some
embodiments, the chemotherapeutic agent is temozolomide or a salt
thereof.
[0129] In some embodiments, the present invention provides a method
of radiosensitizing cancer cells in a mammal in need of radiation
therapy comprising administering to said mammal a compound of
Formula (I) as described herein, or a pharmaceutically acceptable
salt thereof. In some embodiments, said mammal is a human. In some
embodiments, said administration is administration of a
pharmaceutical composition comprising said compound and a
pharmaceutically acceptable carrier.
[0130] In some embodiments, the present invention provides a method
of radiosensitizing cancer cells in a mammal in need of radiation
therapy comprising administering to said mammal a compound selected
from the group consisting of compounds 7-28, 30-46, 50-66, 69, 72,
74-76, and pharmaceutically acceptable salts thereof, as described
herein. In some embodiments, said mammal is a human. In some
embodiments, said administration is administration of a
pharmaceutical composition comprising said compound and a
pharmaceutically acceptable carrier.
[0131] In some embodiments, the invention provides a pharmaceutical
composition comprising a compound of Formula (I) as described
herein, or a pharmaceutically acceptable salt thereof, and a
pharmaceutically acceptable carrier. In some embodiments, the
pharmaceutical composition further comprises a chemotherapeutic
agent as described herein.
[0132] In some embodiments, the invention provides a pharmaceutical
composition comprising compound selected from the group consisting
of compounds 7-28, 30-46, 50-66, 69, 72, 74-76, and
pharmaceutically acceptable salts thereof, as described herein. In
some embodiments, the pharmaceutical composition further comprises
a chemotherapeutic agent as described herein.
[0133] In some embodiments, the cancer cells treated by the chemo
sensitizing and/or radiosensitizing methods of the invention are
selected from the group consisting of ACTH-producing tumors, acute
lymphocytic leukemia, acute nonlymphocytic leukemia, cancer of the
adrenal cortex, bladder cancer, brain cancer, breast cancer,
cervical cancer, chronic lymphocytic leukemia, chronic myelocytic
leukemia, colorectal cancer, cutaneous T-cell lymphoma, endometrial
cancer, esophageal cancer, Ewing's sarcoma, gallbladder cancer,
hairy cell leukemia, head and neck cancer, Hodgkin's lymphoma,
Kaposi's sarcoma, kidney cancer, liver cancer, lung cancer (small
and/or non-small cell), malignant peritoneal effusion, malignant
pleural effusion, melanoma, mesothelioma, multiple myeloma,
neuroblastoma, non-Hodgkin's lymphoma, osteosarcoma, ovarian
cancer, ovary (germ cell) cancer, prostate cancer, pancreatic
cancer, penile cancer, retinoblastoma, skin cancer, soft-tissue
sarcoma, squamous cell carcinomas, stomach cancer, testicular
cancer, thyroid cancer, trophoblastic neoplasms, uterine cancer,
vaginal cancer, cancer of the vulva and Wilm's tumor. In some
embodiments, the cancer cells treated by the chemo sensitizing
and/or radiosensitizing methods of the invention are selected from
the group consisting of brain cancer, melanoma, head and neck
cancer, testicular cancer, ovarian cancer, breast cancer, non small
cell lung cancer, and rectal cancer.
[0134] In some embodiments, the invention provides a method of
treating a mammal having a cancer characterized by having a defect
in the homologous recombination (HR) pathway of double-stranded DNA
repair, comprising administering to said mammal a compound of
Formula (I) as described herein, or a pharmaceutically acceptable
salt thereof. In some embodiments, said mammal is a human. In some
embodiments, said administration is administration of a
pharmaceutical composition comprising said compound and a
pharmaceutically acceptable carrier. In some embodiments, the
cancer cells have a phenotype selected from the group consisting of
i) a BRCA-1 defect, ii) a BRCA-2 defect, iii) a BRCA-1 and BRCA-2
defect, and iv) Fanconi anemia. In some embodiments, the cancer
cells are selected from breast cancer or ovarian cancer.
[0135] In some embodiments, the invention provides a method of
treating a mammal having a cancer characterized by having a defect
in the homologous recombination (HR) pathway of double-stranded DNA
repair, comprising administering to said mammal a compound selected
from the group consisting of compounds 7-28, 30-46, 50-66, 69, 72,
74-76, and pharmaceutically acceptable salts thereof, as described
herein. In some embodiments, said mammal is a human. In some
embodiments, said administration is administration of a
pharmaceutical composition comprising said compound and a
pharmaceutically acceptable carrier. In some embodiments, the
cancer cells have a phenotype selected from the group consisting of
i) a BRCA-1 defect, ii) a BRCA-2 defect, iii) a BRCA-1 and BRCA-2
defect, and iv) Fanconi anemia. In some embodiments, the cancer
cells are selected from breast cancer or ovarian cancer.
Synthetic Procedures for the Disclosed Compounds
##STR00024##
[0136] Procedure A: Preparation of 3-Nitro-Phthalic Acid Dimethyl
Ester, 2
[0137] To a stirred solution of 4-nitro-isobenzofuran-1,3-dione
(150 g, 0.78 mol), 1, in 2 L of MeOH was added 50 mL of
concentrated sulfuric acid. The reaction was heated to reflux for
16 hours. The mixture solution was cooled to room temperature and
then poured into 3 L of ice water and resulted in a heavy white
precipitate. This was triturated for 15 minutes and the
precipitated was filtered off and the solid was washed with water
thoroughly and dried to afford 120 g of 3-nitro-phthalic acid
dimethyl ester, 2, as a white solid (65%). .sup.1H NMR (300 MHz,
DMSO-d.sub.6): 8.54 (d, J=7.25 Hz, 1H), 8.42 (d, J=7.82 Hz, 1H),
7.98 (t, J=8.20 Hz, 1H), 3.99 (s, 3H), 3.98 (s, 3H). .sup.13C NMR:
52.03, 52.29, 111.02, 115.67, 119.08, 131.80, 133.68, 148.80,
167.64, 168.63.
Procedure B: Preparation of 3-Amino-Phthalic Acid Dimethyl Ester,
3
[0138] The compound 2 (205 g, 1.0 mol) was dissolved in 2 L of
MeOH. Catalytic 10% Pd/C was added and the solution was
hydrogenated under H.sub.2 (45 psi) on a Parr hydrogenation
apparatus at room temperature overnight. Filtered through celite
and evaporated to give a quantitative yield of 3-amino-phthalic
acid dimethyl ester, 3. .sup.1H NMR (300 MHz, DMSO-d.sub.6): 7.26
(t, J=7.33 Hz, 1H), 6.94 (d, J=8.34 Hz, 1H), 6.77 (d, J=8.33 Hz,
1H), 6.12 (s, 2H), 3.77 (s, 3H), 3.76 (s, 3H). .sup.13C NMR: 51.51,
51.77, 110.50, 115.16, 118.56, 131.26, 133.16, 148.28, 167.12,
168.11.
Procedure C: Preparation of
2-Chloromethyl-4-Oxo-3,4-Dihydro-Quinazoline-5-Carboxylic Acid
Methyl Ester, 4:
[0139] 100 mL of chloroacetonitrile was set stirring in 130 mL of
1,4 dioxane at room temperature. Dry HCl gas was bubbled through
the solution for thirty minutes followed by the addition of 30 g of
3-amino-1,2-phthalic acid dimethyl ester, 3. The reaction was
refluxed for approximately three hours, resulting in a heavy white
precipitate. The suspension was cooled with an ice bath, filtered
and washed with pentane to remove any residual solvents. 30 g (83%)
of an analytically pure white solid, 4, was isolated. .sup.1H NMR
(300 MHz, DMSO-d.sub.6): 7.88 (t, J 8.33 Hz, 1H), 7.79 (d, J 7.08
Hz, 1H), 7.52 (d, J 7.33 Hz, 1H), 4.60 (s, 2H), 3.84 (s, 3H);
.sup.13C NMR: 42.21, 54.86, 119.95, 127.77, 130.86, 135.71, 136.78,
150.59, 155.70, 162.49, 171.24.
General Procedure D: Preparation of Compounds, 5:
[0140] Displacement of the chloro group of compound 4 with
nucleophiles such as amine using General procedure D provides the
compounds 5. To a solution of the chloro compound 4 in dry DMF or
MeCN is added potassium carbonate and a nucleophile such as an
amine. The reaction mixture is heated to 70.degree. C. for 12 hours
and cooled to room temperature. Water is added to the reaction
mixture, followed by ethyl acetate. The organic layer is collected,
washed with water, brine and dried over sodium sulfate. The
solvents are removed in vacuum. The residue is purified by column
chromatography on silica gel using ethyl acetate/hexanes as eluent
to give the products 5 in 50-95% yield. An example was given in the
preparation of compound 7.
General Procedure E: Preparation of Compounds, 6:
[0141] A 2,9-Dihydro-1,2,7,9-tetraaza-phenalen-3-one ring can be
formed by condensation of the compounds 6 with hydrazine. To a
solution of the compounds 6 in absolute ethanol is added excess
anhydrous hydrazine at room temperature. The solution is refluxed
for overnight and cooled to room temperature. Ice-cold water is
added and white solid is separated. The solid is collected by
vacuum filtration and washed with water and small amount of
methanol to give white solid products 6 in 40-90% yield. An example
was given in the preparation of compound 7.
Example 1
Preparation of
8-(4-hydroxy-piperidin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen--
3-one, 7
[0142] Following the General Procedure D: A solution of MeCN (25
ml), 4-hydroxypiperidine (0.46 mg, 4.5 mmol), 4 (1.0 g, 3.9 mmol),
and potassium carbonate (1 g, 7 mmol) was set refluxing under
nitrogen and stirred overnight. Reaction mixture was evaporated to
dryness and extracted with dichloromethane. Purified with a silica
column using 9:1 dichloromethane/MeOH to afford 1.05 g (84%) of an
off-white solid,
2-(4-Hydroxy-piperidin-1-ylmethyl)-4-oxo-3,4-dihydro-quinazoline-5-carbox-
ylic acid methyl ester, 7a.
[0143] Following the General Procedure E: To a solution of compound
7a (1.0 g, 3.1 mmol) in EtOH (20 mL) when refluxing was added
hydrazine monohydrate (7 mL, large excess) and heated overnight.
Reaction was cooled to RT and H.sub.2O (15 mL) was added resulting
in a heavy white precipitate. Filtered and washed with 1:1
EtOH/H.sub.2O to afford 0.6 g (64%) of an analytically pure white
solid, 7. MP: 168-171.degree. C.; MS (ES+): 300; .sup.1H NMR (300
MHz, CD.sub.3OD): 1.46-1.55 (m, 2H), 1.71-1.75 (m, 2H) 2.15-2.23
(m, 2H) 2.70-2.75 (m, 2H) 3.16-3.18 (m, 1H) 3.25 (s, 2H) 3.47-3.55
(m, 1H) 7.30-7.33 (m, 1H) 7.60-7.64 (m, 2H). Anal. Calcd. for
C.sub.15H.sub.17N.sub.5O.sub.2. 1.7H.sub.2O: C, 56.45; H, 6.06; N,
21.94. Found: C, 56.10; H, 6.00; N, 22.25.
[0144] The compound 7 can be formulated with an acid. For example:
to a solution of 7 (0.6 g, 2.0 mmol) in 10 mL of 1,4 dioxane/DMF
(9:1) at 90.degree. C. was added MsOH (0.14 mL, 2.1 mmol) resulting
in a heavy white precipitate. Filtered and triturated in diethyl
ether to afford 0.5 g (63%) of an off-white solid, mesylate salt of
7. .sup.1H NMR (300 MHz, DMSO-d.sub.6): 1.55-1.58 (m, 2H),
1.78-1.82 (m, 2H), 2.15 (s, 3H), 3.15-3.50 (m, 4H), 3.63-3.65 (m,
1H), 4.04 (s, 2H), 7.24 (d, J=8.5 Hz, 1H), 7.51-7.66 (m, 2H), 11.73
(s, 1H)
[0145] Anal. Calcd. for
C.sub.15H.sub.17N.sub.5O.sub.2.1CH.sub.3SO.sub.3H.2H.sub.2O: C,
44.54; H, 5.84; N, 16.23, S, 7.43. Found: C, 44.48; H, 5.76; N,
16.27, S, 7.60.
[0146] The following compounds were synthesized from the similar
procedures of preparation of compound 7, using the appropriate
corresponding amines.
