U.S. patent number RE47,009 [Application Number 15/182,061] was granted by the patent office on 2018-08-28 for hdac inhibitors and therapeutic methods using the same.
This patent grant is currently assigned to THE CHILDREN'S HOSPITAL OF PHILADELPHIA, THE TRUSTEES OF THE UNIVERSITY OF ILLINOIS. The grantee listed for this patent is THE CHILDREN'S HOSPITAL OF PHILADELPHIA, THE TRUSTEES OF THE UNIVERSITY OF ILLINOIS. Invention is credited to Joel Bergman, Kyle Vincent Butler, Wayne W. Hancock, Jay Hans Kalin, Alan Kozikowski.
United States Patent |
RE47,009 |
Kozikowski , et al. |
August 28, 2018 |
**Please see images for:
( Certificate of Correction ) ** |
HDAC inhibitors and therapeutic methods using the same
Abstract
Histone deacetylases inhibitors (HDACIs) and compositions
containing the same are disclosed. Methods of treating diseases and
conditions wherein inhibition of HDAC provides a benefit, like a
cancer, a neurodegenerative disorder, a peripheral neuropathy, a
neurological disease, traumatic brain injury, stroke, hypertension,
malaria, an autoimmune disease, autism, autism spectrum disorders,
and inflammation, also are disclosed.
Inventors: |
Kozikowski; Alan (Chicago,
IL), Kalin; Jay Hans (Chicago, IL), Butler; Kyle
Vincent (Stanford, CA), Bergman; Joel (Chicago, IL),
Hancock; Wayne W. (Philadelphia, PA) |
Applicant: |
Name |
City |
State |
Country |
Type |
THE CHILDREN'S HOSPITAL OF PHILADELPHIA
THE TRUSTEES OF THE UNIVERSITY OF ILLINOIS |
Philadelphia
Urbana |
PA
IL |
US
US |
|
|
Assignee: |
THE CHILDREN'S HOSPITAL OF
PHILADELPHIA (Philadelphia, PA)
THE TRUSTEES OF THE UNIVERSITY OF ILLINOIS (Urbana,
IL)
|
Family
ID: |
46603265 |
Appl.
No.: |
15/182,061 |
Filed: |
June 14, 2016 |
PCT
Filed: |
January 31, 2012 |
PCT No.: |
PCT/US2012/023332 |
371(c)(1),(2),(4) Date: |
December 20, 2013 |
PCT
Pub. No.: |
WO2012/106343 |
PCT
Pub. Date: |
August 09, 2012 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
Issue Date |
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61438435 |
Feb 1, 2011 |
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61443960 |
Feb 17, 2011 |
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61554653 |
Nov 2, 2011 |
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Reissue of: |
13985760 |
Jan 31, 2012 |
9249087 |
Feb 2, 2016 |
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Current U.S.
Class: |
1/1 |
Current CPC
Class: |
C07D
207/323 (20130101); A61K 45/06 (20130101); C07D
235/06 (20130101); C07D 213/56 (20130101); A61K
51/0455 (20130101); A61K 51/04 (20130101); A61P
35/00 (20180101); C07D 473/00 (20130101); C07D
231/12 (20130101); C07D 207/323 (20130101); C07D
209/08 (20130101); A61K 51/0446 (20130101); A61K
51/0453 (20130101); C07D 209/14 (20130101); C07C
259/10 (20130101); C07D 213/56 (20130101); A61K
49/0002 (20130101); C07C 229/52 (20130101); C07D
209/18 (20130101); C07D 235/06 (20130101); C07D
207/08 (20130101); A61K 51/0455 (20130101); C07D
473/00 (20130101); A61K 51/04 (20130101); A61K
51/0459 (20130101); C07D 209/20 (20130101); A61K
51/0459 (20130101); C07D 209/14 (20130101); A61K
51/0453 (20130101); C07D 231/12 (20130101); A61K
51/0446 (20130101); C07C 259/10 (20130101); C07D
209/18 (20130101); A61K 45/06 (20130101); C07C
229/52 (20130101); C07D 209/08 (20130101); A61K
49/0002 (20130101); C07D 207/08 (20130101); C07D
209/20 (20130101); C07C 2601/08 (20170501); C07C
2101/08 (20130101) |
Current International
Class: |
A61K
31/497 (20060101); A61K 31/405 (20060101); A61K
45/06 (20060101); A61K 49/00 (20060101); C07C
259/10 (20060101); A61K 51/04 (20060101); C07C
229/52 (20060101); C07D 207/30 (20060101); C07D
209/04 (20060101); C07D 207/08 (20060101); C07D
235/06 (20060101); C07D 207/323 (20060101); C07D
209/08 (20060101); C07D 209/14 (20060101); C07D
213/56 (20060101); C07D 231/12 (20060101); C07D
473/00 (20060101); C07D 209/18 (20060101); C07D
209/20 (20060101) |
References Cited
[Referenced By]
U.S. Patent Documents
Foreign Patent Documents
|
|
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0341231 |
|
Nov 1989 |
|
EP |
|
0887348 |
|
Dec 1998 |
|
EP |
|
WO-00/18744 |
|
Apr 2000 |
|
WO |
|
WO-01/70675 |
|
Sep 2001 |
|
WO |
|
WO-03/076422 |
|
Sep 2003 |
|
WO |
|
WO-2007/022638 |
|
Mar 2007 |
|
WO |
|
WO-2008/019025 |
|
Feb 2008 |
|
WO |
|
WO-2008/055068 |
|
May 2008 |
|
WO |
|
WO-2009/046804 |
|
Apr 2009 |
|
WO |
|
WO-2009/055917 |
|
May 2009 |
|
WO |
|
WO-2009/073620 |
|
Jun 2009 |
|
WO |
|
WO-2010/100475 |
|
Sep 2010 |
|
WO |
|
WO-2011/011186 |
|
Jan 2011 |
|
WO |
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WO-2011/021209 |
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Feb 2011 |
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WO |
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WO-2012/178208 |
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Dec 2012 |
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WO |
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Other References
Brown et al., Molecular addition compounds. 4. Unusually slow
reaction of tertiary amines with 9-borabicyclo[3.3.1]nonane (9-BBN)
dimer. New, stable, mildly reactive amine complexes of 9-BBN,
Inorg. Chem., 16(12):3090-4 (1977). cited by applicant .
Chai et al., Metal-mediated inhibition is a viable approach for
inhibiting cellular methionine aminopeptidase, Bioorg. Med. Chem.
Lett., 19(24):6862-4 (2009). cited by applicant .
Dell'Erba et al., alpha-Arylation vs. alpha-arylhydrazonylation of
alkyl aryl ketones with arylazo tert-butyl sulfides, Tetrahedron,
49(1):235-42 (1993). cited by applicant .
Esterbauer, Synthese einiger 4-hydroxy-2-alkenale, Monatsheft fur
Chemie, 102:824-7 (1971). cited by applicant .
Faul et al., A new one step synthesis of maleimides by condensation
of glyoxylate esters with acetamides, Tetrahedron Lett., 40:1109-12
(1999). cited by applicant .
Kawase et al., In vitro susceptibility of helicobacter pylori to
trifluoromethyl ketones, Bioorg. Med. Chem. Lett., 9:193-4 (1999).
cited by applicant .
Kitazume et al., A synethetic approach to seven-membered lactones
by the microbial transformation of ynones having a trifluoromethyl
group, J. Fluorine Chem., 30:189-202(1985). cited by applicant
.
Kon-ya et al., Indole derivatives as potent inhibitors of larval
settlement by the barnacle, Balanus amphitrite, Biosci. Biotech.
Biochem., 58(12):2178-81 (1994). cited by applicant .
Lautens et al., Rhodium-catalyzed heck-type coupling of boronic
acids with activated alkenes in an aqueous emulsion, Synthesis,
12:2006-14 (2004). cited by applicant .
Morand et al., The effect of substituted carboxylic acids on
hepatic cholesterogenesis, J. Med. Chem., 7:504-8 (1964). cited by
applicant .
Nelson et al., Synthesis and antiinflammatory activity of some
1,6-methano[10]annuleneacetic acids, J. Med. Chem., 18(6):593- 6
(1975). cited by applicant .
Remiszewski et al., N-hydroxy-3-phenyl-2-propenamides as novel
inhibitors of human histone deacetylase with in vivo antitumor
activity: discovery of
(2E)-N-hydroxy-3-[4-[[(2-hydroxyethyl)[2-(1H-indol-3-yl)ethyl]amino]me-
thyl]phenyl]-2-propenamide (NVP-LAQ824), J. Med. Chem.,
46(21):4609-24 (2003). cited by applicant .
Smith et al., e-Anilino-4-arylmaleimides: potent and selective
inhibitors of glycogen synthase kinase-3 (GSK-3), Bioorg. Med.
Chem. Lett., 11:635-9 (2001). cited by applicant .
Suzuki, Explorative study on isoform-selective histone deacetylase
inhibitors, Chem. Pharm. Bull., 47(9):897-906 (2009). cited by
applicant .
CAS Registry No. 3360-46-1; STN Entry Date Nov. 16, 1984. cited by
applicant .
CAS Registry No. 854631-63-3; STN Entry Date Jul. 12, 2005. cited
by applicant .
CAS Registry No. 864408-96-8; STN Entry Date Oct. 4, 2005. cited by
applicant .
Cossement et al., [Mivazerol and other benzylimidazoles with
alpha-2 adrenergic properties], J. Pharm. Belg., 49(3):206-15
(1994). cited by applicant .
Examination Report No. 2 for Standard Patent Application,
Australian Patent Application No. 2012212323, dated Feb. 16, 2017.
cited by applicant .
Examination Report, European patent application No. EP 12742236.8,
dated Mar. 9, 2017. cited by applicant .
Yee et al., A novel series of selective leukotriene antagonists:
exploration and optimization of the acidic region in
1,6-disubstituted indoles and indazoles, J. Med. Chem.,
33(9):2437-51 (1990). cited by applicant .
Nyandege, Abner, et al., "Further Studies on the Binding of
N1-Substituted Tryptamines at h5-HT6 Receptors," Bioorganic &
Medicinal Chemistry Letters, 2007, vol. 17, pp. 1691-1694. cited by
applicant .