Preparation of
8-(4-phenyl-piperazin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-
-one, 8
[0147] Synthesized using 1-phenylpiperazine for General Procedure
D. 52% overall yield for last two steps. MS (ES+): 361; .sup.1H NMR
(300 MHz, DMSO-d.sub.6): 2.65-2.68 (m, 4H), 3.19-3.22 (m, 4H) 3.39
(s, 2H); 6.78 (t, J 7.2 Hz, 1H); 6.95 (d, J 8.0 Hz, 2H), 7.19 (t,
J=7.2 Hz, 2H), 7.48-7.51 (m, 1H), 7.62-7.64 (d, J=7.2 Hz, 1H), 7.75
(t, J=8.0 Hz, 1H), 11.23 (s, br, 1H), 11.78 (s, 1H); Anal. Calcd.
for C.sub.20H.sub.20N.sub.6O.sub.1. 2.0H.sub.2O: C, 60.59; H, 6.10;
N, 21.20. Found: C, 60.48; H, 6.05; N, 21.35.
Preparation of
8-(4-benzyl-piperidin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-
-one, 9
[0148] Synthesized using 1-benzylpiperazine for General Procedure
D. 20% overall yield for last two steps. MS (ES-): 372; .sup.1H NMR
(300 MHz, DMSO-d.sub.6): 1.22-1.50 (m, 5H), 2.45-2.55 (m, 4H), 2.85
(d, 2H), 3.28 (s, 2H), 7.14-7.19 (m, 3H), 7.25-7.30 (m, 2H), 7.50
(d, J=7.0 Hz, 1H), 7.62 (d, J=7.7 Hz, 1H), 7.75 (t, J=7.7 Hz 1H),
11.25 (s, br, 1H), 11.76 (s, 1H); Anal. Calcd. for
C.sub.22H.sub.23N.sub.5O.sub.1: C, 70.76; H, 6.21; N, 18.75. Found:
C, 70.36; H, 6.18; N, 18.63.
Preparation of
8-phenoxymethyl-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-one, 10
[0149] Synthesized using phenol for General Procedure D. 60%
overall yield for last two steps. MS (ES+): 293; .sup.1H NMR (300
MHz, DMSO-d.sub.6): 4.90 (s, br, 3H), 7.00 (t, J=6.6 Hz, 1H), 7.08
(d, J=8.2 Hz, 2H), 7.34 (t, J=7.7 Hz, 2H), 7.45 (d, J=7.7 Hz, 1H),
7.65 (d, J=7.7 Hz, 1H), 7.76 (t, J=7.2 Hz, 1H), 11.20 (s, br, 1H),
11.80 (s, 1H). Anal. Calcd. for
C.sub.16H.sub.12N.sub.4O.sub.2.0.75H.sub.2O.0.25N.sub.2H.sub.4: C,
61.24; H, 4.66; N, 20.08. Found: C, 61.06; H, 4.27; N, 20.13.
Preparation of
8-[4-(4-fluoro-phenyl)-3,6-dihydro-2H-pyridin-1-ylmethyl]-2,9-dihydro-1,2-
,7,9-tetraaza-phenalen-3-one, 11
[0150] Synthesized using
4-(4-fluorophenyl)-1,2,3,6-tetrahydropyridine hydrochloride for
General Procedure D. 24% overall yield for last two steps. MS
(ES+): 376; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.51-2.53 (s, br,
2H), 2.77 (t, J=5.4 Hz, 2H), 3.24 (s, br, 2H), 3.46 (s, 2H), 6.16
(m, 1H), 7.16 (t, J=8.8 Hz, 2H), 7.46-7.52 (m, 3H), 7.63 (d, J=7.8
Hz, 1H), 7.44 (t, J=7.8 Hz, 1H), 11.18 (s, br, 1H), 11.79 (s, 1H).
A mesylate salt of 11 was prepared. .sup.1H NMR (400 MHz,
DMSO-d.sub.6): 2.34 (s, 3H), 2.84 (bs, 2H), 3.66 (m, 2H), 4.11 (m,
2H), 4.36 (s, 2H), 6.21 (m, 1H), 7.25 (t, J 8.8 Hz, 2H), 7.43 (d, J
7.4 Hz, 1H), 7.56-7.59 (m, 2H), 7.72 (d, J 7.4 Hz, 1H), 7.82 (t, J
7.5 Hz, 1H), 11.25 (s, br, 1H), 11.76 (s, 1H). Anal. Calcd. for
C.sub.21H.sub.18FN.sub.5O.sub.1.1.0CH.sub.3SOH. 0.2H.sub.2O: C,
55.62; H, 4.75; N, 14.74; S, 6.75. Found: C, 55.65; H, 4.71; N,
14.73; S, 6.74.
Preparation of
8-[4-(4-chloro-phenyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2,7,9-tetraaza-
-phenalen-3-one, 12
[0151] Synthesized using 1-(4-chlorophenyl)-piperazine for General
Procedure D. 23% overall yield for last two steps. A mesylate salt
of 12 was prepared. MS (ES+): 396; .sup.1H NMR (400 MHz,
DMSO-d.sub.6): 2.33 (s, 3H), 4.31 (s, 2H), 7.03 (d, J=9.3 Hz, 2H),
7.31 (d, J=9.3 Hz, 2H), 7.43 (d, J=8.5 Hz, 1H), 7.72 (d, J=8.5 Hz,
1H), 7.82 (t, J=7.9 Hz, 1H), 11.23 (s, br, 1H), 11.90 (s, 1H).
Anal. Calcd. for C.sub.20H.sub.19ClN.sub.6O.sub.1.1.0CH.sub.3SOH:
C, 51.37; H, 4.72; N, 17.12; S, 6.53. Found: C, 51.27; H, 4.91; N,
17.03; S, 6.48.
Preparation of
8-(4-phenyl-3,6-dihydro-2H-pyridin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraa-
za-phenalen-3-one, 13
[0152] Synthesized using 4-phenyl-1,2,3,6-tetrahydro-pyridine for
General Procedure D. 80% overall yield for last two steps. MS
(ES+): 358; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.56 (m, 2H), 2.78
(t, J 5.5 Hz, 2H), 3.25 (d, J 2.6 Hz, 2H), 3.47 (s, 2H), 6.19 (s,
1H), 7.23-7.27 (m, 1H), 7.24 (t, J=7.6 Hz, 2H), 7.15 (d, J=7.1, Hz,
2H), 7.51 (d, J=8.9 Hz, 1H), 7.62 (d, J=7.1 Hz, 1H), 7.75 (t, J=8.0
Hz, 1H), 11.27 (s, br, 1H), 11.78 (s, 1H). A mesylate salt of 13
was prepared. .sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.34 (s, 3H),
2.84-2.88 (m, 2H), 3.65-3.69 (m, 2H), 4.13 (s, 2H), 4.37 (s, 2H),
6.21-6.25 (m, 1H), 7.32-7.44 (m, 4H), 7.53 (d, J 8.6 Hz, 2H), 7.72
(d, J 7.3 Hz, 1H), 7.82 (t, J 8.1 Hz, 1H), 11.30 (s, br, 1H), 11.93
(s, 1H). Anal. Calcd. for C.sub.21H.sub.19N.sub.5O.1.0CH.sub.3SOH.
0.4H.sub.2O: C, 57.35; H, 5.21; N, 15.20; S, 6.96. Found: C, 57.30;
H, 5.16; N, 15.29; S, 7.10.
Preparation of
8-[(3,4-dichloro-benzylamino)-methyl]-2,9-dihydro-1,2,7,9-tetraaza-phenal-
en-3-one, 14
[0153] Synthesized using 3,4-dichlorobenzylamine for General
Procedure D. 10% overall yield for last two steps. A mesylate salt
of 14 was prepared. MS (ES+): 375; .sup.1H NMR (300 MHz,
DMSO-d.sub.6): 2.33 (s, 3H), 4.06 (s, 2H), 4.33 (s, 2H), 7.39 (d,
J=8.0 Hz, 1H), 7.53-7.57 (m, 1H), 7.69-7.88 (m, 4H), 11.31 (s, br,
1H), 11.91 (s, 1H).
Preparation of
8-{[2-(3-Fluoro-phenyl)-ethylamino]-methyl}-2,9-dihydro-1,2,7,9-tetraaza--
phenalen-3-one, 15
[0154] Synthesized using 3-fluorophenethylamine for General
Procedure D. 12% overall yield for last two steps. A mesylate salt
of 15 was prepared. MS (ES+): 338; .sup.1H NMR (300 MHz,
DMSO-d.sub.6): 2.34 (s, 3H), 3.02-3.08 (m, 2H), 3.34-3.38 (m, 2H),
4.14 (s, 2H), 7.08-7.18 (m, 3H), 7.37-7.44 (m, 2H), 7.71 (d, J 7.8
Hz, 1H), 7.82 (t, J 7.8 Hz, 1H), 11.92-11.35 (s, br, 1H), (s,
1H).
Preparation of
8-[(3-trifluoromethyl-benzylamino)-methyl]-2,9-dihydro-1,2,7,9-tetraaza-p-
henalen-3-one, 16
[0155] Synthesized using 3-(trifluoromethyl)benzylamine for General
Procedure D. 14% overall yield for last two steps. A mesylate salt
of 16 was prepared. MS (ES+): 374; .sup.1H NMR (300 MHz,
DMSO-d.sub.6): 2.33 (s, 3H), 4.10 (s, 2H), 4.43 (s, 2H), 7.39 (d, J
7.6 Hz, 1H), 7.69-7.86 (m, 5H), 7.99 (s, 1H), 11.25 (s, br, 1H),
11.91 (s, 1H). Anal. Calcd. for
C.sub.19H.sub.18F.sub.3N.sub.5O.1.0CH.sub.3SOH.1.0H.sub.2O: C,
46.82; H, 4.14; N, 14.37; S, 6.58. Found: C, 46.81; H, 4.17; N,
14.64; S, 6.35.
Preparation of
8-(1,4-dioxa-8-aza-spiro[4.5]dec-8-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-
-phenalen-3-one, 17
[0156] Synthesized using 4-piperidone ethylene ketal for General
Procedure D. 10% overall yield for last two steps. MS (ES-): 370;
.sup.1H NMR (300 MHz, DMSO-d.sub.6): 1.69-1.71 (m, 4H), 2.57 (s,
br, 4H), 3.35 (s, 2H), 3.87 (s, 4H), 7.51 (d, J 7.8 Hz, 1H), 7.62
(d, J=7.7 Hz, 1H), 7.74 (t, J=7.8 Hz, 1H), 11.23 (s, br, 1H), 11.76
(s, 1H). Anal. Calcd. for
C.sub.17H.sub.19N.sub.5O.sub.3.0.2H.sub.2O: C, 59.19; H, 5.67; N,
20.30. Found: C, 59.03; H, 5.60; N, 20.63.
Preparation of
8-{[2-(3,4-dichloro-phenyl)-ethylamino]-methyl}2,9-dihydro-1,2,7,9-tetraa-
za-phenalen-3-one, 18
[0157] Synthesized using 3,4-dichlorophenethylamine for General
Procedure D. 17% overall yield for last two steps. A mesylate salt
of 18 was prepared. MS (ES-): 387; .sup.1H NMR (300 MHz,
DMSO-d.sub.6): 2.36 (s, 3H), 3.04 (t, J=8.2 Hz, 2H), 3.37 (t, J=8.1
Hz, 2H), 4.14 (s, 2H), 7.30-7.43 (m, 2H), 7.61-7.75 (m, 3H),
7.79-7.84 (m, 1H), 11.31 (s, br, 1H), 11.91 (s, 1H).
Preparation of
8-{[2-(3-trifluoromethyl-phenyl)-ethylamino]-methyl}-2,9-dihydro-1,2,7,9--
tetraaza-phenalen-3-one, 19
[0158] Synthesized using 2-(3-Trifluoromethyl-phenyl)-ethylamine
for General Procedure D. 39% overall yield for last two steps. A
mesylate salt of 19 was prepared. MS (ES-): 387; .sup.1H NMR (300
MHz, DMSO-d.sub.6): 3.74 (s, 3H), 3.13 (t, J=8.1 Hz, 2H), 3.30 (t,
J 8.2 Hz, 2H), 4.15 (s, 2H), 7.40-7.43 (m, 1H), 7.62-7.72 (m, 4H),
7.79-7.85 (m, 1H), 11.35 (s, br, 1H), 11.92 (s, 1H).
Preparation of
8-[(1-Aza-bicyclo[2.2.2]oct-3-ylamino)-methyl]-2,9-dihydro-1,2,7,9-tetraa-
za-phenalen-3-one, 20
[0159] Synthesized using (S)-(-)-3-aminoquinuclidine for General
Procedure D. 23% overall yield for last two steps. A mesylate salt
of 20 was prepared. MS (ES+): 325; .sup.1H NMR (300 MHz,
DMSO-d.sub.6): 1.97-2.03 (m, 3H), 2.20-2.35 (m, 1H), 2.35-2.44 (m,
2H), 2.42 (s, 3H), 3.72-3.80 (m, 6H), 4.15-4.21 (m, 1H), 4.38 (s,
2H), 7.46 (d, J 7.6, 1H) 7.69-7.72 (m, 1H), 7.78-7.84 (m, 1H), 8.63
(s, br, 3H).
Preparation of
8-(4-ethyl-piperazin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3--
one, 21
[0160] Synthesized using ethylpiperazine for General Procedure D.
35% overall yield for last two steps. A mesylate salt of 21 was
prepared. MS (ES+): 313; .sup.1H NMR (300 MHz, DMSO-d.sub.6): 1.25,
(t, J 7.4 Hz, 3H), 2.41 (s, 6H), 2.51-3.87 (m, 10H), 3.87 (s, 2H),
7.70 (d, J=8.0 Hz, 1H), 7.81 (d, J=7.9 Hz, 1H), 7.91 (t, J=8.1 Hz,
1H), 9.82 (s, 1H), 11.96 (s, 1H). .sup.13C NMR (DMSO-d.sub.6):
157.40, 155.99, 140.65, 135.96, 133.84, 126.72, 119.71, 118.65,
115.85, 56.09, 50.30, 49.05, 48.66, 8.51. Anal. Calcd. for
C.sub.16H.sub.20N.sub.6O.2.0CH.sub.3SO.sub.3H.1.2H.sub.2O.: C,
40.84; H, 5.43; N, 15.79. Found: C, 41.09; H, 5.82; N, 15.97.
Preparation of
8-(4-methyl-piperazin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-
-one, 22
[0161] Synthesized using methylpiperazine for General Procedure D.
29% overall yield for last two steps. A mesylate salt of 22 was
prepared. MS (ES+): 299; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.38
(s, 3H), 2.58-2.63 (m, 2H), 3.09-3.18 (m, 4H), 3.40-3.45 (m, 2H),
3.51 (s, 2H), 7.50 (d, J 7.8 Hz, 1H), 7.67 (d, J 7.8 Hz, 1H), 7.79
(t, J 7.8 Hz, 1H), 9.53 (s, br, 1H), 11.85 (s, 1H). Anal. Calcd.
for C.sub.15H.sub.18N.sub.6O.1.15CH.sub.3SO.sub.3H. 1.0H.sub.2O.:
C, 45.44; H, 5.81; N, 19.69; S, 8.64. Found: C, 45.18; H, 5.88; N,
19.83; S, 8.68.
Preparation of
8-(4-benzyl-[1,4]diazepan-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenal-
en-3-one. 23
[0162] Synthesized using 1-benzyl-[1,4]diazepane for General
Procedure D. 24% overall yield for last two steps. MP:
140-142.degree. C.; MS (ES-): 387; .sup.1H NMR (400 MHz,
CDCl.sub.3): 1.88 (m, 2H), 2.77 (m, 4H), 2.89 (m, 4H), 3.62 (s,
2H), 3.69 (s, 2H), 7.20-7.42 (m, 6H), 7.45 (s, br, 1H), 7.74 (t, J
7.8 Hz, 1H), 7.87 (d, J 7.6 Hz, 1H), 11.50 (s, br, 1H); Anal.
Calcd. for C.sub.22H.sub.24N.sub.6O.1.35H.sub.2O.: C, 64.01; H,
6.52; N, 20.36. Found: C, 64.18; H, 6.59; N, 20.46.
[0163] An HCl salt of 23 was prepared: to a solution of 23 (0.5 g)
in 20 mL of dioxane was bubbled HCl gas for 30 min. The solution
was stirred at room temperature overnight. After filtration, the
precipitate was washed with dioxane to afford 0.25 g (48%) of
analytically pure off white solid, an HCl salt of 23. .sup.1H NMR
(400 MHz, D.sub.2O): 2.08 (m, 2H), 3.36 (m, 4H), 3.56 (m, 4H), 4.04
(s, 2H), 4.24 (s, 2H), 7.02 (d, 1H), 7.20-7.35 (m, 5H); 7.36 (d,
1H), 7.45 (t, 1H); Anal. Calcd. for C.sub.22H.sub.24N.sub.6O.2.0
HCl. 1.15H.sub.2O.: C, 54.81; H, 5.92; N, 17.43. Found: C, 54.81;
H, 5.92; N, 17.36.
Preparation of
4-(3-oxo-2,9-dihydro-3H-1,2,7,9-tetraaza-phenalen-8-ylmethyl)-[1,4]diazep-
ane-1-carboxylic acid tert-butyl ester, 24
[0164] Synthesized using [1,4]diazepane-1-carboxylic acid t-butyl
ester for General Procedure D. 30% overall yield for last two
steps. MP: 219-221.degree. C.; MS (ES-): 397; .sup.1H NMR. (400
MHz, CDCl.sub.3): 1.46 (s, 9H); 1.88 (m, 2H); 2.83 (m, 4H); 3.50
(m, 4H); 3.59 (s, 2H); 7.63 (m, 1H), 7.72-7.86 (m, 3H), 11.90 (s,
br, 1H). Anal. Calcd. for
C.sub.20H.sub.26N.sub.6O.sub.3.0.5H.sub.2O.: C, 58.95; H, 6.68; N,
20.62. Found: C, 58.83; H, 6.69; N, 20.60.
Preparation of
8-[4-(4-fluoro-benzyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-1,2,7,9-tetr-
aaza-phenalen-3-one, 25
[0165] Synthesized using 1-(4-fluoro-benzyl)-[1,4]diazepane for
General Procedure D. 35% overall yield for last two steps. MP:
163-165.degree. C.; MS (ES-): 405; .sup.1H NMR (400 MHz,
CDCl.sub.3): 1.87 (m, 2H), 2.72 (m, 4H), 2.88 (m, 4H), 3.63 (s,
2H), 3.65 (s, 2H), 6.99 (t, J=8.4 Hz, 2H), 7.30 (m, 3H) 7.61 (s,
br, 1H), 7.78 (m, 1H); 7.93 (d, J=7.3 Hz 1H), 10.82 (s, br, 1H).
Anal. Calcd. for C.sub.22H.sub.23N.sub.6O.1.5H.sub.2O.: C, 60.96;
H, 6.05; N, 19.39. Found: C, 61.07; H, 5.97; N, 19.59.
[0166] A mesylate salt of 25 was prepared. .sup.1H NMR (400 MHz,
D.sub.2O): 2.06 (m, 2H), 2.70 (s, 3H), 3.06 (m, 2H), 3.24 (m, 2H),
3.46 (m, 4H), 3.65 (s, 4H), 3.74 (s, 2H), 4.33 (s, 2H), 7.25 (m,
3H), 7.46 (m, 3H), 7.62 (t, J=8.4 Hz, 1H). Anal. Calcd. for
C.sub.22H.sub.23FN.sub.6O.1.3CH.sub.3SO.sub.3H.0.5C.sub.4H.sub.8O.sub.2.2-
.0H.sub.2O.: C, 49.70; H, 5.97; N, 13.74; S, 6.82. Found: C, 49.40;
H, 5.97; N, 13.37; S, 6.65.
Preparation of
8-[1,4]diazepan-1-ylmethyl-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-one,
26
[0167] Synthesized from compound 24. To a solution of 24 (1.5 g,
3.7 mmol) in 30 mL of CH.sub.2Cl.sub.2 was added 6 mL of TFA while
stirring at room temperature. After 30 minutes, the solvents were
evaporated and the residue was washed with acetonitrile to afford
1.0 g (90%) of analytically pure white solid. MP: 147-149.degree.
C.; MS (ES-): 297; .sup.1H NMR (400 MHz, D.sub.2O): 1.96 (m, 2H),
2.82 (t, 2H), 3.01 (t, 2H), 3.28 (t, 4H), 3.53 (s, 2H), 7.22 (d,
1H), 7.47 (d, 1H), 7.61 (t, 1H). Anal. Calcd. for
C.sub.15H.sub.18N.sub.6O.1.1CF.sub.3CO.sub.2H.1.0H.sub.2O.: C,
46.76; H, 4.81; N, 19.02. Found: C, 46.64; H, 4.98; N, 19.02.
Preparation of
8-[4-(2-trifluoromethyl-benzoyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-1,-
2,7,9-tetraaza-phenalen-3-one, 27
[0168] Synthesized from compound 26. To a solution of compound 26
(0.2 g, 0.6 mmol) in 5 mL of CH.sub.2Cl.sub.2 was added 1 mmol of
TEA and 0.8 mmol of 2-trifluoromethyl-benzoyl chloride. The
reaction was stirred overnight at room temperature. After the
solvents were evaporated, the residue was purified with
semi-preparative HPLC to afford a solid (15% yield). MP:
140-142.degree. C.; MS (ES-): 469; .sup.1H NMR (400 MHz,
CDCl.sub.3): 1.92-2.10 (m, 2H), 2.91-3.10 (m, 4H), 3.36-3.44 (m,
2H), 3.64-3.74 (m, 2H), 3.93 (m, 2H), 7.38 (m, 1H), 7.57 (m, 3H),
7.79 (m, 2H), 7.93 (m, 1H). Anal. Calcd. for
C.sub.23H.sub.21F.sub.3N.sub.6O.sub.2.0.9 HCl: C, 54.89; H, 4.39;
N, 16.70. Found: C, 54.93; H, 4.43; N, 16.34.
Preparation of
8-[4-(3-chloro-benzoyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-1,2,7,9-tet-
raaza-phenalen-3-one, 28
[0169] Synthesized from compound 26. To a solution of compound 26
(0.2 g, 0.6 mmol) in 5 mL of CH.sub.2Cl.sub.2 was added 1 mmol of
TEA and 0.8 mmol of 3-chloro-benzoyl chloride. The reaction was
stirred overnight at room temperature. After the solvents were
evaporated, the residue was purified with semi-preparative HPLC to
afford a solid (16% yield). MP: 147-149.degree. C.; MS (ES-): 436;
.sup.1H NMR (400 MHz, CDCl.sub.3): 1.88-2.08 (m, 2H), 2.86-3.07 (m,
4H), 3.52-3.71 (m, 4H), 3.81-3.89 (m, 2H), 7.33-7.43 (m, 4H), 7.62
(d, 1H), 7.81 (t, 1H), 7.90 (t, 1H). Anal. Calcd. for
C.sub.22H.sub.21C.sub.1N.sub.6O.sub.2.0.7H.sub.2O: C, 54.89; H,
4.39; N, 16.70. Found: C, 54.93; H, 4.43; N, 16.34.
Preparation of
8-(4-pyridin-2-yl-piperazin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phen-
alen-3-one 30
[0170] Synthesized using 1-pyridin-2-yl-piperazine for General
Procedure D. 20% overall yield for last two steps. A mesylate salt
of 30 was prepared. MS (ES-): 360; .sup.1H NMR (400 MHz,
DMSO-d.sub.6): 2.37 (s, 6H), 3.52 (s, br, 4H), 3.93 (s, br, 4H),
4.30 (s, 2H), 6.93 (t, J 6.6 Hz, 1H), 7.25 (d, J 8.6 Hz, 1H), 7.47
(d, J 7.8 Hz, 1H), 7.73 (d, J 7.8 Hz, 1H), 7.82-7.91 (m, 2H),
8.16-8.18 (m, 1H), 11.96 (s, 1H). Anal. Calcd. for
C.sub.19H.sub.19N.sub.7O.1.9CH.sub.3SO.sub.3H.1.2H.sub.2O.: C,
44.38; H, 5.17; N, 17.33; S, 10.77. Found: C, 44.21; H, 5.19; N,
17.28; S, 10.68.
Preparation of
8-{[2-(2-fluoro-phenyl)-ethylamino]-methyl}-2,9-dihydro-1,2,7,9-tetraaza--
phenalen-3-one, 31
[0171] Synthesized using 2-(2-fluoro-phenyl)-ethylamine for General
Procedure D. 20% overall yield for last two steps. A mesylate salt
of 31 was prepared. MS (ES-): 336; .sup.1H NMR (400 MHz,
DMSO-d.sub.6): 2.41 (s, 5H), 3.02 (t, J 7.6 Hz, 2H), 3.32 (t, J 8.3
Hz, 2H), 4.16 (s, 2H), 7.19 (t, J 8.8 Hz, 2H), 7.32-7.35 (m, 2H),
7.42 (d, J 7.8 Hz, 1H), 7.71 (d, J=7.8 Hz, 1H), 7.82 (t, J=8.1 Hz,
1H), 9.10 (s, br, 1H), 11.92 (s, 1H). Anal. Calcd. for
C.sub.18H.sub.16FN.sub.5O.1.75CH.sub.3SO.sub.3H.0.75H.sub.2O: C,
45.70; H, 4.76; N, 13.49; S, 10.81. Found: C, 45.45; H, 4.69; N,
13.42; S, 11.10.