Tang, H., et al., "Novel Inhibitors of Human Histone Deacetylase
(HDAC) Identified by QSAR Modeling of Known Inhibitors, Virtual
Screening, and Experimental Validation," Journal of Chemical
Information Modeling., 2009, vol. 49, No. 2, pp. 461-476. cited by
applicant .
International Search Report in international patent application No.
PCT/US2012/023332, dated Aug. 7, 2012. cited by applicant .
Examination Report, European patent application No. 12742236.8,
dated Apr. 29, 2015. cited by applicant .
Headley et al., Effects of solvents on the tautomerization of
N,N-dimethylglycine, J. Phys. Org. Chem., 10:898-900 (1997). cited
by applicant .
Takagi et al., Synthesis of indolyl aryl 1,2-diketones, Pharm.
Bull., 5(6):617-8 (1957). cited by applicant.
|
Primary Examiner: Railey; Johnny F
Attorney, Agent or Firm: Marshall, Gerstein & Borun
LLP
Government Interests
STATEMENT OF GOVERNMENT INTEREST
This invention was made with government support under NIH Grant No.
P01AI073489 awarded by the National Institutes of Health. The
government has certain rights in the invention.
Parent Case Text
CROSS-REFERENCE TO RELATED APPLICATIONS
This application is the U.S. National phase of PCT/US2012/023332,
filed Jan. 31, 2012, which claims the benefit of U.S. provisional
application No. 61/438,435, filed Feb. 1, 2011, U.S. provisional
application No. 61/443,960, filed Feb. 17, 2011, and U.S.
provisional application No. 61/554,653, filed Nov. 2, 2011, each
incorporated herein by reference in its entirety.
Claims
What is claimed:
1. A compound having a structural formula Cap-L-M wherein Cap is
selected from the group consisting of ##STR00164## wherein ring
##STR00165## is an aliphatic or aromatic five or six membered ring,
W, X, Y, and Z independently are selected from the group consisting
of null, C(R.sup.1).sub.2, .[.O, S,.]. and NR.sup.1, ring
##STR00166## is an aliphatic or aromatic six membered ring, wherein
D, E, F, and G independently are selected from the group consisting
of C(R.sup.1).sub.2, .[.O, and S,.]. or ring ##STR00167## is an
aliphatic or aromatic five membered ring, wherein D, E, F, and G
are selected from the group consisting of null, C(R.sup.1).sub.2,
and NR.sup.1, R.sup.1, independently, is selected from the group
consisting of null, hydrogen, C.sub.1-6alkyl, C.sub.1-6heteroalkyl,
C.sub.2-6alkenyl, C.sub.1-6perfluoroalkyl,
C.sub.1-6perfluoroalkoxy, .[.aryl, heteroaryl,.].
C.sub.3-10cycloalkyl, C.sub.3-10heterocycloalkyl,
C.sub.1-6alkylenearyl, C.sub.1-6alkyleneheteroaryl,
C.sub.1-6alkyleneheterocycloalkyl, C.sub.1-6alkylenecycloalkyl,
##STR00168## OR.sup.a, halo, N(R.sup.a).sub.2, SR.sup.a, SOR.sup.a,
SO.sub.2R.sup.a, CN, C(.dbd.O)R.sup.a, CF.sub.3, OCF.sub.3,
NO.sub.2, OC(.dbd.O)R.sup.a, SO.sub.2N(R.sup.a).sub.2,
OSO.sub.2CF.sub.3, C(.dbd.O)OR.sup.a, C(.dbd.O)N(R.sup.a).sub.2,
C.sub.1-6alkyleneN(R.sup.a).sub.2,
C.sub.1-6alkyleneC(.dbd.O)R.sup.a, C.sub.1-6alkyleneOR.sup.a,
C.sub.1-6alkyleneSR.sup.a,
C.sub.1-6alkyleneNR.sup.aSO.sub.2R.sup.a,
C.sub.1-6alkyleneSOR.sup.a, C.sub.1-6alkyleneCN,
C.sub.1-6heteroalkyleneCN, C.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
C.sub.1-6alkyleneOC(.dbd.O)N(R.sup.a).sub.2,
C.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
C.sub.1-6alkyleneNR.sup.aC(.dbd.O)R.sup.a, .[.C.sub.1-6alkylene
C(.dbd.O)N(R.sup.a).sub.2.].
.Iadd.C.sub.1-6alkyleneC(.dbd.O)N(R.sup.a).sub.2.Iaddend.,
C.sub.1-6alkyleneOC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
C(.dbd.O)NR.sup.aSO.sub.2R.sup.a, C(.dbd.O)C.sub.1-6alkylenearyl,
C(.dbd.O)NR.sup.aC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
OC.sub.1-6alkyleneN(R.sup.a).sub.2, OC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
NR.sup.aC.sub.1-6alkyleneN(R.sup.a).sub.2,
NR.sup.aC(.dbd.O)R.sup.a, NR.sup.aC(.dbd.O)N(R.sup.a).sub.2,
N(SO.sub.2C.sub.1-6alkyl).sub.2, and
NR.sup.a(SO.sub.2C.sub.1-6alkyl), and R.sup.a, independently, is
selected from the group consisting of hydrogen, C.sub.1-6alkyl,
C.sub.1-6heteroalkyl, C.sub.1-6alkyleneNH.sub.2,
C.sub.1-6alkyleneNH(C.sub.1-6alkyl),
C.sub.1-6alkyleneN(C.sub.1-6alkyl).sub.2,
C.sub.1-6alkyleneNH(C.sub.1-6alkyl).sub.2,
C.sub.1-6alkyleneNHC(.dbd.O)(C.sub.1-6alkyl),
C.sub.1-6alkyleneC(.dbd.O)NH.sub.2, C.sub.1-6alkyleneOH,
C.sub.1-6alkyleneCN, C.sub.1-6heteroalkyleneCN,
C.sub.1-6alkyleneOC.sub.1-6alkyl, C.sub.1-6alkyleneSH,
C.sub.1-6alkyleneSC.sub.1-6alkyl,
C.sub.1-6alkyleneNH(SO.sub.2C.sub.1-6alkyl), aryl, heteroaryl,
C.sub.3-8cycloalkyl, and C.sub.3-10heterocycloalkyl, wherein linker
L is attached to any atom D, E, F, or G, .Iadd.and .Iaddend.
##STR00169## wherein ring ##STR00170## is an aliphatic or aromatic
five or six membered ring, E, F, W, X, Y, Z, R.sup.1, and R.sup.a
are as defined above, .[.and ##STR00171## wherein A is C, N, O, S,
B, or P, and L is attached to A, R.sup.2, R.sup.3 and R.sup.4
independently are selected from the group consisting of null,
hydrogen, C.sub.1-6alkyl, C.sub.1-6heteroalkyl, C.sub.2-6alkenyl,
C.sub.1-6perfluoroalkyl, C.sub.1-6perfluoroalkoxy, aryl,
heteroaryl, C.sub.3-10cycloalkyl, C.sub.3-8heterocycloalkyl,
C.sub.1-6alkylenearyl, C.sub.1-6alkyleneheteroaryl,
C.sub.1-6alkyleneheterocycloalkyl, C.sub.1-6alkylenecycloalkyl,
##STR00172## OR.sup.a, halo, N(R.sup.a).sub.2, SR.sup.a, SOR.sup.a,
SO.sub.2R.sup.a, CN, C(.dbd.O)R.sup.a, OC(.dbd.O)R.sup.a,
C(.dbd.O)OR.sup.a, C.sub.1-6alkyleneN(R.sup.a).sub.2,
C.sub.1-6alkyleneOR.sup.a, C.sub.1-6alkyleneSR.sup.a,
C.sub.1-6alkyleneC(.dbd.O)OR.sup.a, C(.dbd.O)N(R.sup.a).sub.2,
C(.dbd.O)NR.sup.aC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
OC.sub.1-6alkyleneN(R.sup.a).sub.2, OC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
NR.sup.aC.sub.1-6alkyleneN(R.sup.a).sub.2,
NR.sup.aC(.dbd.O)R.sup.a, NR.sup.aC(.dbd.O)N(R.sup.a).sub.2,
N(SO.sub.2C.sub.1-6alkyl).sub.2, NR.sup.a(SO.sub.2C.sub.1-6alkyl),
nitro, and SO.sub.2N(R.sup.a).sup.2, and R.sup.a is defined
above;.]. .Iadd.wherein at least one of E, F, W, X, Y, and Z is
NR.sup.1; .Iaddend. L is .[.selected from the group consisting of
null, C.sub.1-8alkylene, R.sup.a substituted C.sub.1-8alkylene,
NR.sup.a, C(.dbd.O), aryl, C(.dbd.O)aryl,
C(.dbd.O)C.sub.1-6alkylene, C.sub.1-8alkyleneNR.sup.a,
C.sub.1-6alkylenearyleneC.sub.1-6alkylene, C.sub.2-6alkenylene,
C.sub.4-8alkdienylene, C.sub.1-6alkylenearylene,
C.sub.1-6alkyleneheteroarylene, R.sup.a substituted
C.sub.1-6alkyleneheteroarylene, and
C.sub.2-6alkenylenearyleneC.sub.1-6alkylene, and R.sup.a is defined
above.]. .Iadd.CH.sub.2--C.sub.6H.sub.5.Iaddend.; M is .Iadd.