Preparation of
8-[4-(4-fluoro-phenyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2,7,9-tetraaza-
-phenalen-3-one, 32
[0172] Synthesized using 4-(4-fluoro-phenyl)-piperazine for General
Procedure D. 57% overall yield for last two steps. A mesylate salt
of 32 was prepared. MS (ES-): 377; .sup.1H NMR (400 MHz,
DMSO-d.sub.6): 2.40 (s, 5H), 3.45 (s, br, 4H), 3.59 (s, br, 4H),
4.37 (s, 2H), 7.03-7.15 (m, 4H), 7.44 (d, J=7.8 Hz, 1H), 7.72 (d,
J=7.8 Hz, 1H), 7.83 (t, J=7.8 Hz, 1H), 9.8 (s, br, 1H), 11.94 (s,
1H). Anal. Calcd. for
C.sub.20H.sub.19FN.sub.6O.1.65CH.sub.3SO.sub.3H: C, 46.85; H, 5.01;
N, 15.14; S, 9.53. Found: C, 46.74; H, 5.15; N, 15.14; S, 9.53.
Preparation of
8-{[2-(4-Fluoro-phenyl)-ethylamino]-methyl}-2,9-dihydro-1,2,7,9-tetraaza--
phenalen-3-one, 33
[0173] Synthesized using 2-(4-fluoro-phenyl)-ethylamine for General
Procedure D. 19% overall yield for last two steps. A mesylate salt
of 33 was prepared. MS (ES-): 336; .sup.1H NMR (400 MHz,
DMSO-d.sub.6): 2.38 (s, 6H), 3.06-3.10 (m, 2H), 3.30-3.34 (m, 2H),
4.18 (s, 2H), 7.19-7.22 (m, 2H), 7.34-7.42 (m, 3H), 7.71 (d, J=8.6
Hz, 1H), 7.82 (t, J 7.8 Hz, 1H), 9.6 (s, br, 1H), 11.92 (s, 1H).
Anal. Calcd. for C.sub.18H.sub.16FN.sub.5O.2.0 CH.sub.3SO.sub.3H:
C, 45.36; H, 4.57; N, 13.22; S, 12.11. Found: C, 45.34; H, 4.58; N,
13.16; S, 11.88.
Preparation of
8-(4-acetyl-[1,4]diazepan-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenal-
en-3-one 34
[0174] Synthesized using [1,4]diazepane-1-yl-ethanone for General
Procedure D. 16% overall yield for last two steps. MP:
191-193.degree. C.; MS (ES-): 339; .sup.1H NMR (400 MHz,
CDCl.sub.3): 2.11 (s, 3H), 2.84-2.93 (m, 4H), 3.56-3.76 (m, 6H),
7.66 (m, 1H), 7.83-7.92 (m, 2H), 9.3 (s, br, 1H), 11.3 (s, br, 1H).
Anal. Calcd. for C.sub.17H.sub.20N.sub.6O.sub.2.0.6H.sub.2O: C,
58.14; H, 6.08; N, 23.93. Found: C, 58.09; H, 6.18; N, 24.08.
Preparation of
8-(phenethylamino-methyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-one,
35
[0175] Synthesized using phenetyhlamine for General Procedure D.
29% overall yield for last two steps. A mesylate salt of 35 was
prepared. MS (ES-): 358; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.32
(s, 3H), 3.00-3.04 (m, 2H), 3.31-3.36 (m, 2H), 4.15 (s, 1H),
7.27-7.42 (m, 6H), 7.71 (d, J=7.8 Hz, 1H), 7.82 (t, J=7.8 Hz, 1H),
9.70 (s, br, 1H), 11.92 (s, 1H). Anal. Calcd. for
C.sub.18H.sub.17N.sub.5O.1.0CH.sub.3SO.sub.3H.1.8H.sub.2O: C,
50.95; H, 5.54; N, 15.64; S, 7.16. Found: C, 50.95; H, 5.54; N,
15.64; S, 7.16.
Preparation of
8-(4-phenyl-piperidin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-
-one. 36
[0176] Synthesized using 4-phenyl-piperidine for General Procedure
D. 33% overall yield for last two steps. MS (ES-): 318; .sup.1H NMR
(400 MHz, DMSO-d.sub.6): 1.87-1.93 (m, 4H), 2.37-2.46 (m, 2H), 2.56
(m, 1H), 3.10-3.14 (m, 2H), 3.54 (s, 2H), 7.17-7.34 (m, 5H), 7.56
(bs, 1H), 7.76 (t, J 7.8 Hz, 1H), 7.93 (d, J 7.8 Hz, 1H), 11.10 (s,
br, 1H), 11.76 (s, 1H). A mesylate salt of 36 was prepared. .sup.1H
NMR (400 MHz, D.sub.2O): 2.08 (m, 4H), 2.95 (m, 1H), 3.34 (m, 2H),
3.84 (m, 2H), 4.23 (s, 2H), 7.21-7.39 (m, 6H), 7.59 (m, 1H), 7.70
(m, 1H). Anal. Calcd. for
C.sub.21H.sub.21N.sub.5O.1.3CH.sub.3SO.sub.3H.0.5H.sub.2O: C,
54.29; H, 5.56; N, 14.19; S, 8.45. Found: C, 54.03; H, 5.65; N,
13.98; S, 8.64.
Preparation of
8-(1,3-dihydro-isoindol-2-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen-
-3-one. 37
[0177] Synthesized using isoindoline for General Procedure D. 40%
overall yield for last two steps. MS (ES-): 316; .sup.1H NMR (400
MHz, DMSO-d.sub.6): 3.77 (s, 2H), 4.04 (s, 4H), 7.20-7.30 (m, 4H),
7.49 (d, J=7.8 Hz, 1H), 7.6 (d, J=7.8 Hz, 1H), 7.74 (t, J=7.8 Hz,
1H), 11.34 (s, br, 1H), 11.78 (s, 1H). A mesylate salt of 37 was
prepared. .sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.34 (s, 3H), 4.64
(s, 2H), 4.87 (s, 4H), 7.39-7.46 (m, 5H), 7.72 (d, J=7.8 Hz, 1H),
7.83 (t, J=8.1 Hz, 1H), 11.30 (s, br, 1H), 11.95 (s, 1H). Anal.
Calcd. for
C.sub.18H.sub.15N.sub.5O.1.25CH.sub.3SO.sub.3H.2.0H.sub.2O: C,
48.83; H, 5.11; N, 14.79; S, 8.46. Found: C, 48.80; H, 5.11; N,
14.97; S, 8.71.
Preparation of
8-(4-benzenesulfonyl-[1,4]diazepan-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraa-
za-phenalen-3-one, 38
[0178] Synthesized from compound 26. To a solution of 26 (0.2 g,
0.67 mmol) in 5 mL of CH.sub.2Cl.sub.2 was added TEA (2 mmol) and
benzensulfonyl chloride (1 mmol). The mixture was stirred at room
temperature over night. After the solvents were evaporated, the
residue was poured into 10 mL of H.sub.2O and the product was
purified by preparative HPLC to afford analytically pure white
solid (5% yield). MP: 265-268.degree. C.; MS (ES-): 437; .sup.1H
NMR (400 MHz, DMSO-d.sub.6): 1.79 (m, 2H), 2.50 (m, 41), 2.79 (m,
4H), 3.51 (s, 2H), 7.44 (d, 1H), 7.62-7.79 (m, 71), 11.1 (s, br,
1H), 11.75 (s, 1H). Anal. Calcd. for
C.sub.21H.sub.22N.sub.5O.sub.3S. 0.5H.sub.2O: C, 56.36; H, 5.18; N,
18.78; S, 7.17. Found: C, 56.44; H, 5.12; N, 19.00; S, 7.19.
[0179] A mesylate salt of 38 was prepared. MS (ES+): 439; .sup.1H
NMR (400 MHz, D.sub.2O): 2.18 (m, 2H), 2.35 (s, 6H), 3.36 (m, 2H),
3.65 (m, 6H), 4.3 (s, 2H), 7.24 (d, 1H), 7.51-7.71 (m, 7H). Anal.
Calcd. for C.sub.21H.sub.22N.sub.6O.sub.3S.
1.8CH.sub.3SO.sub.3H.1.0H.sub.2O: C, 43.50; H, 5.00; N, 13.35; S,
14.26. Found: C, 43.61; H, 5.00; N, 13.15; S, 14.59.
Preparation of
8-(4-pyridin-4-yl-piperazin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phen-
alen-3-one 39
[0180] Synthesized using 1-(4-pyridyl)piperazine for General
Procedure D. 10% overall yield for last two steps. MS (ES-): 360;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.80 (t, J=5.0 Hz, 4H), 3.61
(t, J 5.0 Hz, 4H), 3.99 (s, 2H), 6.83 (d, J 7.1 Hz, 2H), 7.42-7.45
(m, 1H), 7.73-7.81 (m, 2H), 8.26 (d, J 7.1 Hz, 2H), 11.20 (s, br,
1H), 11.90 (s, 1H). An HCl salt of 39 was prepared. .sup.1H NMR
(400 MHz, D.sub.2O): 2.74-2.77 (m, 4H), 3.43 (s, 2H), 3.35-3.69 (m,
4H), 6.93 (d, J=7.1 Hz, 2H), 7.13 (d, J=8.0 Hz, 1H), 7.37 (d, J=7.8
Hz, 1H), 7.58 (t, J=7.8 Hz, 1H), 7.92 (d, J=7.1 Hz, 2H). Anal.
Calcd. for C.sub.19H.sub.19N.sub.7O.1.0 HCl. 2.5H.sub.2O: C, 51.53;
H, 5.69; N, 22.14; Cl, 8.00. Found: C, 51.46; H, 5.69; N, 21.90;
Cl, 8.27.
Preparation of
8-(4-benzyl-piperazin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-
-one 40
[0181] Synthesized using 4-benzyl-piperazine for General Procedure
D. 12% overall yield for last two steps. MS (ES-): 373; .sup.1H NMR
(400 MHz, DMSO-d.sub.6): 2.44 (s, br, 4H), 3.35 (s, br, 4H), 3.48
(s, 2H), 7.23-7.34 (m, 5H), 7.49 (d, J 8.8 Hz, 1H), 7.62 (d, J=7.8
Hz, 1H), 7.74 (t, J=7.8 Hz, 1H), 11.10 (s, br, 1H), 11.77 (s, 1H).
An HCl salt of 40 was prepared. .sup.1H NMR (400 MHz, D.sub.2O):
2.54-2.70 (m, 2H), 3.10-3.50 (m, 6H), 3.48 (s, 2H), 4.35 (s, 2H),
7.20 (d, J=8.1 Hz, 1H), 7.45 (t, J=7.8 Hz, 1H), 7.47-7.51 (m, 5H),
7.62 (t, J=8.1 Hz, 1H). Anal. Calcd. for
C.sub.21H.sub.22N.sub.6O.1.0 HCl. 2.5H.sub.2O: C, 55.32; H, 6.19;
N, 18.43. Found: C, 55.54; H, 6.08; N, 18.32.
Preparation of
8-(4-methyl-[1,4]diazepan-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-phenal-
en-3-one 41
[0182] Synthesized using 1-methyl-[1,4]diazepane for General
Procedure D. 24% overall yield for last two steps. MS (ES-): 311;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.75 (m, 2H), 2.26 (s, 3H),
2.55 (m, 4H), 2.79 (m, 4H), 3.48 (s, 2H), 7.52 (d, J 8.2 Hz, 1H),
7.64 (d, J=7.2 Hz, 1H), 7.75 (t, J=8.1 Hz, 1H), 11.55 (s, 1H).
Anal. Calcd. for C.sub.16H.sub.20N.sub.6O.0.95H.sub.2O: C, 58.33;
H, 6.70; N, 25.51. Found: C, 58.32; H, 6.65; N, 25.53.
Preparation of
8-[4-(1H-indol-3-yl)-piperidin-1-ylmethyl]-2,9-dihydro-1,2,7,9-tetraaza-p-
henalen-3-one, 42
[0183] Synthesized using 3-piperidin-4-yl-1H-indole for General
Procedure D. 19% overall yield for last two steps. MS (ES-): 397;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.83-1.94 (m, 4H), 2.31 (m,
2H), 2.50 (s, 2H), 2.79-2.99 (m, 3H), 6.96-7.09 (m, 3H), 7.32 (d,
J=8.1 Hz, 1H), 7.54-7.63 (m, 3H), 7.75 (t, J=7.3 Hz, 1H), 10.79 (s,
1H), 11.80 (s, 1H). A mesylate salt of 42 was prepared. .sup.1H NMR
(400 MHz, DMSO-d.sub.6): 2.15 (m, 4H), 2.32 (s, 3H), 3.11 (m, 1H),
3.52 (m, 2H), 3.73 (m, 2H), 4.29 (s, 2H), 7.10-7.18 (m, 3H), 7.36
(d, 1H); 7.46 (d, J 8.2 Hz, 1H), 7.69-7.83 (m, 3H), 10.91 (s, 1H),
11.93 (s, 1H). Anal. Calcd. for
C.sub.23H.sub.22N.sub.6O.1.0CH.sub.3SO.sub.3H.1.25H.sub.2O: C,
55.23; H, 5.62; N, 16.91; S, 6.14. Found: C, 55.27; H, 5.53; N,
16.95; S, 6.00.
Preparation of
8-[(2-pyridin-4-yl-ethylamino)-methyl]-2,9-dihydro-1,2,7,9-tetraaza-phena-
len-3-one 43
[0184] Synthesized using 4-ethylamino-pyridine for General
Procedure D. 10% overall yield for last two steps. An HCl salt of
43 was prepared. MS (ES-): 319; .sup.1H NMR (400 MHz, D.sub.2O):
3.28 (t, J 7.8 Hz, 2H), 3.53 (t, J 7.8 Hz, 2H), 4.09 (s, 2H), 7.02
(d, J=8.0 Hz, 1H), 7.35 (d, J=8.0 Hz, 1H), 7.52 (t, J=8.0 Hz, 1H),
7.70 (d, J=5.3 Hz, 2H), 8.52 (d, J=5.3 Hz, 21-1). Anal. Calcd. for
C.sub.17H.sub.16N.sub.6O.1.3 HCl. 2.6H.sub.2O.0.1N.sub.2H.sub.4: C,
47.52; H, 5.38; N, 20.27. Found: C, 47.12; H, 5.26; N, 20.67.
Preparation of
8-(3,4-dihydro-1H-isoquinolin-2-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-ph-
enalen-3-one, 44
[0185] Synthesized using 1,2,3,4-tetrahydro-isoquinoline for
General Procedure D. 30% overall yield for last two steps. MS
(ES-): 330; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.81-2.90 (m, 4H);
3.52 (s, 2H), 3.72 (s, 2H), 7.05-7.25 (m, 4H), 7.51 (d, J=7.8 Hz,
1H), 7.63 (d, J=8.0 Hz, 1H), 7.74 (t, J=8.0 Hz, 1H), 11.30 (s, br,
1H), 11.91 (s, 1H). Anal. Calcd. for C.sub.19H.sub.17N.sub.5O: C,
68.87; H, 5.17; N, 21.13. Found: C, 68.34; H, 5.19; N, 21.30.
[0186] A mesylate salt of 44 was prepared. MS (ES-): 330; .sup.1H
NMR (400 MHz, D.sub.2O): 2.80 (s, 3H), 3.31 (t, 2H), 3.85 (m, 2H),
4.47 (s, 2H), 4.68 (s, 2H), 7.23 (d, J 7.8 Hz, 1H), 7.28-7.42 (m,
4H), 7.67 (d, J 8.0 Hz, 1H); 7.80 (t, J 7.9 Hz, 1H). Anal. Calcd.
for C.sub.19H.sub.17N.sub.5O.1.12CH.sub.3SO.sub.3H.2.0H.sub.2O: C,
50.87; H, 5.41; N, 14.74; S, 7.56. Found: C, 50.89; H, 5.47; N,
14.84; S, 7.63.
Preparation of
8-(5,6-Dimethoxy-3,4-dihydro-1H-isoquinolin-2-ylmethyl)-2,9-dihydro-1,2,7-
,9-tetraaza-phenalen-3-one, 45
[0187] Synthesized using
5,6-dimethoxy-1,2,3,4-tetrahydro-isoquinoline for General Procedure
D. 29% overall yield for last two steps. MS (ES-): 311; .sup.1H NMR
(400 MHz, DMSO-d.sub.6): 2.79 (s, 4H), 3.49 (s, 2H), 3.61 (s, 2H),
3.67 (s, 3H), 3.70 (s, 3H), 6.69 (d, J=8.8 Hz, 2H), 7.48 (d, J=7.6
Hz, 1H), 7.63 (d, J=7.8 Hz, 1H), 7.74 (t, J=7.6 Hz, 1H), 11.55 (s,
1H). Anal. Calcd. for C.sub.21H.sub.21N.sub.5O.sub.3: C, 64.44; H,
5.41; N, 17.89. Found: C, 64.24; H, 5.43; N, 17.98.
[0188] A mesylate salt of 45 was prepared. MS (ES-): 330; .sup.1H
NMR (400 MHz, D.sub.2O): 2.82 (s, 3H), 3.21 (t, 2H), 3.65-3.85 (m,
8H), 4.48 (s, 2H), 4.60 (s, 2H), 6.75 (s, 1H), 6.83 (s, 1H), 7.38
(d, 1H), 7.71 (d, 1H), 7.82 (t, 1H). Anal. Calcd. for
C.sub.21H.sub.21N.sub.5O.sub.3. 1.18CH.sub.3SO.sub.3H.1.75H.sub.2O:
C, 49.70; H, 5.49; N, 13.07; S, 7.03. Found: C, 49.77; H, 5.49; N,
13.17; S, 7.03.
Preparation of
8-[4-(3-Trifluoromethyl-benzenesulfonyl)-[1,4]diazepan-1-ylmethyl]-2,
dihydro-1,2,7,9-tetra-phenalen-3-one 46
[0189] Synthesized from compound 26. To a solution of 26 (0.2 g,
0.67 mmol) in 5 mL of CH.sub.2Cl.sub.2 was added TEA (2 mmol) and
3-trifluoromethyl-benzenesulfonyl chloride (1 mmol). The mixture
was stirred at room temperature over night. After the solvents were
evaporated, the residue was poured into 10 mL of H.sub.2O and the
product was purified by preparative HPLC to afford analytically
pure white solid (15% yield). MS (ES+): 507; .sup.1H NMR (400 MHz,
DMSO-d.sub.5): 1.82 (m, 2H), 2.73-2.81 (m, 4H), 3.25-3.42 (m, 61),
7.44 (d, J=7.8 Hz, 1H), 7.63 (d, J=7.2 Hz, 1H), 7.74 (t, J=7.8 Hz,
1H), 7.89 (t, J=8.2 Hz, 1H), 8.04-8.13 (m, 3H), 11.10 (s, br, 1H),
11.75 (s, 1H). Anal. Calcd. for
C.sub.22H.sub.21F.sub.3N.sub.6O.sub.3S. 1.1H.sub.2O: C, 50.21; H,
4.44; N, 15.97; S, 6.09. Found: C, 50.19; H, 4.54; N, 15.50; S,
5.97.
##STR00025##
General Procedure F: Preparation of Compounds 47a and 47B:
[0190] Displacement of the chloro group of compound 4 with
piperazine or [1,4]diazepane using General procedure F provides the
compound 47A or 47B. To a stirring solution of 4 (1 eq) in
acetonitrile was added piperazine or [1,4]diazepane (large excess)
under a blanket of nitrogen. The solution was allowed to stir
overnight and then evaporated to dryness. The crude material was
purified via silica plug with 9:1 dichloromethane:methanol to
afford a white solid,
4-Oxo-2-piperazin-1-ylmethyl-3,4-dihydro-quinazoline-5-carboxylic
acid methyl ester, 47A or
2-[1,4]diazepan-1-ylmethyl-4-oxo-3,4-dihydro-quinazoline-5-carboxylic
acid methyl ester, 47B.
General Procedure G: Preparation of compounds 48A and 48B:
[0191] A reaction of amine 47A or 47B with various sulfonyl
chloride yields sulfonyl amide 48A or 48B. To a stirring solution
of 47A or 47B (1.0 eq) in pyridine was added various sulfonyl
chloride (1.1 eq). The reaction was allowed to stir overnight and
then was evaporated to dryness. The residue was then extracted with
dichloromethane and washed with brine. The product was evaporated
to dryness and used without further purification.
General Procedure E: Preparation of Compounds 49A and 49B:
[0192] A 2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-one ring can be
formed by condensation of the compound 48A or 48B with hydrazine.
To a solution of the compounds 6 in absolute ethanol is added
excess anhydrous hydrazine at room temperature. The solution is
refluxed for overnight and cooled to room temperature. Ice-cold
water is added and white solid is separated. The solid is collected
by vacuum filtration and washed with water and small amount of
methanol to give white solid products 6 in 40-90% of yield. An
example was given in the preparation of compounds 49A and 49B.
Example 2
Preparation of
8-[4-(4-methoxy-benzenesulfonyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2,7,-
9-tetraaza-phenalen-3-one, 50
[0193] To a stirring solution of 4 (2.2 g, 8.73 mmol, 1 eq) in 200
mL of acetonitrile was added piperazine (14 g, 0.162 mol, large
excess) under a blanket of nitrogen. The solution was allowed to
stir overnight and then evaporated to dryness. The crude material
was purified via silica plug with 9:1 dichloromethane:methanol to
afford 2.0 g of a fluffy white solid,
4-Oxo-2-piperazin-1-ylmethyl-3,4-dihydro-quinazoline-5-carboxylic
acid methyl ester, 47A. MS (ES-): 301; .sup.1H NMR. (400 MHz,
DMSO-d.sub.6): 2.40-2.43 (m, 4H), 2.69-2.72 (m, 4H), 3.41 (s, 2H),
3.83 (s, 3H), 7.44 (d, J 7.2 Hz, 1H), 7.74 (d, J=8.2 Hz, 1H), 7.82
(t, J=7.8 Hz, 1H).
[0194] To a stirring solution of 47A (170 mg, 0.56 mmol, 1 eq) in 5
mL of pyridine was added 4-methoxybenzene sulfonyl chloride (130
mg, 0.62 mmol, 1.1 eq) resulting in a bright yellow solution. The
reaction was allowed to stir overnight and then was evaporated to
dryness. The waxy residue was then extracted with dichloromethane
and washed with brine. The crude material was dissolved in 10 mL of
EtOH and 5 mL of hydrazine monohydrate (large excess). This
solution was refluxed overnight resulting in a heavy white
precipitate which was filtered, washed with ethyl ether and dried
to give an off white solid. This solid was then purified via
chromatography to afford 112 mg of analytically pure compound 50. A
mesylate salt of 50 was prepared. 8% overall yield for last three
steps. MS (ES+): 455; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.34 (s,
3H), 3.19 (bs, 4H), 3.44 (bs, 4H), 3.89 (s, 3H), 4.20 (s, 2H), 7.25
(d, J=9.0 Hz, 2H), 7.72 (d, J=7.8 Hz, 1H), 7.70-7.83 (m, 4H), 11.20
(s, br, 1H), 11.93 (s, 1H). Anal. Calcd. for
C.sub.21H.sub.22N.sub.6O.sub.4S. 1.5
CH.sub.3SO.sub.3H.3.0H.sub.2O.0.1N.sub.2H.sub.4: C, 41.20; H, 5.29;
N, 13.24; S, 12.22. Found: C, 41.07; H, 5.09; N, 13.53; S,
12.62.
[0195] The following compounds were synthesized from the similar
procedures of preparation of compound 50, using the appropriate
corresponding sulfonyl chloride.
Preparation of
8-[4-(3-fluoro-benzenesulfonyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2,7,9-
-tetraaza-phenalen-3-one, 51
[0196] Synthesized using 3-fluoro-benzenesulfonyl chloride and
compound 47A for General Procedure G. A mesylate salt of 51 was
prepared. 35% overall yield for last three steps. MS (ES-): 441;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.31 (s, 3H), 3.25 (bs, 4H),
3.39 (bs, 4H), 4.15 (s, 2H), 7.42 (d, J=7.8 Hz, 1H), 7.65-7.71 (m,
4H), 7.78-7.82 (m, 2H), 11.78 (s, 1H). Anal. Calcd. for
C.sub.20H.sub.19N.sub.6O.sub.3S. 1.25CH.sub.3SO.sub.3H.2.4H.sub.2O:
C, 42.43; H, 4.87; N, 13.87; S, 11.91. Found: C, 42.13; H, 4.79; N,
13.48; S, 11.89.