##STR00173## .Iaddend. .[.selected from the group consisting of
--C(.dbd.O)N(R.sup.b)OH,
--O(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
--N(R.sup.b)(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
--N(R.sup.b)(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
arylC(.dbd.O)NHOH, --N(OH)C(.dbd.O)R.sup.b,
heteroarylC(.dbd.O)NHOH,
--C.sub.3-6cycloalkylN--C(.dbd.O)CH.sub.2SH, --B(OR.sup.b).sub.2,
SO.sub.2NHR.sup.b, --NHSO.sub.2NHR.sup.b, NHSO.sub.2C.sub.1-6alkyl,
--SO.sub.2C.sub.1-6alkyl, --SR.sup.c, ##STR00174##
--C(.dbd.O)R.sup.e, --P(.dbd.O)(OR.sup.f).sub.2,
--NH--P(.dbd.O)(OR.sup.f).sub.2, ##STR00175##
--C(.dbd.O)(C(R.sup.b).sup.2).sub.1-6SH,
--C(.dbd.O)C(.dbd.O)NHR.sup.b, --C(.dbd.O)NHN(R.sup.b).sub.2,
--C(.dbd.O)NH(CH.sub.2).sub.1-3N(R.sup.b).sub.2, ##STR00176##
--S--(C.dbd.O)C.sub.1-6alkyl, C.sub.3-10heterocycloalkyl optionally
substituted with oxo (.dbd.O), thioxo (.dbd.S), or both, aryl
optionally substituted with one or more of C.sub.1-6alkyl,
--C(.dbd.O)R.sup.d, --NH.sub.2, and --SH, heteroaryl optionally
substituted with --NH.sub.2, --SH, or both, --N(H)C(.dbd.O)SH,
--NHC(.dbd.O)NHR.sup.d, --NHC(.dbd.O)CH.sub.2R.sup.d,
--NHC(.dbd.O)(CH.sub.2).sub.1-6SH, --NHC(.dbd.O)CH.sub.2Hal,
--NHC(.dbd.S)NHR.sup.d, --NHC(.dbd.S)CH.sub.2R.sup.d,
--C(.dbd.S)NHR.sup.d, --C(.dbd.S)CH.sub.2R.sup.d,
--NHC(.dbd.S)CH.sub.2R.sup.d, --NHC(.dbd.S)CH.sub.2Hal, and
--(C.dbd.O)C.sub.1-6alkyl; R.sup.b, independently, is selected from
the group consisting of hydrogen, (C.dbd.O)CH.sub.3,
C.sub.1-6alkyl, CF.sub.3, CH.sub.2F, and aryl, or two R.sup.b
groups are taken together with the carbon to which they attached to
form a C.sub.3-8cycloalkyl group; R.sup.c is selected from hydrogen
or (C.dbd.O)CH.sub.3; R.sup.d is NH.sub.2 or OH; R.sup.e is
selected from the group consisting of OH, N(R.sup.b).sup.2,
NH(OCH.sub.3), N(CH.sub.3)OH, C.sub.1-6alkyl, CF.sub.3, aryl,
heteroaryl, C.sub.3-8cycloalkyl, NHSO.sub.2CH.sub.3,
NHSO.sub.2CF.sub.3, and C.sub.1-6haloalkyl; R.sup.f independently
is hydrogen, methyl, or ethyl; and Hal is halo.]., or a
pharmaceutically acceptable salt, hydrate, or prodrug thereof.
2. The compound of claim 1 wherein the bicyclic ring system
##STR00177## is selected from a residue of the group consisting of
.[.thionaphthene, isothionaphthene,.]. indole, indolenine,
1H-isoindole, cyclopenta[b]pyridine, .[.pyrano[3,4-b]pyrrole,.].
indazole, .[.benzisoxazole, benzoxazole, anthranil,.]. naphthalene,
tetralin, decalin, .[.2H-1-benzopyran, coumarin, coumarin-4-one,
isochromen-1-one, isochromen-3-one,.]. quinoline, isoquinoline,
cinnoline, quinazoline, .[.naphthyridine, pyrido[3,4-b]-pyridine,
pyrido[3,2-b]-pyridine, pyrido[4,3-b]-pyridine, 2H-1,3-benzoxazine,
1H-2,3-benzoxazine, 4H-3,1-benzoxazine, 2H-1,2-benzoxazine,
4H-1,4-benzoxazine,.]. purine, indoline, .[.benzo(b)furan,.].
indene, pteridine, quinoxaline, benzimidazole, .[.benzthiazole,.].
and phthalzine, and wherein the linker L is attached to any atom D,
E, F, G, W, X, Y, or Z.
3. The compound of claim 1 wherein ##STR00178## is selected from
the group consisting of .[.cyclohexyl, cyclohexenyl, cyclopentyl,
cyclopentenyl, cycloheptyl, cycloheptenyl, phenyl, furanyl,.].
2H-pyrrolyl, .[.thienyl,.]. pyrrolyl, .[.oxazolyl, thiazolyl,.].
imidazolyl, pyrazolyl, .[.isoxazolyl, isothiazolyl,
1,2,3,-oxadiazolyl,.]. 1,2,3,-triazolyl, .[.1,2,5-oxadiazolyl,.].
2-pyrrolinyl, 3-pyrrolinyl, pyrrolidinyl,
.[.1,3-dioxolanyl,oxazolyl,.]. 2-imidazolinyl, imidazolidinyl,
2-pyrazolinyl, pyrazolidinyl, 3H-pyrrolyl, .[.1,3-dithiolyl,
3H-1,2-oxathiolyl, 3H-1,2,3-dioxazolyl, 1,3,2-dioxazolyl,
1,2-dithiolyl, 5H-1,2,5-oxathiazolyl, 1,3-oxathiolyl, 2H-pyranyl,
4H-pyranyl, piperidinyl, 1,4-dioxanyl, morpholinyl, 1,4-dithianyl,
thiomorpholinyl,.]. pyrimidinyl, piperazinyl, .[.2-pyronyl,
4-pyronyl, 1,2-dioxinyl, 1,3-dioxinyl, 1,2,4-triazinyl,.].
1,2,3-triazinyl, .[.4H-1,3-oxazinyl, 2H-1,3-oxazinyl,
6H-1,3-oxazinyl, 6H-1,2-oxazinyl, 4H-1,4-oxazinyl, 2H-1,2-oxazinyl,
1,4-oxazinyl, p-isoxazinyl, o-isoxazinyl,.]. pyridinyl,
.[.1,2,6-oxathiazinyl, 1,2,5-oxathiazinyl, 1,4,2-oxadiazinyl,.].
pyridazinyl, .Iadd.and .Iaddend.pyrazinyl.[., azacycloheptyl,
azacycloheptenyl, oxacycloheptyl, and thiacycloheptyl.]., wherein
linker L is attached to any atom .[.U, V.]. .Iadd.E, F.Iaddend., W,
X, Y, or Z.
4. The compound of claim 1 wherein the Cap group has a structure:
##STR00179## either unsubstituted or substituted with one or more
R.sup.1.
.[.5. The compound of claim 1 wherein ##STR00180## .].
6. The compound of claim 1 wherein R.sup.1 is selected from the
group consisting of null, hydrogen, OR.sup.a, halo, C.sub.1-6alkyl,
.[.aryl,.]. heterocycloalkyl, --(CH.sub.2).sub.1-4heterocycloalkyl,
--(CH.sub.2).sub.1-4N(R.sup.a).sub.2, ##STR00181## or
--C(.dbd.O)N(CH.sub.2).sub.1-4N(R.sup.a).sub.2.
7. The compound of claim 1 wherein R.sup.a is selected from the
group consisting of .[.null,.]. hydrogen, C.sub.1-6alkyl,
C.sub.1-6heteroalkyl, aryl, and heteroaryl.
.[.8. The compound of claim 1 wherein L is null,
--(CH.sub.2).sub.1-6--, ##STR00182## optionally substituted with
halo, CF.sub.3, or CN, ##STR00183##
--CH.sub.2--CH.dbd.CH--CH.dbd.CH--,
--(CH.sub.2).sub.2--CH.dbd.CH--CH.dbd.CH.sub.2--,
--(CH.sub.2).sub.0-6--NH--, ##STR00184## .].
.[.9. The compound of claim 1 wherein M is ##STR00185##
--SC(.dbd.O)tBu, --SC(.dbd.O)CF.sub.3,
--S(CH.sub.2).sub.1-3C(.dbd.O)CF.sub.3, --CH.sub.2SAc, ##STR00186##
--NH--C(.dbd.O)CH.sub.2SH, ##STR00187## ##STR00188## .].
10. The compound of claim 1 wherein the Cap group is selected from
the group consisting of ##STR00189## ##STR00190##
11. A composition comprising (a) compound of claim 1, (b) an
optional second therapeutic agent useful in the treatment of a
disease or condition wherein inhibition of histone deacetylase
provides a benefit, and (c) an excipient and/or pharmaceutically
acceptable carrier.
12. A compound selected from the group consisting of ##STR00191##
##STR00192## ##STR00193## ##STR00194##
13. A compound having a structure ##STR00195##
14. A compound having a structure ##STR00196## wherein Cap is
selected from the group consisting of ##STR00197##
Description
FIELD OF THE INVENTION
The present invention relates to histone deacetylase (HDAC)
inhibitors, to pharmaceutical compositions comprising one or more
of the HDAC inhibitors, to methods of increasing the sensitivity of
cancer cells to the cytotoxic effects of radiotherapy and/or
chemotherapy comprising contacting the cell with one or more of the
HDAC inhibitors, and to therapeutic methods of treating conditions
and diseases wherein inhibition of HDAC provides a benefit, for
example, a cancer, an inflammation, a neurological disease, a
neurodegenerative disorder, stroke, traumatic brain injury,
allograft rejection, autoimmune diseases, and malaria, comprising
administering a therapeutically effective amount of a present HDAC
inhibitor to an individual in need thereof.
BACKGROUND OF THE INVENTION
Inhibitors of HDACs modulate transcription and induce cell growth
arrest, differentiation, and apoptosis. HDAC inhibitors (HDACIs)
also enhance the cytotoxic effects of therapeutic agents used in
cancer treatment, including radiation and chemotherapeutic drugs.
Moreover, recent research indicates that transcriptional
dysregulation may contribute to the molecular pathogenesis of
certain neurodegenerative disorders, such as Huntington's disease,
Rett syndrome, Charcot-Marie-Tooth disease (CMT) and other
peripheral neuropathies, spinal muscular atrophy, amyotropic
lateral sclerosis, and ischemia. For example, suberoylanilide
hydroxamic acid (SAHA) has been shown to penetrate into the brain
to dramatically improve motor impairment in a mouse model of
Huntington's disease, thereby validating research directed to
HDACIs in the treatment of neurodegenerative diseases. Furthermore,
selective HDAC6 inhibitors have been shown to rescue the CMT
phenotype, restore proper mitochondrial motility, and correct the
axonal transport defects observed in transgenic mice. Selective
HDAC6 inhibitors also induce the re-innervation of muscles and
increase the number of observed neuromuscular junctions in these
same models (C. d'Ydewalle et al., Nature Medicine 2011).