Preparation of
8-[4-(toluene-4-sulfonyl)-piperazin-1-ylmethyl]-1,2,79-tetraaza-phenalen--
3-one, 52
[0197] Synthesized using tolune-4-sulfonyl chloride and compound
47A for General Procedure G. A mesylate salt of 52 was prepared.
38% overall yield for last three steps. MS (ES-): 438; .sup.1H NMR
(400 MHz, DMSO-d.sub.6): 2.36 (s, 3H), 2.45 (s, 3H), 3.20 (bs, 4H),
3.46 (bs, 4H), 4.22 (s, 2H), 7.43 (d, J=7.8 Hz, 1H), 7.54 (d, J=7.8
Hz, 2H), 7.68-7.81 (m, 4H), 11.90 (s, 1H). Anal. Calcd. for
C.sub.21H.sub.22N.sub.6O.sub.3S. 1.3C.sub.11SO.sub.3H. 4.0H.sub.2O:
C, 42.25; H, 5.58; N, 13.22; S, 11.61. Found: C, 42.63; H, 5.53; N,
13.40; S, 11.90.
Preparation of
8-(4-benzenesulfonyl-piperazin-1-ylmethyl)-2,9-dihydro-1,2,7,9-tetraaza-p-
henalen-3-one, 53
[0198] Synthesized using benzensulfonyl chloride and compound 47A
for General Procedure G. A mesylate salt of 53 was prepared. 30%
overall yield for last three steps. MS (ES-): 438; .sup.1H NMR (400
MHz, DMSO-d.sub.6): 2.70 (s, 3H), 3.36 (bs, 4H), 3.51 (bs, 4H),
4.14 (s, 2H), 7.11 (d, J=8.0 Hz, 1H), 7.40-7.70 (m, 7H), 11.90 (s,
1H). Anal. Calcd. for C.sub.20H.sub.20N.sub.6O.sub.3S.
1.2CH.sub.3SO.sub.3H.2.5H.sub.2O.0.08N.sub.2H.sub.4: C, 43.36; H,
5.17; N, 14.67; S, 12.01. Found: C, 43.00; H, 5.17; N, 15.05; S,
12.40.
Preparation of
8-[4-(3-trifluoromethyl-benzenesulfonyl)-piperazin-1-ylmethyl]-2,9-dihydr-
o-1,2,7,9-tetraaza-phenalen-3-one, 54
[0199] Synthesized using 3-trifluoro-benzensulfonyl chloride and
compound 47A for General Procedure G. A mesylate salt of 54 was
prepared. 15% overall yield for last three steps. MS (ES-): 438;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.32 (s, 3H), 3.26-3.35 (m,
8H), 4.10 (s, 2H), 7.43 (d, J 8.0 Hz, 1H), 7.69-7.80 (m, 2H),
7.98-8.26 (m, 4H), 11.92 (s, 1H)
[0200] Anal. Calcd. for C.sub.21H.sub.19F.sub.3N.sub.6O.sub.3S.
1.3CH.sub.3SO.sub.3H.2.0H.sub.2O: C, 40.99; H, 4.35; N, 12.86; S,
11.29. Found: C, 40.71; H, 4.60; N, 12.68; S, 11.50.
Preparation of
8-[4-(4-chloro-benzenesulfonyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2,7,9-
-tetraaza-phenalen-3-one, 55
[0201] Synthesized using 4-chlorobenzensulfonyl chloride and
compound 47A for General Procedure G. A mesylate salt of 55 was
prepared. 15% overall yield for last three steps. MS (ES-): 458;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.31 (s, 3H), 3.18 (bs, 4H),
3.40 (bs, 4H), 3.98 (s, 2H), 7.43 (d, J 7.7 Hz, 1H), 7.68-7.84 (m,
5H), 11.90 (s, 1H). Anal. Calcd. for
C.sub.20H.sub.19C.sub.1N.sub.6O.sub.3S.
1.3CH.sub.3SO.sub.3H.2.0H.sub.2O: C, 41.27; H, 4.59; N, 13.56; S,
11.90. Found: C, 41.07; H, 4.66; N, 13.30; S, 11.89.
Preparation of
8-[4-(4-fluoro-benzenesulfonyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2,7,9-
-tetraaza-phenalen-3-one, 56
[0202] Synthesized using 4-fluorobenzensulfonyl chloride and
compound 47A for General Procedure G. An HCl salt of 56 was
prepared. 42% overall yield for last three steps. MS (ES-): 441;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.31 (s, 3H), 3.18 (bs, 4H),
3.40 (bs, 4H), 3.98 (s, 2H), 7.43 (d, J=7.8 Hz, 1H), 7.68-7.84 (m,
5H), 11.90 (s, 1H).
Preparation of
8-[4-(4-isopropyl-benzenesulfonyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2,-
7,9-tetraaza-phenalen-3-one, 57
[0203] Synthesized using 4-isopropylbenzensulfonyl chloride and
compound 47A for General Procedure G. A mesylate salt of 57 was
prepared. 22% overall yield for last three steps. MS (ES-): 465;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.26 (d, J=6.8 Hz, 6H), 2.33
(s, 3H), 3.01-3.05 (m, 1H), 3.16-3.32 (m, 10H), 7.41 (d, J 8.1 Hz,
1H), 7.59 (d, J 8.6 Hz, 2H), 7.68-7.80 (m, 4H), 11.89 (s, 1H).
Anal. Calcd. for C.sub.23H.sub.26N.sub.6O.sub.3S.
1.35CH.sub.3SO.sub.3H. 1.75H.sub.2O.0.1N.sub.2H.sub.4: C, 46.35; H,
5.64; N, 13.76; S, 11.94. Found: C, 46.01; H, 5.62; N, 13.80; S,
12.33.
Preparation of
8-[4-(4-tert-butyl-benzenesulfonyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2-
,7,9-tetraaza-phenalen-3-one, 58
[0204] Synthesized using 4-tertbutylbenzensulfonyl chloride and
compound 47A for General Procedure G. A mesylate salt of 58 was
prepared. 23% overall yield for last three steps. MS (ES-): 480;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.25 (s, 9H), 2.21 (s, 3H),
3.05-3.15 (m, 8H), 3.99 (bs, 2H), 7.32 (d, J=8.1 Hz, 1H), 7.59-7.72
(m, 6H), 11.81 (s, 1H). Anal. Calcd. for
C.sub.24H.sub.28N.sub.6O.sub.3S. 1.5CH.sub.3SO.sub.3H.2.75H.sub.2O:
C, 45.42; H, 5.90; N, 12.46; S, 11.89. Found: C, 45.23; H, 5.76; N,
12.84; S, 12.17.
Preparation of
8-[4-(4-isopropyl-benzenesulfonyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro--
1,2,7,9-tetraaza-phenalen-3-one, 59
[0205] Synthesized using 4-isopropylbenzensulfonyl chloride and
compound 47B for General Procedure G. 22% overall yield for last
two steps. MS (ES-): 479; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.23
(d, 6H), 1.79 (m, 2H), 2.40-2.55 (m, 4H), 2.71-2.90 (m, 4H), 3.00
(m, 1H), 3.48 (s, 2H), 7.48 (m, 3H), 7.73 (m, 4H), 11.80 (s, 1H).
Anal. Calcd. for C.sub.24H.sub.28N.sub.6O.sub.3S: C, 59.98; H,
5.87; N, 17.49; S, 6.67. Found: C, 60.02; H, 5.85; N, 17.55; S,
6.52.
Preparation of
8-[4-(4-chloro-benzenesulfonyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-1,2-
,7,9-tetraaza-phenalen-3-one, 60
[0206] Synthesized using 4-chloro-benzenesulfony chloride and
compound 47B for General Procedure G. 8% overall yield for last
three steps. MS (ES-): 472; .sup.1H NMR (400 MHz, DMSO-d.sub.6):
1.80 (m, 2H), 2.73-2.78 (m, 4H), 3.50 (m, 4H), 3.69 (s, 2H), 7.45
(d, J=8.2 Hz, 1H), 7.71-7.83 (m, 6H), 10.95 (s, br, 1H), 11.76 (s,
1H). A mesylate salt of 60 was prepared. .sup.1H NMR (400 MHz,
D.sub.2O): 1.92 (m, 2H), 2.73 (s, 5H), 3.50-3.77 (m, 8H), 4.36 (s,
2H), 7.49 (d, J=7.2 Hz, 1H), 7.75 (t, J=8.1 Hz, 2H), 7.78-7.93 (m,
4H). Anal. Calcd. for C.sub.21H.sub.21C.sub.1N.sub.6O.sub.3S.
1.61CH.sub.3SO.sub.3H: C, 39.57; H, 4.99; N, 12.25; S, 12.20.
Found: C, 39.50; H, 5.29; N, 12.57; S, 12.47.
Preparation of
8-[4-(3-fluoro-benzenesulfonyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-1,2-
,7,9-tetraaza-phenalen-3-one, 61
[0207] Synthesized using 3-fluoro-benzenesulfony chloride and
compound 47B for General Procedure G. 16% overall yield for last
two steps. MS (ES+): 457; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.79
(m, 2H), 2.70-2.81 (m, 4H), 3.26-3.40 (m, 4H), 3.48 (s, 2H), 7.45
(d, J 7.3 Hz, 1H); 7.55-7.74 (m, 6H), 11.10 (s, br, 1H), 11.75 (s,
1H). Anal. Calcd. for C.sub.21H.sub.21FN.sub.6O.sub.3S.
1.15H.sub.2O: C, 52.85; H, 4.92; N, 17.61; S, 6.72. Found: C,
52.88; H, 4.93; N, 17.43; S, 6.48.
Preparation of
8-[4-(4-methoxy-benzenesulfonyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-1,-
2,7,9-tetraaza-phenalen-3-one, 62
[0208] Synthesized using 4-methoxy-benzensulfonyl chloride and
compound 47B for General Procedure G. 21% overall yield for last
two steps. MS (ES+): 469; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.78
(m, 2H), 2.72-2.79 (m, 4H), 3.30-3.39 (m, 4H), 3.48 (s, 2H), 3.84
(s, 3H), 7.14 (d, J 8.2 Hz, 2H), 7.48 (d, J 8.1 Hz, 1H), 7.63 (d, J
7.2 Hz, 1H); 7.09-7.22 (m, 3H), 11.10 (s, br, 1H), 11.80 (s, 1H).
Anal. Calcd. for C.sub.22H.sub.24N.sub.6O.sub.4S. 1.0H.sub.2O: C,
54.31; H, 5.39; N, 17.27; S, 6.59. Found: C, 54.38; H, 5.34; N,
17.28; S, 6.19.
Preparation of
8-[4-(4-tert-butyl-benzenesulfonyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-
-1,2,7,9-tetraaza-phenalen-3-one, 63
[0209] Synthesized using 4-t-butyl-benzenesulfony chloride and
compound 47B for General Procedure G. 14% overall yield for last
two steps. MS (ES-): 493; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.31
(s, 9H), 1.79 (m, 2H), 2.73-2.86 (m, 4H), 3.26-3.41 (m, 4H), 3.48
(s, 2H), 7.45 (d, J=8.6 Hz, 1H), 7.62-7.76 (m, 6H), 11.20 (s, br,
1H), 11.80 (s, 1H). Anal. Calcd. for
C.sub.25H.sub.30N.sub.6O.sub.3S: C, 60.71; H, 6.11; N, 16.99; S,
6.48. Found: C, 60.78; H, 6.10; N, 17.08; S, 6.36.
Preparation of
8-[4-(4-amino-benzenesulfonyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-1,2,-
7,9-tetraaza-phenalen-3-one, 64
[0210] Synthesized using 4-nitro-benzenesulfony chloride and
compound 47B for General Procedure G. 14% overall yield for last
two steps. MS (ES-): 452; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.76
(m, 2H), 2.71-2.79 (m, 4H), 3.21-3.31 (m, 4H), 3.46 (s, 2H), 6.01
(s, 2H), 6.64 (d, J=8.6 Hz, 2H), 7.39 (d, J=8.6 Hz, 2H), 7.48 (d,
J=8.0 Hz, 1H), 7.63 (d, J=7.9 Hz, 1H), 7.74 (t, J=7.8 Hz, 1H),
11.10 (s, br, 1H), 11.75 (s, 1H). Anal. Calcd. for
C.sub.21H.sub.23N.sub.7O.sub.3S. 0.5H.sub.2O: C, 54.53; H, 5.23; N,
21.20; S, 6.93. Found: C, 54.50; H, 5.24; N, 20.84; S, 6.74.
Preparation of
8-[4-(biphenyl-4-sulfonyl)-[1,4]diazepan-1-ylmethyl]-2,9-dihydro-1,2,7,9--
tetraaza-phenalen-3-one, 65
[0211] Synthesized using biphenyl-4-sulfony chloride and compound
47B for General Procedure G. 10% overall yield for last two steps.
MS (ES-): 513; .sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.82 (m, 21),
2.73-2.83 (m, 4H), 3.29-3.41 (m, 41), 3.48 (s, 21), 7.47-7.53 (m,
4H), 7.62 (d, J=8.1 Hz, 1H), 7.68-7.78 (m, 3H), 7.82-7.93 (m, 4H),
11.00 (s, br, 1H), 11.75 (s, 1H). Anal. Calcd. for
C.sub.27H.sub.26N.sub.6O.sub.3S. 2.3H.sub.2O: C, 58.32; H, 5.55; N,
15.11; S, 5.77. Found: C, 58.24; H, 4.89; N, 15.10; S, 5.79.
Preparation of
8-[4-(4-amino-benzenesulfonyl)-piperazin-1-ylmethyl]-2,9-dihydro-1,2,7,9--
tetraaza-phenalen-3-one, 66
[0212] Synthesized using 4-nitrobenzene sulfonyl chloride and
compound 47A for General Procedure G. A mesylate salt of 66 was
prepared. 35% overall yield for last three steps. MS (ES-): 438;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 2.32 (s, 3H), 3.13 (bs, 4H),
3.42 (bs, 4H), 4.18 (s, 2H), 6.71 (d, J=8.8 Hz, 2H), 7.40-7.43 (m,
3H), 7.70-7.80 (m, 2H), 11.20 (s, br, 1H), 11.92 (s, 1H). Anal.
Calcd. for C.sub.20H.sub.21N.sub.7O.sub.3S.
1.3CH.sub.3SO.sub.3H.2.75H.sub.2O: C, 41.67; H, 5.20; N, 15.97; S,
12.01. Found: C, 41.76; H, 5.25; N, 15.92; S, 12.22.
##STR00026##
General Procedure L to prepare compounds 71. To a stirring solution
of 70 (1.0 eq) in THF under nitrogen was added TEA (1 mL, excess)
and either sulfonyl chloride or acid chloride (1.2 eq). The
reaction was allowed to stir for four hours after which time it was
evaporated and extracted with CH.sub.2Cl.sub.2/H.sub.2O, dried and
condensed. Crude material was further purified via column
chromatography using 9:1CH.sub.2Cl.sub.2/MeOH to afford
analytically pure products 71.
Example 3
Preparation of
(3-oxo-2,9-dihydro-3H-1,2,7,9-tetraaza-phenalen-8-ylmethyl)-carbamic
acid tert-butyl ester, 69
[0213] Procedure H to prepare
2-aminomethyl-4-oxo-3,4-dihydro-quinazoline-5-carboxylic acid
methyl ester, 67. To a solution of 25 mL of 7N NH.sub.3 (large
excess) in MeOH at 0.degree. C. was added compound 4 (1.0 g, 4.0
mmol) in a sealed tube. The mixture was then heated to 60.degree.
C. for 4 hours. The mixture was evaporated to dryness, dissolved
and re-evaporated in 2.times.50 mL of CH.sub.2Cl.sub.2. Product was
used as is without further purification.
[0214] Procedure I to prepare
2-(tert-butoxycarbonylamino-methyl)-4-oxo-3,4-dihydro-quinazoline-5-carbo-
xylic acid methyl ester, 68. To a solution of 50 mL
CH.sub.2Cl.sub.2 of with 2 mL of TEA (excess), catalytic DMAP and
compound 67 (from Procedure H) was added boc anhydride (2.6 g, 3
eq) at room temperature. Reaction was allowed to stir for 60
minutes, during which time all solids went into solution. The
solution was evaporated to dryness and purified via column
chromatography using CH.sub.2Cl.sub.2 and 5% MeOH to afford 0.5 g
of analytically pure compound, 68.
[0215] Procedure J to prepare 69.5 g of compound 68 was dissolved
in 10 mL of hydrazine monohydrate and 25 mL of ethanol. The mixture
was refluxed for four hours until no starting material was detected
by TLC. Reaction was cooled, poured over 100 mL of cold water and
extracted with 2.times.25 mL of EtOAc. Organic layers were dried
with brine and then magnesium sulfate. Purified via column
chromatography using 9:1 CH.sub.2Cl.sub.2/MeOH to afford 2.7 g of
analytically pure compound 69. MS (ES-): 314; .sup.1H NMR (400 MHz,
CDCl.sub.3): 1.45 (s, 9H), 3.90 (s, 2H), 6.15 (bs, 1H), 6.94-7.30
(m, 3H), 12.38-12.43 (m, br, 2H). Anal. Calcd. for
C.sub.15H.sub.17N.sub.5O.sub.3.0.2H.sub.2O: C, 56.49; H, 5.50; N,
21.96. Found: C, 56.61; H, 5.60; N, 21.85.
Preparation of
8-aminomethyl-2,9-dihydro-1,2,7,9-tetraaza-phenalen-3-one, 70
[0216] Procedure K to prepare 70. 250 mg of compound 69 was
dissolved in 10 mL of CH.sub.2Cl.sub.2 along with 4 mL of TFA. The
reaction was allowed to stir at room temperature overnight
resulting in a heavy white precipitate, which was filtered off and
washed with CH.sub.2Cl.sub.2 and dried under vacuum to afford a
quantitative yield of analytically pure material, a TFA salt of
compound 70. MS (ES+): 216; .sup.1H NMR (400 MHz, D.sub.2O): 3.97
(s, 2H), 6.91 (d, J 8.2 Hz, 1H), 7.23 (d, J 7.8 Hz, 1H), 7.43 (t, J
8.0 Hz, 1H). Anal. Calcd. for
C.sub.10H.sub.9N.sub.5O.1.3CF.sub.3COOH. 0.2H.sub.2O: C, 41.27; H,
2.86; N, 19.10. Found: C, 41.00; H, 3.04; N, 19.25.
Preparation of
4-methyl-N-(3-oxo-2,9-dihydro-3H-1,2,7,9-tetraaza-phenalen-8-ylmethyl)-be-
nzenesulfonamide, 72
[0217] Synthesized using 4-methylbenzene sulfonyl chloride and
compound 70 for General Procedure L. 20% yield for compound 72. MS
(ES-): 368; .sup.1H NMR (400 MHz, CDCl.sub.3): 2.27 (s, 3H), 3.93
(s, 2H), 7.28-7.43 (m, 3H), 7.67-7.78 (m, 4H), 11.43 (s, 1H), 11.83
(s, 1H). Anal. Calcd. for C.sub.17H.sub.15N.sub.5O.sub.3S: C,
55.27; H, 4.09; N, 18.96; S, 8.68. Found: C, 54.93; H, 4.09; N,
18.63; S, 8.33.
Preparation of
N-(3-oxo-2,9-dihydro-3H-1,2,7,9-tetraaza-phenalen-8-ylmethyl)-benzenesulf-
onamide, 74
[0218] Synthesized using benzene sulfonyl chloride and compound 70
for General Procedure L. 25% yield for compound 74. MS (ES-): 354;
.sup.1H NMR (400 MHz, CDCl.sub.3): 3.95 (d, J 5.0 Hz, 2H), 7.44 (d,
J 7.8 Hz, 1H), 7.45-7.60 (m, 3H), 7.67-7.77 (m, 2H), 7.91-7.93 (m,
2H), 8.24 (t, J=8.8 Hz, 1H), 11.24 (s, 1H), 11.83 (s, 1H). Anal.
Calcd. for C.sub.16H.sub.13N.sub.5O.sub.3S. 1.0H.sub.2O: C, 51.47;
H, 4.05; N, 18.76; S, 8.59. Found: C, 51.17; H, 4.20; N, 18.73; S,
8.31.
Preparation of
N-(3-oxo-2,9-dihydro-3H-1,2,7,9-tetraaza-phenalen-8-ylmethyl)-acetamide,
75
[0219] Synthesized using acetic anhydride and compound 70 for
General Procedure L. 22% yield for compound 75. MS (ES-): 256;
.sup.1H NMR (400 MHz, DMSO-d.sub.6): 1.91 (s, 3H), 4.05 (s, 2H),
7.32 (d, J=8.1 Hz, 1H), 7.49-7.81 (m, 2H), 8.40 (t, J=8.3 Hz, 1H).
11.25 (s, 1H), 11.75 (s, 1H). Anal. Calcd. for
C.sub.12H.sub.11N.sub.5O.sub.2.0.5H.sub.2O: C, 54.13; H, 4.54; N,
26.30. Found: C, 54.14; H, 4.52; N, 26.00.
Preparation of
4-nitro-N-(3-oxo-2,9-dihydro-3H-1,2,7,9-tetraaza-phenalen-8-ylmethyl)-ben-
zamide, 76
[0220] Synthesized using 4-nitro-benzoyl chloride and compound 70
for General Procedure L. 25% yield for compound 76. MS (ES-): 363;
.sup.1H NMR (400 MHz, CDCl.sub.3): 3.99 (s, 2H), 7.18-7.20 (m, 1H),
7.34-7.38 (m, 1H), 7.75-7.90 (m, 2H), 8.20-8.32 (m, 2H), 8.40-8.48
(m, 3H).
In Vitro PARP Inhibitory Potency--IC.sub.50
[0221] A convenient method to determine IC.sub.50 of a PARP
inhibitor compound is a PARP assay using purified recombinant human
PARP from Trevigan (Gaithersburg, Md.), as follows: The PARP enzyme
assay is set up on ice in a volume of 100 microliters consisting of
100 mM Tris-HCl (pH 8.0), 1 mM MgCl.sub.2, 28 mM KCl, 28 mM NaCl,
3.0 .mu.g/ml of DNase I-activated herring sperm DNA (Sigma, Mo.),
30 micromolar [.sup.3H]nicotinamide adenine dinucleotide (62.5
mci/mmole), 15 micrograms/ml PARP enzyme, and various
concentrations of the compounds to be tested. The reaction is
initiated by adding enzyme and incubating the mixture at 25.degree.
C. After 2 minutes of incubation, the reaction is terminated by
adding 500 microliters of ice cold 30% (w/v) trichloroacetic acid.
The precipitate formed is transferred onto a glass fiber filter
(Packard Unifilter-GF/C) and washed three times with 70% ethanol.
After the filter is dried, the radioactivity is determined by
scintillation counting. The compounds of this invention were found
to have potent enzymatic activity in the range of a few nanomolar
to 20 micromolar in IC.sub.50 in this inhibition assay.
[0222] Using the PARP assay described above, approximate IC50
values were obtained for the following compounds:
TABLE-US-00001 TABLE I Compound Structure IC50 nM 7 ##STR00027## 35
8 ##STR00028## 23 9 ##STR00029## 35 10 ##STR00030## 19 11
##STR00031## 6 12 ##STR00032## 9 13 ##STR00033## 12 14 ##STR00034##
18 15 ##STR00035## 32 16 ##STR00036## 21 17 ##STR00037## 20 18
##STR00038## 17 19 ##STR00039## 18 20 ##STR00040## 35 21
##STR00041## n/a 22 ##STR00042## 35 23 ##STR00043## 39 24
##STR00044## 51 25 ##STR00045## 26 26 ##STR00046## 41 27
##STR00047## 43 28 ##STR00048## 29 30 ##STR00049## 13 31
##STR00050## 28 32 ##STR00051## 31 33 ##STR00052## n/a 34
##STR00053## 49 35 ##STR00054## 44 36 ##STR00055## 19 37
##STR00056## 12 38 ##STR00057## 20 39 ##STR00058## 15 40
##STR00059## 39 41 ##STR00060## 42 42 ##STR00061## 13 43
##STR00062## 38 44 ##STR00063## 21 45 ##STR00064## 49 46
##STR00065## 11 50 ##STR00066## 52 51 ##STR00067## 15 52
##STR00068## 21 53 ##STR00069## 23 54 ##STR00070## 14 55
##STR00071## 18 56 ##STR00072## 27 57 ##STR00073## 17 58
##STR00074## 13 59 ##STR00075## 23 60 ##STR00076## 24 61
##STR00077## 27 62 ##STR00078## 22 63 ##STR00079## 19 64
##STR00080## 15 65 ##STR00081## 22 66 ##STR00082## 45 69
##STR00083## 47 72 ##STR00084## 171 74 ##STR00085## 23 75
##STR00086## 30 76 ##STR00087## 10
Efficacy In Vivo for Compound 13
1) Mouse Intracranial Model of B16 Melanoma:
[0223] The murine melanoma cell line B16 of C.sub.57BL/6J
(H-2.sup.b/H-2.sup.b) origin was cultured in RPMI-1640 containing
10% fetal calf serum (Invitrogen, Milan, Italy), 2 mM L-glutamine,
100 units/ml penicillin and 100 .mu.g/ml streptomycin (Flow
Laboratories, Mc Lean, Va.), at 37.degree. C. in a 5% CO.sub.2
humidified atmosphere. TMZ was provided by Schering-Plough Research
Institute (Kenilworth, N.J.). Compound 13 was dissolved in 70 mM
PBS without potassium.