A recent review summarized evidence that aberrant histone
acetyltransferase (HAT) and HDAC activity may be a common
underlying mechanism contributing to neurodegeneration. Moreover,
from a mouse model of depression, the therapeutic potential of
HDACs in treating depression is discussed. See WO 2008/019025,
designating the United States, incorporated herein in its
entirety.
Eleven isozymes in the HDAC family of enzymes, which can be grouped
into classes by their evolutionary relationships, have been
identified. Structure and function appear to be conserved among
members of the various classes. The HDAC family is made up of class
I HDACs, including HDAC1, 2, 3, and 8; class IIa, including HDAC4,
5, 7, and 9; class IIb, including HDAC6 and 10; and a class IV
enzyme, HDAC11 (A. J. de Ruijter et al., The Biochemical Journal
2003, 370(Pt), 737-749).
The class I HDACs are found primarily in the nucleus and are
expressed in all tissue types, except for the muscle cell-specific
HDAC8. The class I HDACs interact with many key transcription
factors regulating gene expression, including CoREST and NuRD.
Class IIa HDACs have tissue specific expression, and are found in
both the nucleus and cytoplasm. Unlike the other isozymes, the
class IIb HDAC6 does not extensively associate with transcription
factors, and acts as a deacetylase on non-histone proteins,
including .alpha.-tubulin, HSP90, cortactin, and the peroxiredoxins
(O. Witt et al., Cancer Letters 2008; R. B. Parmigiana et al., PNAS
2008).
HDACs form multiprotein complexes with many regulatory proteins
inside the cell. For example, HDAC4, 5, and 7 actually lack
intrinsic deacetylase ability, and gain activity only by
interacting with HDAC3. Each isozyme interacts with a specific
series of regulatory proteins and transcription factors and has a
specific set of substrates, and thus each regulates a specific
series of genes and proteins (O. Witt et al., Cancer Letters 2008).
The design of selective HDAC isozyme inhibitors allows preferential
inhibition of only the isozyme(s) relevant to a particular disease
or condition, thereby reducing the probability of counterproductive
and/or adverse effects resulting from an unwanted and undesired
inhibition of other HDAC isozymes.
HDAC6 is the most abundant histone deacetylase isozyme in the human
body, and along with HDAC7, is the most commonly expressed isozyme
in the brain (A. J. de Ruijter et al., The Biochemical Journal
2003, 370(Pt), 737-749). Structurally significant features of HDAC6
include two deacetylase domains and a zinc finger motif. It is most
commonly found in the cytoplasm, but can be shuttled into the
nucleus via its nuclear export signal. A cytoplasmic retention
signal, which sequesters the enzyme in the cytoplasm, also was
found (A. Valenzuela-Fernandez et al., Trends in Cell Biology 2008,
18(6), 291-297). The functions of HDAC6 are unlike any of the other
HDAC isozymes. Many non-histone substrates are deacetylated by
HDAC6, including .alpha.-tubulin, HSP90, cortactin, and
peroxiredoxins (O. Witt et al., Cancer Letters 2008; R. B.
Parmigiani et al., PNAS USA 2008, 105 (28), 9633-9638).
The design of HDACIs focuses on the three major domains of the
enzyme molecule. A zinc binding group (ZBG) of the HDACI typically
is a hydroxamic acid, benzamide, or thiol, although other
functional groups have been used. This ZBG moiety of the inhibitor
chelates the zinc cofactor found in the active site of the enzyme.
The ZBG moiety typically is bonded to a lipophilic linker group,
which occupies a narrow channel leading from the HDAC surface to
the active site. This linker, in turn, is bonded to a surface
recognition, or `cap`, moiety, which typically is an aromatic group
that interacts with residues at the surface of the enzyme (K. V.
Butler et al., Current Pharmaceutical Design 2008, 14(6),
505-528).
Consideration of each structural element is important in the design
of HDACIs (37). Alteration of the ZBG has profound effects on
inhibitor potency. The most potent inhibitors frequently feature a
hydroxamic acid ZBG, though other groups such as ketones, amides,
and thiols effectively inhibit the enzyme with lower potency. Low
molecular weight compounds having carboxylic acid ZBGs, such as
valproic acid, inhibit HDACs at micromolar potency, but have
profound effects in vivo when given in high doses. Hydroxamic acids
chelate zinc in a bidentate fashion and hydrogen bond with H142 and
H143 (HDAC8), as revealed by crystal structure data. Computational
studies have refined this description of chelation, demonstrating
that the hydroxamic acid carbonyl coordinates zinc more strongly
than the hydroxyl group. Energetically favorable interactions
between hydroxamic acid and the active site provide for high
potency inhibition, but the hydroxamic acid functional group has
some undesirable properties from a drug design perspective.
Hydroxamic acids are potentially mutagenic, and have given positive
results in the Ames test. Hydroxamic acids also are promiscuous
zinc chelators and potently inhibit many zinc-containing
metalloproteins. The replacement of hydroxamic acid with a
different potent ZBG is an active area of research in HDACI design
and development.
The linker region typically is a hydrophobic aryl or alkyl
scaffold, which occupies the hydrophobic HDAC catalytic channel.
Most reported HDACIs, including SAHA, feature an alkyl chain
linker, mimicking the lysine alkyl chain. Aromatic groups are
frequently included in the linker region of an HDACI, for example,
in the HDACIs panbinostat and belinostat.
Manipulation of the cap group can greatly increase potency because
this group has the potential to interact with multiple residues at
the enzyme surface. The most potent inhibitors typically feature an
aryl cap group scaffold. Biaryl and heteroaromatic cap group
scaffolds have been extensively investigated. The large
tetrapeptide motif of apicidin imparts high potency, and gives high
potency with a number of diverse ZBGs. The tetrapeptide cap group
motif is common to natural product HDACIs, and has been engineered
to produce libraries of tetrapeptide HDACIs for use in screening
protocols.
Currently, at least eleven HDACIs are in clinical development.
These HDACIs can be divided into at least five chemical classes,
illustrated below, based on their structure, and in most cases they
broadly and nonselectively inhibit class I/II HDACs with varying
efficiency. These five chemical classes are hydroxamates, cyclic
tetrapeptides, cyclic peptides, short-chain fatty acids, and
benzamides. Typically, known HDACIs fail to show prominent HDAC
isozyme selectivity, which as stated above can cause serious
problems in a clinical setting, especially in the treatment of
diseases and conditions wherein a prolonged drug administration of
an HDACI is required. For example, it has been found that some
HDACIs enhance lung and microglial inflammation (TSA and SAHA), as
well as high glucose-induced inflammation. If this effect is linked
to specific HDAC isozymes, the use of certain HDACIs would be
contraindicated in various diseases and conditions, such as
diabetes and asthma.
##STR00001## ##STR00002##
Additional HDACI's include
##STR00003## ##STR00004##
The following table summarizes some HDACIs that presently are in
clinical trials.
TABLE-US-00001 TABLE I Inhibitor Indications SAHA T-cell lymphoma
(Approved) Romidepsin T-cell lymphoma (Approved) Multiple myeloma
(Phase III) Peripheral T-cell lymphoma (Phase III) Refractory renal
cell cancer (Phase II) Valproic Acid Bipolar disorder (Approved)
Acute myeloid leukemia (Phase I/II with all trans- retinoic acid)
PCI-24781 Leukemia (Phase I/II) ITF-2357 Hodgkins lymphoma (Phase
II) Follicular lymphoma (Phase III, with yttrium-90- ibritumomab)
Juvenile arthritis (Phase II) Myeloproliferative Diseases (Phase
II) MS-275 Melanoma Lymphoma (halted due to dose limiting
toxicities) Advanced acute leukemias (Phase 1) Combination trials
with DNA methyltransferase inhibitors and 5-azacitidine in
non-small cell lung cancer (Preclinical) Panbinostat T-cell
lymphoma (Phase II) Prostate cancer (Phase I with docetaxel)
Belinostat Solid tumors (Phase I) Mesothelioma (Abandoned) MGCD0103
Solid tumors (Phase II with gemcitabine) Diffuse large B-cell
lymphoma (Phase II) EVP-0334 Parkinson's disease (Phase I)
Clinical trial information relating to HDACIs is published in J.
Tan et al., Journal of hematology & oncology. 3:5 (2010) and L.
Wang et al., Nat Rev Drug Discov. 8:969-81 (2009).
HDAC-regulated factors have been implicated in the mechanisms of
major central nervous system (CNS) disorders. In Parkinson's
disease (PD), .alpha.-synuclein binds to histones and inhibits HAT
activity, causing neurodegeneration. Application of HDACIs to PD
neurons blocks .alpha.-synuclein toxicity. Dysregulation of histone
acetylation, involving CBP, a neuroprotective transcription factor
with histone acetyltransferase activity, has been found in
Huntington's disease (HD), Alzheimer's disease (AD), and
Rubinstein-Taybi syndrome (T. Abel et al., Curr. Opin. in
Pharmacol. 2008, 8(1), 57-64). In a cellular model of AD, cell
death was accompanied by loss of CBP function and histone
deacetylation. The mutant HD protein, htt, interacts with CBP,
inhibiting the HAT activity and causing cell death. Treatment with
an HDACI helps to restore histone acetylation, protecting against
neurodegeneration and improving motor performance in a mouse model
of HD (C. Rouaux et al., Biochem. Pharmacol. 2004, 68(6),
1157-1164).
Various studies directed to the application of HDACIs in the
context of CNS disorders have implicated the class II HDACs,
particularly HDAC6, as potential therapeutic targets. One
investigation revealed that inhibition of HDAC6 could be beneficial
as a treatment for HD, a disease for which no pharmacological
treatment is available. The mutant htt protein found in HD disrupts
intracellular transport of the pro-survival and pro-growth nerve
factor, BDNF, along the microtubule network, causing neuronal
toxicity. Inhibition of HDAC6 promotes transport of BDNF by
promoting tubulin hyperacetylation. TSA (trichostatin A), a
nonselective HDAC inhibitor, was found to facilitate transport and
release of BNDF-containing vesicles (J. P. Dompierre et al., J
Neurosci 2007, 27(13), 3571-3583). These results provide a
biological basis for the identification and development of HDACIs,
and particularly HDAC6 selective inhibitors, as a treatment for HD
and other neurodegenerative disorders.
HDACIs prevent or delay neuronal dysfunction and death in in vitro
and in vivo models thereby indicating that HDACIs are broadly
neuroprotective. For example, HDACIs have shown therapeutic
efficacy in the polyglutamine-expansion disorder Huntington's
disease. While the neuroprotective mechanisms of the HDACIs in
rodent models are not yet understood, it is clear that HDACIs
induce the expression of certain genes that confer neuroprotection.
The upregulation of HSP-70 and Bcl-2 through the inhibition of HDAC
has been observed in the cortex and striatum of rats after focal
cerebral ischemia. HSP-70 expression has been found to result in
neuroprotection in a number of disease models including Alzheimer's
disease (AD), Parkinson's disease (PD), and Huntington's disease
(HD). In addition, HDAC6 inhibition leads to the acetylation of
peroxiredoxin and increases its antioxidant activity which may
contribute to the neuroprotective effects of these compounds (R. B.
Parmigiana et al., PNAS 2008).
Studies also provide good evidence that HDACI-induced p21cip1/waf1
expression may play a significant role in HDACI-mediated
neuroprotection. It recently was reported that p21cip1/waf1
overexpression protects neurons from oxidative stress-induced
death, that p21cip1/waf1 is induced in the rodent brain by HDAC
inhibition, and that homozygous loss of p21cip1/waf1 exacerbates
damage in a mouse MCAO/reperfusion model of ischemic stroke. In a
similar study, the HDAC inhibitor TSA was shown to increase
gelsolin expression in neurons, and that gelsolin expression is
necessary for neuroprotection in an oxygen/glucose deprivation
model of neurodegeneration and a mouse MCAO/reperfusion model of
ischemic stroke.
Alternatively, unrelated to histone acetylation and gene
upregulation, proteins such as .alpha.-tubulin and HSP90 are
targets for acetylation and become acetylated when HDACs are
inhibited. In tumor cells, the acetylation of HSP90 has been shown
to decrease the ability of HSP90 to interact with certain client
proteins and thereby abrogate chaperone function. With regard to
stroke and traumatic brain injury (TBI), as well as several other
neurodegenerative diseases, the inhibition of HSP90 is predicted to
have a positive effect on neuronal survival. Indeed, the
pharmacological HSP90 inhibitor, Geldanamycin, and its analogs have
been shown to be neuroprotective in a number of stroke models.
HSP90 inhibition and the consequent release of heat-shock factor
(HSF) to the nucleus may also, in part, explain an upregulation of
HSP70 in the brain during focal ischemia and HDACI treatment.
In addition, HDACIs are useful in the treatment of cancers. For
example, histone acetylation and deacetylation play important roles
in chromatin folding and maintenance (Kornberg et al., Bjorklund et
al., Cell, 1999, 96:759-767; Struhl et al., Cell, 1998, 94:1-4).
Acetylated chromatin is more open and has been implicated in the
increased radiation sensitivities observed in some cell types
(Oleinick et al., Int. J. Radiat. Biol. 1994, 66:523-529).
Furthermore, certain radiation-resistant human cancer cells treated
with the HDACI inhibitor TSA were sensitized to the damaging
effects of ionizing radiation. Thus, HDACIs appear useful as
radiation sensitizing agents.
WO 2008/055068, designating the U.S. and incorporated herein in its
entirety, discloses numerous diseases and conditions treatable by
HDACIs, including the underlying science and reasoning supporting
such treatments.
HDAC6 therefore has emerged as an attractive target for drug
development and research. (C. M. Grozinger et al., Proc. Natl.
Acad. Sci. USA 1999, 96, 4868-73; and C. Boyault et al., Oncogene
2007, 26, 5468-76.) Presently, HDAC6 inhibition is believed to
offer potential therapies for autoimmunity, cancer, and many
neurodegenerative conditions. (S. Minucci et al., Nat. Rev. Cancer.
2006, 6, 38-51; L. Wang et al., Nat. Rev. Drug Discov. 2009, 8,
969-81; J. P. Dompierre et al., J. Neurosci. 2007, 27, 3571-83; and
A. G. Kazantsev et al., Nat. Rev. Drug Discov. 2008, 7, 854-68.)
Selective inhibition of HDAC6 by small molecule or genetic tools
has been demonstrated to promote survival and re-growth of neurons
following injury, offering the possibility for pharmacological
intervention in both CNS injury and neurodegenerative conditions.
(M. A. Rivieccio et al., Proc. Natl. Acad. Sci. USA 2009, 106,
19599-604.) Unlike other histone deacetylases, inhibition of HDAC6
does not appear to be associated with any toxicity, making it an
excellent drug target. (O. Witt et al., Cancer Lett 2009, 277,
8-21.) Tubacin, an HDAC6 selective inhibitor, used in models of
disease, has helped to validate, in part, HDAC6 as a drug target,
but its non-drug-like structure, high lipophilicity (ClogP=6.36
(KOWWIN)) and tedious synthesis make it more useful as a research
tool than a drug. (S. Haggarty et al., Proc. Natl. Acad. Sci. USA
2003, 100, 4389-94.) Other compounds also have a modest preference
for inhibiting HDAC6. (S. Schafer et al., ChemMedChem 2009, 4,
283-90; Y. Itoh et al., J. Med. Chem. 2007, 50, 5425-38; and S.
Manku et al., Bioorg. Med. Chem. Lett. 2009, 19, 1866-70.)
In summary, extensive evidence supports a therapeutic role for
HDACIs in the treatment of a variety of conditions and diseases,
such as cancers and CNS diseases and degenerations. However,
despite exhibiting overall beneficial effects, like beneficial
neuroprotective effects, for example, HDACIs known to date have
little specificity with regard to HDAC inhibition, and therefore
inhibit all zinc-dependent histone deacetylases. It is still
unknown which is (are) the salient HDAC(s) that mediate(s)
neuroprotection when inhibited. Emerging evidence suggests that at
least some of the HDAC isozymes are absolutely required for the
maintenance and survival of neurons, e.g., HDAC1. Additionally,
adverse side effect issues have been noted with nonspecific HDAC
inhibition. Thus, the clinical efficacy of present-day nonspecific
HDACIs for stroke, neurodegenerative disorders, neurological
diseases, and other diseases and conditions ultimately may be
limited. It is important therefore to design, synthesize, and test
compounds capable of serving as potent, and preferably
isozyme-selective, HDACIs that are able to ameliorate the effects
of neurological disease, neurodegenerative disorder, traumatic
brain injury, cancer, inflammation, malaria, autoimmune diseases,
immunosuppressive therapy, and other conditions and diseases
mediated by HDACs.
An important advance in the art would be the discovery of HDACIs,
and particularly selective HDAC6 inhibitors, that are useful in the
treatment of diseases wherein HDAC inhibition provides a benefit,
such as cancers, neurological diseases, traumatic brain injury,
neurodegenerative disorders and other peripheral neuropathies,
stroke, hypertension, malaria, allograft rejection, rheumatoid
arthritis, and inflammations. Accordingly, a significant need
exists in the art for efficacious compounds, compositions, and
methods useful in the treatment of such diseases, alone or in
conjunction with other therapies used to treat these diseases and
conditions. The present invention is directed to meeting this
need.
SUMMARY OF THE INVENTION
The present invention relates to HDACIs, pharmaceutical
compositions comprising the HDACIs, and methods of treating
diseases and conditions wherein inhibition of HDAC provides a
benefit, such as a cancer, a neurological disease, a
neurodegenerative disorder, a peripheral neuropathy, stroke,
hypertension, an inflammation, traumatic brain injury, rheumatoid
arthritis, allograft rejection, autoimmune diseases, and malaria,
comprising administering a therapeutically effective amount of an
HDACI to an individual in need thereof. The present invention also
relates to a method of increasing the sensitivity of a cancer cell
to radiotherapy and/or chemotherapy. The present invention also
allows for the use of these HDACIs inhibitors in combination with
other drugs and/or therapeutic approaches. In some embodiments, the
present HDACIs exhibit selectivity for particular HDAC isozymes,
such as HDAC6, over other HDAC isozymes.