[0224] For intracranial transplantation, cells (10.sup.4 in 0.03 ml
of RPMI-1640) were injected intracranially (ic) through the
center-middle area of the frontal bone to a 2 mm depth, using a 0.1
ml glass microsyringe and a 27-gauge disposable needle. Murine
melanoma B16 cells (10.sup.4) were injected ic into male B6D2F1
(C.sub.57BL/6.times.DBA/2) mice. Before tumor challenge, animals
were anesthetized with ketamine (100 mg/kg) and xylazine (5 mg/kg)
in 0.9% NaCl solution (10 ml/kg/ip). Histological evaluation of
tumor growth in the brain was performed 1-5 days after tumor
challenge, in order to determine the timing of treatment.
[0225] The compound 13 was administered per os 15 min before TMZ.
Control mice were always injected with drug vehicles. In
tumor-bearing mice treatment started 48 h after challenge, when
tumor infiltration in the surrounding brain tissue was
histologically evident. Mice were treated with compound 13 by oral
gavage once a day for five days, at the doses of 10 mg/kg.
[0226] In tumor-bearing mice, treatment started on day 2 after
challenge, when tumor infiltration in the surrounding brain tissue
was histologically evident. Mice were treated daily with compound
13 plus TMZ for 5 days and monitored for mortality for 90 days.
Median survival times (MST) were determined and the percentage of
increase in lifespan (ILS) was calculated as: {[MST (days) of
treated mice/MST (days) of control mice]-1}.times.100. Efficacy of
treatments was evaluated by comparing survival curves between
treated and control groups.
[0227] All procedures involving mice and care were performed in
compliance with national and international guidelines (European
Economy Community Council Directive 86/109, OLJ318, Dec. 1, 1987
and NIH Guide for care and use of laboratory animals, 1985).
[0228] Survival curves were generated by Kaplan-Meier product-limit
estimate and statistical differences between the various groups (8
animals/group) were evaluated by log-rank analysis with Yates
correction (software Primer of Biostatistics, McGraw-Hill, New
York, N.Y.). Statistical significance was determined at a p=0.05
level. Differences were considered statistically significant when
P<0.05.
[0229] The results indicate oral administration of 10 mg/kg
compound 13 significantly increased the survival time of mice
treated with compound 13+TMZ combination and was significantly
higher than that observed in animals receiving TMZ as single agent
(P<0.0001). No significant differences in survival times were
observed between control and TMZ treated groups (FIG. 1).
2) Intracranial Xenograft Model of SJGBM2 Glioma in Mice:
[0230] The compound 13 was tested in the intracranial xenograft
model of SJGBM2 glioma in mice (Tentori, et al. Clin. Cancer Reser.
2003, 9, 5370). For this purpose compound 13 was given once at 15
min pre-TMZ at 10 mg/kg, po.
[0231] A dose of 10 mg/kg compound 13 was found to be efficacious
(FIG. 2). Its combination with TMZ increased MTS from 22.5 d (TMZ
alone) to 25 d (P=0.002).
Efficacy In Vivo for Compound 37
1) Mouse Intracranial Model of B16 Melanoma:
[0232] The experiment was performed as described above for Compound
13. It was investigated whether oral administration of Compound 37
(5 mg/kg or 12.5 mg/kg), might increase the efficacy of TMZ against
B16 melanoma growing at the CNS site. In mice bearing B16 melanoma,
the results indicated that the mean survival time of the groups
treated with Compound 37 12.5 mg/kg+TMZ combination was
significantly higher than that observed in animals receiving TMZ as
single agent (FIG. 3).
2) Intracranial Xenograft Model of SJGBM2 Glioma in Mice:
[0233] The efficacy of Compound 37 was then investigated using an
orthotopic model of a human glioblastoma multiforme xenograft
(SJGBM2) in nude mice. The response of SJGBM2 to TMZ, used as
single agent or in combination with Compound 37 (10 mg/kg or 20
mg/kg) for five days or in combination with Compound 37 (MGI25036)
10 mg/kg for five days followed by a 14-day treatment with Compound
37 100 mg/kg as single agent, is shown in FIG. 4. The results
indicate that oral administration of Compound 37 (10 mg/kg or 20
mg/kg)+TMZ significantly prolonged survival of tumor bearing mice
with respect to controls or to animals treated with TMZ. It should
be noted that in this tumor model TMZ was ineffective. Treatment
with 10 mg/kg Compound 37+TMZ for five days followed by a high dose
of Compound 37 (100 mg/kg) for 14 days significantly increased
animal survival with respect to 10 mg/kg Compound 37+TMZ for five
days.
3) Enhancement of Radiation Treatment of Head and Neck Squamous
Cell Carcinoma
[0234] Human HNSCC cell line JHU012 was used, having been
previously genetically characterized and originally derived at the
Johns Hopkins University Head and Neck Laboratories from human
tumor explants. The cell line was maintained in RPMI 1640 medium
with 10% fetal bovine serum and 1% penicillin/streptomycin at 5%
CO.sub.2 in 37.degree. C. humidified incubators. Experiments were
performed on 6-week-old male BALB/c nude mice nu/nu. The animals
were randomly divided into the following treatment groups: Group
1--controls, Group 2--Radiation alone (2 Gray (gy)/day for 2 days),
Group 3--100 mg/kg Compound 37 alone orally (PO) qdx17, Group 4--30
mg/kg Compound 37 PO+Radiation, Group 5--100 mg/kg Compound 37
PO+Radiation, with each group consisting of 8 mice. Mice were
anesthetized by intraperitoneal injection of 3-5 mL
tribromoethanol. Tumors were established at the right flank by
subcutaneous injection of 1.times.10.sup.7 cells. Fourteen days
post cell injection tumors were surgically exposed and measured in
3 dimensions using calipers. Compound 37 was then dosed orally in
treatment Groups 3-5. In Groups 4 and 5, animals received Compound
37 15 minutes prior to radiation (2 gy/day for 2 days). At day 31
post tumor cell inoculation, tumors were again surgically exposed
and measured in 3 dimensions using calipers A significant
inhibition of tumor growth was observed in Group 5 treated with 100
mg/kg orally administered Compound 37+Radiation (tumor volume at
end of experiment=209.04 mm.sup.3) compared to the control Group 1
(tumor volume at end of experiment=585.9 mm.sup.3 p<0.01). (FIG.
5) Compound 37 at 30 mg/kg in combination with radiation had no
significant effect on tumor growth inhibition compared to radiation
alone (FIG. 5). In addition, 100 mg/kg Compound 37 PO qdx17 alone
had no significant effect on tumor growth inhibition compared to
vehicle controls (FIG. 5). This indicates an enhanced effect when
the higher dose of Compound 37 was combined with radiation as
opposed to either treatment modality alone.
4) Effect of Compound 37 on Tumor Growth in Mice Bearing BRCA-1
Deficient Tumors
[0235] 1.times.10.sup.6 BRCA-1 null cells were injected
subcutaneously on the right flank of female nu/nu mice (6-7 weeks
old; Harlan Sprague Dawley, Indianapolis Ind.). After approximately
10-14 days, the tumors were approximately 100 mm.sup.3. Mice were
sorted into groups so that mean tumor size was similar among groups
with minimum standard deviations. Dosing started the day after
sorting and tumor volume was monitored three times per week. Tumors
were measured in two diameters and volume calculated by
(1.times.w).sup.2/2. Mice were removed from the study when tumors
reached 1500 mm.sup.3. "Time to Endpoint" or TTE (the number of
days it takes for the tumor to reach 1500 mm.sup.3 or greater) is
the endpoint of the study. Compound 37 was weighed out every 2-3
days and solubilized in sterile bottled water (J. T. Baker,
Ultrapure Bioreagent 4221-02) to 10 mg/ml. The compound was dosed
orally, daily for 28 days from start of the study--day 1. A
positive control was utilized, using a well known PARP inhibitor
shown to be effective as a stand alone agent in the BRCA models
(Bryant et al). The positive control agent was dosed at 25 mg/kg IP
qdx5 from start of experiment. 100 mg/kg Compound 37 was effective
in significantly retarding tumor growth in the BRCA-1 null model
both times tested. When the dosing of Compound 37 was stopped at
day 28, the tumors start to grow approximately 10-14 days later.
Compound 37 not only significantly delayed tumor growth compared to
vehicle controls but also delayed tumor growth compared to the
positive control (p<0.05) in both experiments.
[0236] A study was conducted to compare the bioavailablity and
brain plasma levels of various mammals administered with the
disclosed compounds and a similar prior art compound. The prior art
compound has the following formula:
##STR00088##
The Comparative Study was Conducted as Follows:
[0237] PARP inhibitors in water solutions were dosed either by
bolus (<1 min) intravenous injection, or by oral gavage. For
dogs, intravenous and oral dosing was performed in a crossover
design with a one-week washout period between dose routes. The
screening dose was 30 mg/kg for each compound. For mice, three
animals per time point were sacrificed by CO.sub.2 asphyxia and
blood collected by cardiac puncture. For rats and dogs, serial
blood samples were taken at various time points from the indicated
number of animals. For rats, the volume of blood sampled was
immediately replaced with 2.times. volume of 1:1 donor rat
blood:heparinized saline. The blood samples were transferred to
heparinized containers, briefly mixed, and stored on ice until
centrifugation to prepare plasma. The plasma was transferred to
fresh containers and stored at .ltoreq.-70.degree. C. until
bioanalysis. In some cases brains or tumor tissue were collected
after sacrifice and stored at .ltoreq.-70.degree. C. until
bioanalysis.
[0238] Plasma samples were processed by precipitation with
acetonitrile, evaporation and reconstitution. Brain and tumor
tissue samples were homogenized with phosphate buffered saline, pH
7.4, precipitated with acetonitrile, followed by evaporation and
reconstitution. The reconstituted samples were analyzed vs. matrix
calibration standards by LC--MS/MS. The bioanalytical method
performance was verified by the performance of quality control
samples. Generally, the plasma lower limit of quantitation was 5
ng/mL. Tissue lower limits of quantitation depended on the degree
of dilution during homogenization, but usually were 15 to 20
ng/g.
[0239] Plasma, brain, and tumor concentration data were processed
by noncompartmental pharmacokinetic analysis using WinNonlin
Professional Version 4.1. AUC was calculated using the Linear/Log
rule. Time points for the Lambda Z phase were selected by visual
inspection. The slopes of terminal phases were calculated by
unweighted linear regression.
[0240] Selective PARP inhibitors were tested for basic plasma and
tissue pharmacokinetic properties in mice, rats, and dogs. After
assessment, this family compounds appear to be orally bioavailable
in all species and to penetrate brain and tumor tissue. Table 1
summarizes the oral bioavailability (PO) for compounds 8, 13, 36
and 37 and the comparative compound in mice and rats and
brain/plasma ratio (B/P) for these five compounds in mice and
rats.
[0241] The results of the comparative study are summarized in Table
II. The results show that while the prior art compound has good
bioavailability the prior art compound has a ratio of brain to
plasma levels that is very low. Unexpectedly, the disclosed
compounds of Formula (I) have a good ratio of brain to plasma level
compared to the prior art compound. These results show the
disclosed compounds are unexpectedly available to the central
nervous system where needed for therapeutic benefit as compared to
the prior art compound.
TABLE-US-00002 TABLE II Comparison of Bioavailabity (PO) and Ratio
of Brain to Plasma levels (B/P) for selected compounds of Formula
(I) relative to a related prior art compound. B/P in Compound PO in
mice mice PO in rats B/P in rats ##STR00089## Comparative Compound
77% <5% 77% <5% ##STR00090## 8 49% 49% 58% 40% ##STR00091##
13 61% 46% 51% 42% ##STR00092## 36 75% 30-64% 50% 71-117%
##STR00093## 37 81% 26% 45% 36%
[0242] The invention being thus described, it will be obvious that
the same may be varied in many ways. Such variations are not to be
regarded as a departure from the spirit and scope of the invention,
and all such modifications are intended to be included within the
scope of the following claims.
INCORPORATION BY REFERENCE
[0243] All publications, patents, and pre-grant patent application
publications cited in this specification are herein incorporated by
reference, and for any and all purposes, as if each individual
publication or patent application were specifically and
individually indicated to be incorporated by reference. In the case
of inconsistencies the present invention will prevail.
* * * * *