More particularly, the present invention relates to HDACIs having a
structural formula: Cap-L-M
wherein Cap is selected from the group consisting of
##STR00005##
wherein ring
##STR00006## is an aliphatic or aromatic five or six membered
ring,
W, X, Y, and Z independently are selected from the group consisting
of null, C(R.sup.1).sub.2, O, S, and NR.sup.1,
ring
##STR00007## is an aliphatic or aromatic five or six membered
ring,
D, E, F, and G independently are selected from the group consisting
of null, C(R.sup.1).sub.2, O, S, and NR.sup.1,
R.sup.1, independently, is selected from the group consisting of
null, hydrogen, C.sub.1-6alkyl, C.sub.1-6heteroalkyl,
C.sub.2-6alkenyl, C.sub.1-6perfluoroalkyl, C.sub.1-6
perfluoroalkoxy, aryl, heteroaryl, C.sub.3-10cycloalkyl,
C.sub.3-10heterocycloalkyl, C.sub.1-6alkylenearyl,
C.sub.1-6alkyleneheteroaryl, C.sub.1-6alkyleneheterocycloalkyl,
C.sub.1-6alkylenecycloalkyl,
##STR00008## OR.sup.a, halo, N(R.sup.a).sub.2, SR.sup.a, SOR.sup.a,
SO.sub.2R.sup.a, CN, C(.dbd.O)R.sup.a, CF.sub.3, OCF.sub.3,
NO.sub.2, OC(.dbd.O)R.sup.a, SO.sub.2N(R.sup.a).sub.2,
OSO.sub.2CF.sub.3, C(.dbd.O)OR.sup.a, C(.dbd.O)N(R.sup.a).sub.2,
C.sub.1-6alkyleneN(R.sup.a).sub.2,
C.sub.1-6alkyleneC(.dbd.O)R.sup.a, C.sub.1-6alkyleneOR.sup.a,
C.sub.1-6alkyleneSR.sup.a,
C.sub.1-6alkyleneNR.sup.aSO.sub.2R.sup.a,
C.sub.1-6alkyleneSOR.sup.a, C.sub.1-6alkyleneCN,
C.sub.1-6heteroalkyleneCN, C.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
C.sub.1-6alkyleneOC(.dbd.O)N(R.sup.a).sub.2,
C.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
C.sub.1-6alkyleneNR.sup.aC(.dbd.O)R.sup.a, C.sub.1-6alkylene
C(.dbd.O)N(R.sup.a).sub.2,
C.sub.1-6alkyleneOC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
C(.dbd.O)NR.sup.aSO.sub.2R.sup.a, C(.dbd.O)C.sub.1-6alkylenearyl,
C(.dbd.O)NR.sup.aC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
OC.sub.1-6alkyleneN(R.sup.a).sub.2, OC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
NR.sup.aC.sub.1-6alkyleneN(R.sup.a).sub.2,
NR.sup.aC(.dbd.O)R.sup.a, NR.sup.aC(.dbd.O)N(R.sup.a).sub.2,
N(SO.sub.2C.sub.1-6alkyl).sub.2, and
NR.sup.a(SO.sub.2C.sub.1-6alkyl), and
R.sup.a, independently, is selected from the group consisting of
hydrogen, C.sub.1-6alkyl, C.sub.1-6heteroalkyl,
C.sub.1-6alkyleneNH.sub.2, C.sub.1-6alkyleneNH(C.sub.1-6alkyl),
C.sub.1-6alkyleneN(C.sub.1-6alkyl).sub.2,
C.sub.1-6alkyleneNH(C.sub.1-6alkyl).sub.2,
C.sub.1-6alkyleneNHC(.dbd.O)(C.sub.1-6alkyl),
C.sub.1-6alkyleneC(.dbd.O)NH.sub.2, C.sub.1-6alkyleneOH,
C.sub.1-6alkyleneCN, C.sub.1-6heteroalkyleneCN,
C.sub.1-6alkyleneOC.sub.1-6 alkyl, C.sub.1-6alkyleneSH,
C.sub.1-6alkyleneSC.sub.1-6alkyl,
C.sub.1-6alkyleneNH(SO.sub.2C.sub.1-6alkyl), aryl, heteroaryl,
C.sub.3-8cycloalkyl, and C.sub.3-10heterocycloalkyl,
##STR00009##
wherein ring
##STR00010## is an aliphatic or aromatic five or six membered
ring,
E, F, W, X, Y, Z, R.sup.1, and R.sup.a are as defined above,
and
##STR00011##
wherein A is C, N, O, S, B, or P, and L is attached to A,
R.sup.2, R.sup.3 and R.sup.4 independently are selected from the
group consisting of null, hydrogen, C.sub.1-6alkyl,
C.sub.1-6heteroalkyl, C.sub.2-6alkenyl, C.sub.1-6perfluoroalkyl,
C.sub.1-6perfluoroalkoxy, aryl, heteroaryl, C.sub.3-10cycloalkyl,
C.sub.3-8heterocycloalkyl, C.sub.1-6alkylenearyl,
C.sub.1-6alkyleneheteroaryl, C.sub.1-6alkyleneheterocycloalkyl,
C.sub.1-6alkylenecycloalkyl,
##STR00012## OR.sup.a, halo, N(R.sup.a).sub.2, SR.sup.a, SOR.sup.a,
SO.sub.2R.sup.a, CN, C(.dbd.O)R.sup.a, OC(.dbd.O)R.sup.a,
C(.dbd.O)OR.sup.a, C.sub.1-6alkyleneN(R.sup.a).sub.2,
C.sub.1-6alkyleneOR.sup.a, C.sub.1-6alkyleneSR.sup.a,
C.sub.1-6alkyleneC(.dbd.O)OR.sup.a, C(.dbd.O)N(R.sup.a).sub.2,
C(.dbd.O)NR.sup.aC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
OC.sub.1-6alkyleneN(R.sup.a).sub.2, OC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
NR.sup.aC.sub.1-6alkyleneN(R.sup.a).sub.2,
NR.sup.aC(.dbd.O)R.sup.a, NR.sup.aC(.dbd.O)N(R.sup.a).sub.2,
N(SO.sub.2C.sub.1-6alkyl).sub.2, NR.sup.a(SO.sub.2C.sub.1-6alkyl),
nitro, and SO.sub.2N(R.sup.a).sup.2, and
R.sup.a is defined above;
L is selected from the group consisting of null, C.sub.1-8alkylene,
R.sup.a substituted C.sub.1-8alkylene, NR.sup.a, C(.dbd.O), aryl,
C(.dbd.O)aryl, C(.dbd.O)C.sub.1-6alkylene,
C.sub.1-8alkyleneNR.sup.a,
C.sub.1-6alkylenearyleneC.sub.1-6alkylene, C.sub.2-6alkenylene,
C.sub.4-8alkdienylene, C.sub.1-6alkylenearylene,
C.sub.1-6alkyleneheteroarylene, R.sup.a substituted
C.sub.1-6alkyleneheteroarylene, and
C.sub.2-6alkenylenearyleneC.sub.1-6alkylene, and R.sup.a is defined
above;
M is selected from the group consisting of
--C(.dbd.O)N(R.sup.b)OH,
--O(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
--N(R.sup.b)(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
--N(R.sup.b)(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
arylC(.dbd.O)NHOH,
--N(OH)C(.dbd.O)R.sup.b,
heteroarylC(.dbd.O)NHOH,
--C.sub.3-6cycloalkylN--C(.dbd.O)CH.sub.2SH,
--B(OR.sup.b).sub.2,
--SO.sub.2NHR.sup.b,
--NHSO.sub.2NHR.sup.b,
--NHSO.sub.2C.sub.1-6alkyl,
--SO.sub.2C.sub.1-6alkyl,
--SR.sup.c,
##STR00013##
--C(.dbd.O)R.sup.e,
--P(.dbd.O)(OR.sup.f).sub.2,
--NH--P(.dbd.O)(OR.sup.f).sub.2,
##STR00014##
--C(.dbd.O)(C(R.sup.b).sup.2).sub.1-6SH,
--C(.dbd.O)C(.dbd.O)NHR.sup.b,
--C(.dbd.O)NHN(R.sup.b).sub.2,
--C(.dbd.O)NH(CH.sub.2).sub.1-3N(R.sup.b).sub.2,
##STR00015##
--S--(C.dbd.O)C.sub.1-6alkyl,
C.sub.3-10heterocycloalkyl optionally substituted with oxo
(.dbd.O), thioxo (.dbd.S), or both,
aryl optionally substituted with one or more of C.sub.1-6alkyl,
--C(.dbd.O)R.sup.d, --NH.sub.2, and --SH,
heteroaryl optionally substituted with --NH.sub.2, --SH, or
both,
--N(H)C(.dbd.O)SH,
--NHC(.dbd.O)NHR.sup.d,
--NHC(.dbd.O)CH.sub.2R.sup.d,
--NHC(.dbd.O)(CH.sub.2).sub.1-6SH,
--NHC(.dbd.O)CH.sub.2Hal,
--NHC(.dbd.S)NHR.sup.d,
--NHC(.dbd.S)CH.sub.2R.sup.d,
--C(.dbd.S)NHR.sup.d,
--C(.dbd.S)CH.sub.2R.sup.d,
--NHC(.dbd.S)CH.sub.2R.sup.d,
--NHC(.dbd.S)CH.sub.2Hal, and
--(C.dbd.O)C.sub.1-6alkyl;
R.sup.b, independently, is selected from the group consisting of
hydrogen, (C.dbd.O)CH.sub.3, C.sub.1-6alkyl, CF.sub.3, CH.sub.2F,
and aryl, or two R.sup.b groups are taken together with the carbon
to which they attached to form a C.sub.3-8cycloalkyl group;
R.sup.c is selected from hydrogen or (C.dbd.O)CH.sub.3;
R.sup.d is NH.sub.2 or OH;
R.sup.e is selected from the group consisting of OH,
N(R.sup.b).sup.2, NH(OCH.sub.3), N(CH.sub.3)OH, C.sub.1-6alkyl,
CF.sub.3, aryl, heteroaryl, C.sub.3-8cycloalkyl,
NHSO.sub.2CH.sub.3, NHSO.sub.2CF.sub.3, and C.sub.1-6haloalkyl;
R.sup.f independently is hydrogen, methyl, or ethyl; and
Hal is halo,
or a pharmaceutically acceptable salt, hydrate, prodrug
thereof.
In another embodiment, the present invention provides a method of
treating a condition or disease by administering a therapeutically
effective amount of a present HDACI to an individual in need
thereof. The disease or condition of interest is treatable by
inhibition of HDAC, for example, a cancer, a neurodegenerative
disorder, a traumatic brain injury, a neurological disease,
peripheral neuropathy, an inflammation, stroke, hypertension, an
autoimmune disease, allograft rejection, and malaria.
The present HDACIs contain, but are not limited to, a bidentate
chelate as the zinc binding group (ZBG). Preferably, a present
HDACI contains a relatively short linker group between the ZBG and
the aromatic surface recognition group, e.g., contains a 0 to 5
carbon atom chain. The surface recognition group is a bicyclic,
monocyclic, or acyclic moiety, such as indolyl, i.e.,
##STR00016## i.e., pyridinyl, i.e.,
##STR00017## or phenyl, i.e.,
##STR00018##
It has been found that a degree of isoform selectivity for an HDACI
can be achieved by manipulating the surface recognition group in
concert with the ZBG. In particular, a combination of steric and
electronic properties of the surface recognition group modulates
the ability of the compounds to target different isoforms via
interactions with an HDAC surface. Such considerations led to the
present HDACIs having a surface recognition group that exhibits
selectivity in the inhibition of HDAC6.
Another embodiment of the present invention provides a method of
treating a cancer comprising administering to an individual in need
thereof, such as a human, a therapeutically effective amount of a
present HDACI. A present HDACI can be administered as the sole
anticancer therapy, or in conjunction with a therapeutically
effective amount of a second anticancer agent, such as radiation
and/or chemotherapy.
Another embodiment of the present invention provides a method of
increasing the sensitivity of a cancer cell to the cytotoxic
effects of radiotherapy and/or chemotherapy comprising contacting
the cell with an effective amount of a present HDACI. In certain
embodiments, the cell is an in vivo cell.
In another embodiment, the present invention provides a method of
treating a neurological disease comprising administering to an
individual in need thereof, such as a human, a therapeutically
effective amount of a present HDACI. The present invention also
relates to a method of treating neurodegenerative disorders,
peripheral neuropathies, and traumatic brain injuries comprising
administering a therapeutically effective amount of a present HDACI
to an individual in need thereof. In each embodiment, a present
HDACI can be the sole therapeutic agent or can be administered with
additional therapeutic agents known to treat the disease or
condition of interest.
The present invention also provides a method of treating malaria
and other parasitic infections comprising administering a
therapeutically effective amount of a present HDACI to an
individual in need thereof. In certain embodiments, the individual
is a human. In certain embodiments, said method further comprises
optionally coadministering a second antimalarial compound (e.g.,
chloroquine).
In yet another embodiment, the present invention provides a method
of inducing immunosuppression in an individual comprising
administration of a therapeutically effective amount of a present
HDACI to an individual in need thereof, for example, an individual
receiving a transplant. This method further comprises optionally
coadministering a second immunosuppressant (e.g., cyclosporin) or
therapeutic agent.
In still another embodiment, the present invention provides a
method of treating inflammatory diseases and conditions, e.g.,
arthritis and rheumatic diseases, comprising administration of a
therapeutically effective amount of a present HDACI to an
individual in need thereof. The method further contemplates
optional coadministration of a second anti-inflammatory drug or
therapeutic agent.
In another embodiment, the present invention also provides a
pharmaceutical composition comprising a present HDACI and a
pharmaceutically acceptable excipient.
Another embodiment of the present invention is to utilize a present
HDACI and an optional second therapeutically active agent in a
method of treating an individual for a disease or condition wherein
inhibition of HDAC provides a benefit.
In a further embodiment, the invention provides for use of a
composition comprising a present HDACI and an optional second
therapeutic agent for the manufacture of a medicament for treating
a disease or condition of interest, e.g., a cancer,
neurodegeneration, or autoimmunity.
Still another embodiment of the present invention is to provide a
kit for human pharmaceutical use comprising (a) a container, (b1) a
packaged composition comprising a present HDACI, and, optionally,
(b2) a packaged composition comprising a second therapeutic agent
useful in the treatment of a disease or condition of interest, and
(c) a package insert containing directions for use of the
composition or compositions, administered simultaneously or
sequentially, in the treatment of the disease or condition of
interest.
A present HDACI and the second therapeutic agent can be
administered together as a single-unit dose or separately as
multi-unit doses, wherein a present HDACI is administered before
the second therapeutic agent, or vice versa. It is envisioned that
one or more dose of a present HDACI and/or one or more dose of a
second therapeutic agent can be administered.
In one embodiment, a present HDACI and a second therapeutic agent
are administered simultaneously. In related embodiments, a present
HDACI and second therapeutic agent are administered from a single
composition or from separate compositions. Ina further embodiment,
a present HDACI and a second therapeutic agent are administered
sequentially. A present HDACI can be administered in an amount of
about 0.005 to about 500 milligrams per dose, about 0.05 to about
250 milligrams per dose, or about 0.5 to about 100 milligrams per
dose.
Compounds of the invention inhibit HDAC and are useful research
tools for in vitro study of histone deacetylases and their role in
biological processes.
These and other novel aspects of the present invention will become
apparent from the following detailed description of the preferred
embodiments.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 contains bar graphs of concentrations of TSA vs. survival (%
control) for the HCA oxidative stress assay; and
FIG. 2 contains bar graphs of concentrations of various inventive
HDACIs vs. survival (% control) for the HCA oxidative stress
assay.
DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS
The present invention is directed to novel HDACIs and their use in
therapeutic treatments of, for example, cancers, inflammations,
traumatic brain injuries, neurodegenerative disorders, neurological
diseases, peripheral neuropathies, strokes, hypertension,
autoimmune diseases, inflammatory diseases, and malaria. The
present HDACIs also increase the sensitivity of a cancer cell to
the cytotoxic effects of radiotherapy and/or chemotherapy. In some
embodiments, the present HDACIs selectively inhibit HDAC6 over
other HDAC isozymes.
The present invention is described in connection with preferred
embodiments. However, it should be appreciated that the invention
is not limited to the disclosed embodiments. It is understood that,
given the description of the embodiments of the invention herein,
various modifications can be made by a person skilled in the art.
Such modifications are encompassed by the claims below.
The term "a disease or condition wherein inhibition of HDAC
provides a benefit" pertains to a condition in which HDAC and/or
the action of HDAC is important or necessary, e.g., for the onset,
progress, expression of that disease or condition, or a disease or
a condition which is known to be treated by an HDAC inhibitor (such
as, e.g., TSA, pivalolyloxymethylbutane (AN-9; Pivanex), FK-228
(Depsipeptide), PXD-101, NVP-LAQ824, SAHA, MS-275, and or
MGCD0103). Examples of such conditions include, but are not limited
to, cancer, psoriasis, fibroproliferative disorders (e.g., liver
fibrosis), smooth muscle proliferative disorders (e.g.,
atherosclerosis, restenosis), neurodegenerative diseases (e.g.,
Alzheimer's, Parkinson's, Huntington's chorea, amyotropic lateral
sclerosis, spino-cerebellar degeneration, Rett syndrome),
peripheral neuropathies (Charcot-Marie-Tooth disease, Giant Axonal
Neuropathy (GAN)), inflammatory diseases (e.g., osteoarthritis,
rheumatoid arthritis, colitis), diseases involving angiogenesis
(e.g., cancer, rheumatoid arthritis, psoriasis, diabetic
retinopathy), hematopoietic disorders (e.g., anemia, sickle cell
anemia, thalasseimia), fungal infections, parasitic infections
(e.g., malaria, trypanosomiasis, helminthiasis, protozoal
infections), bacterial infections, viral infections, and conditions
treatable by immune modulation (e.g., multiple sclerosis,
autoimmune diabetes, lupus, atopic dermatitis, allergies, asthma,
allergic rhinitis, inflammatory bowel disease; and for improving
grafting of transplants). One of ordinary skill in the art is
readily able to determine whether a compound treats a disease or
condition mediated by HDAC for any particular cell type, for
example, by assays which conveniently can be used to assess the
activity of particular compounds.
The term "second therapeutic agent" refers to a therapeutic agent
different from a present HDACI and that is known to treat the
disease or condition of interest. For example, when a cancer is the
disease or condition of interest, the second therapeutic agent can
be a known chemotherapeutic drug, like taxol, or radiation, for
example.
The term "HDAC" refers to a family of enzymes that remove acetyl
groups from a protein, for example, the -amino groups of lysine
residues at the N-terminus of a histone. The HDAC can be a human
HDAC, including, HDAC1, HDAC2, HDAC3, HDAC4, HDAC5, HDAC6, HDAC7,
HDAC8, HDAC9, HDAC10, and HDAC11. The HDAC also can be derived from
a protozoal or fungal source.
HDAC inhibitors (HDACIs) typically contain three structural
elements which are analogous to the structure of acetyllysine.
These three structural elements are a zinc binding group (M), which
is responsible for chelation of zinc in the active site, a linker
region (L), which binds to the hydrophobic channel that connects
the active site to the outer enzyme surface, and a capping group
(Cap), which interacts with residues at the outer enzyme
surface.
As used herein, the terms "treat," "treating," "treatment," and the
like refer to eliminating, reducing, relieving, reversing, and/or
ameliorating a disease or condition and/or symptoms associated
therewith. Although not precluded, treating a disease or condition
does not require that the disease, condition, or symptoms
associated therewith be completely eliminated, including the
treatment of acute or chronic signs, symptoms and/or malfunctions.
As used herein, the terms "treat," "treating," "treatment," and the
like may include "prophylactic treatment," which refers to reducing
the probability of redeveloping a disease or condition, or of a
recurrence of a previously-controlled disease or condition, in a
subject who does not have, but is at risk of or is susceptible to,
redeveloping a disease or condition or a recurrence of the disease
or condition, "treatment" therefore also includes relapse
prophylaxis or phase prophylaxis. The term "treat" and synonyms
contemplate administering a therapeutically effective amount of a
compound of the invention to an individual in need of such
treatment. A treatment can be orientated symptomatically, for
example, to suppress symptoms. It can be effected over a short
period, be oriented over a medium term, or can be a long-term
treatment, for example within the context of a maintenance
therapy.
The term "therapeutically effective amount" or "effective dose" as
used herein refers to an amount of the active ingredient(s) that,
when administered, is (are) sufficient, to efficaciously deliver
the active ingredient(s) for the treatment of condition or disease
of interest to an individual in need thereof. In the case of a
cancer or other proliferation disorder, the therapeutically
effective amount of the agent may reduce (i.e., retard to some
extent and preferably stop) unwanted cellular proliferation; reduce
the number of cancer cells; reduce the tumor size; inhibit (i.e.,
retard to some extent and preferably stop) cancer cell infiltration
into peripheral organs; inhibit (i.e., retard to some extent and
preferably stop) tumor metastasis; inhibit, to some extent, tumor
growth; reduce HDAC signaling in the target cells; and/or relieve,
to some extent, one or more of the symptoms associated with the
cancer. To extent the administered compound or composition prevents
growth and/or kills existing cancer cells, it may be cytostatic
and/or cytotoxic.
The term "container" means any receptacle and closure therefor
suitable for storing, shipping, dispensing, and/or handling a
pharmaceutical product.
The term "insert" means information accompanying a pharmaceutical
product that provides a description of how to administer the
product, along with the safety and efficacy data required to allow
the physician, pharmacist, and patient to make an informed decision
regarding use of the product. The package insert generally is
regarded as the "label" for a pharmaceutical product.
"Concurrent administration," "administered in combination,"
"simultaneous administration," and similar phrases mean that two or
more agents are administered concurrently to the subject being
treated. By "concurrently," it is meant that each agent is
administered either simultaneously or sequentially in any order at
different points in time. However, if not administered
simultaneously, it is meant that they are administered to an
individual in a sequence and sufficiently close in time so as to
provide the desired therapeutic effect and can act in concert. For
example, a present HDACI can be administered at the same time or
sequentially in any order at different points in time as a second
therapeutic agent. A present HDACI and the second therapeutic agent
can be administered separately, in any appropriate form and by any
suitable route. When a present HDACI and the second therapeutic
agent are not administered concurrently, it is understood that they
can be administered in any order to a subject in need thereof. For
example, a present HDACI can be administered prior to (e.g., 5
minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4
hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1
week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12
weeks before), concomitantly with, or subsequent to (e.g., 5
minutes, 15 minutes, 30 minutes, 45 minutes, 1 hour, 2 hours, 4
hours, 6 hours, 12 hours, 24 hours, 48 hours, 72 hours, 96 hours, 1
week, 2 weeks, 3 weeks, 4 weeks, 5 weeks, 6 weeks, 8 weeks, or 12
weeks after) the administration of a second therapeutic agent
treatment modality (e.g., radiotherapy), to an individual in need
thereof. In various embodiments, a present HDACI and the second
therapeutic agent are administered 1 minute apart, 10 minutes
apart, 30 minutes apart, less than 1 hour apart, 1 hour apart, 1
hour to 2 hours apart, 2 hours to 3 hours apart, 3 hours to 4 hours
apart, 4 hours to 5 hours apart, 5 hours to 6 hours apart, 6 hours
to 7 hours apart, 7 hours to 8 hours apart, 8 hours to 9 hours
apart, 9 hours to 10 hours apart, 10 hours to 11 hours apart, 11
hours to 12 hours apart, no more than 24 hours apart or no more
than 48 hours apart. In one embodiment, the components of the
combination therapies are administered at 1 minute to 24 hours
apart.
The use of the terms "a", "an", "the", and similar referents in the
context of describing the invention (especially in the context of
the claims) are to be construed to cover both the singular and the
plural, unless otherwise indicated. Recitation of ranges of values
herein merely serve as a shorthand method of referring individually
to each separate value falling within the range, unless otherwise
indicated herein, and each separate value and subrange is
incorporated into the specification as if it were individually
recited herein. The use of any and all examples, or exemplary
language (e.g., "such as" and "like") provided herein, is intended
to better illustrate the invention and is not a limitation on the
scope of the invention unless otherwise claimed. No language in the
specification should be construed as indicating any non-claimed
element as essential to the practice of the invention.
In particular, the present invention is directed to HDACIs,
compositions comprising a present HDACI, and therapeutic uses of
the HDACIs of the following structural formula: Cap-L-M
wherein Cap is selected from the group consisting of
##STR00019##
wherein ring
##STR00020## is an aliphatic or aromatic five or six membered
ring,
W, X, Y, and Z independently are selected from the group consisting
of null, C(R.sup.1).sub.2, O, S, and NR.sup.1,
ring
##STR00021## is an aliphatic or aromatic five or six membered
ring,
D, E, F, and G independently are selected from the group consisting
of null, C(R.sup.1).sub.2, O, S, and NR.sup.1,
R.sup.1, independently, is selected from the group consisting of
null, hydrogen, C.sub.1-6alkyl, C.sub.1-6heteroalkyl,
C.sub.2-6alkenyl, C.sub.1-6perfluoroalkyl,
C.sub.1-6perfluoroalkoxy, aryl, heteroaryl, C.sub.3-10cycloalkyl,
C.sub.3-10heterocycloalkyl, C.sub.1-6alkylenearyl,
C.sub.1-6alkyleneheteroaryl, C.sub.1-6alkyleneheterocycloalkyl,
C.sub.1-6alkylenecycloalkyl,
##STR00022## OR.sup.a, halo, N(R.sup.a).sub.2, SR.sup.a, SOR.sup.a,
SO.sub.2R.sup.a, CN, C(.dbd.O)R.sup.a, CF.sub.3, OCF.sub.3,
NO.sub.2, OC(.dbd.O)R.sup.a, SO.sub.2N(R.sup.a).sub.2,
OSO.sub.2CF.sub.3, C(.dbd.O)OR.sup.a, C(.dbd.O)N(R.sup.a).sub.2,
C.sub.1-6alkyleneN(R.sup.a).sub.2,
C.sub.1-6alkyleneC(.dbd.O)R.sup.a, C.sub.1-6alkyleneOR.sup.a,
C.sub.1-6alkyleneSR.sup.a,
C.sub.1-6alkyleneNR.sup.aSO.sub.2R.sup.a,
C.sub.1-6alkyleneSOR.sup.a, C.sub.1-6alkyleneCN,
C.sub.1-6heteroalkyleneCN, C.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
C.sub.1-6alkyleneOC(.dbd.O)N(R.sup.a).sub.2,
C.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
C.sub.1-6alkyleneNR.sup.aC(.dbd.O)R.sup.a, C.sub.1-6alkylene
C(.dbd.O)N(R.sup.a).sub.2,
C.sub.1-6alkyleneOC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
C(.dbd.O)NR.sup.aSO.sub.2R.sup.a, C(.dbd.O)C.sub.1-6alkylenearyl,
C(.dbd.O)NR.sup.aC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
OC.sub.1-6alkyleneN(R.sup.a).sub.2, OC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
NR.sup.aC.sub.1-6alkyleneN(R.sup.a).sub.2,
NR.sup.aC(.dbd.O)R.sup.a, NR.sup.aC(.dbd.O)N(R.sup.a).sub.2,
N(SO.sub.2C.sub.1-6alkyl).sub.2, and
NR.sup.a(SO.sub.2C.sub.1-6alkyl), and
R.sup.a, independently, is selected from the group consisting of
hydrogen, C.sub.1-6alkyl, C.sub.1-6heteroalkyl,
C.sub.1-6alkyleneNH.sub.2, C.sub.1-6alkyleneNH(C.sub.1-6alkyl),
C.sub.1-6alkyleneN(C.sub.1-6alkyl).sub.2,
C.sub.1-6alkyleneNH(C.sub.1-6alkyl).sub.2,
C.sub.1-6alkyleneNHC(.dbd.O)(C.sub.1-6alkyl),
C.sub.1-6alkyleneC(.dbd.O)NH.sub.2, C.sub.1-6alkyleneOH,
C.sub.1-6alkyleneCN, C.sub.1-6heteroalkyleneCN,
C.sub.1-6alkyleneOC.sub.1-6 alkyl, C.sub.1-6alkyleneSH,
C.sub.1-6alkyleneSC.sub.1-6alkyl,
C.sub.1-6alkyleneNH(SO.sub.2C.sub.1-6alkyl), aryl, heteroaryl,
C.sub.3-8cycloalkyl, and C.sub.3-10heterocycloalkyl,
##STR00023##
wherein ring
##STR00024## is an aliphatic or aromatic five or six membered
ring,
E, F, W, X, Y, Z, R.sup.1, and R.sup.a are as defined above,
and
##STR00025##
wherein A is C, N, O, S, B, or P, and L is attached to A,
R.sup.2, R.sup.3 and R.sup.4 independently are selected from the
group consisting of null, hydrogen, C.sub.1-6alkyl,
C.sub.1-6heteroalkyl, C.sub.2-6alkenyl, C.sub.1-6perfluoroalkyl,
C.sub.1-6perfluoroalkoxy, aryl, heteroaryl, C.sub.3-10cycloalkyl,
C.sub.3-8heterocycloalkyl, C.sub.1-6alkylenearyl,
C.sub.1-6alkyleneheteroaryl, C.sub.1-6alkyleneheterocycloalkyl,
C.sub.1-6alkylenecycloalkyl,
##STR00026## OR.sup.a, halo, N(R.sup.a).sub.2, SR.sup.a, SOR.sup.a,
SO.sub.2R.sup.a, CN, C(.dbd.O)R.sup.a, OC(.dbd.O)R.sup.a,
C(.dbd.O)OR.sup.a, C.sub.1-6alkylerleN(R.sup.a).sub.2,
C.sub.1-6alkyleneOR.sup.a, C.sub.1-6alkyleneSR.sup.a,
C.sub.1-6alkyleneC(.dbd.O)OR.sup.a, C(.dbd.O)N(R.sup.a).sub.2,
C(.dbd.O)NR.sup.aC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneC(.dbd.O)OR.sup.a,
OC.sub.1-6alkyleneN(R.sup.a).sub.2, OC.sub.1-6alkyleneOR.sup.a,
OC.sub.1-6alkyleneNR.sup.aC(.dbd.O)OR.sup.a,
NR.sup.aC.sub.1-6alkyleneN(R.sup.a).sub.2,
NR.sup.aC(.dbd.O)R.sup.a, NR.sup.aC(.dbd.O)N(R.sup.a).sub.2,
N(SO.sub.2C.sub.1-6alkyl).sub.2, NR.sup.a(SO.sub.2C.sub.1-6alkyl),
nitro, and SO.sub.2N(R.sup.a).sup.2, and
R.sup.a is defined above;
L is selected from the group consisting of null, C.sub.1-8alkylene,
R.sup.a substituted C.sub.1-8alkylene, NR.sup.a, C(.dbd.O), aryl,
C(.dbd.O)aryl, C(.dbd.O)C.sub.1-6alkylene,
C.sub.1-8alkyleneNR.sup.a,
C.sub.1-6alkylenearyleneC.sub.1-6alkylene, C.sub.2-6alkenylene,
C.sub.4-8alkdienylene, C.sub.1-6alkylenearylene,
C.sub.1-6alkyleneheteroarylene, R.sup.a substituted
C.sub.1-6alkyleneheteroarylene, and
C.sub.2-6alkenylenearyleneC.sub.1-6alkylene, and R.sup.a is defined
above;
M is selected from the group consisting of
--C(.dbd.O)N(R.sup.b)OH,
--O(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
--N(R.sup.b)(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
--N(R.sup.b)(CH.sub.2).sub.1-6C(.dbd.O)N(R.sup.b)OR.sup.b,
arylC(.dbd.O)NHOH,
--N(OH)C(.dbd.O)R.sup.b,
heteroarylC(.dbd.O)NHOH,
--C.sub.3-6cycloalkylN--C(.dbd.O)CH.sub.2SH,
--B(OR.sup.b).sub.2,
--SO.sub.2NHR.sup.b,
--NHSO.sub.2NHR.sup.b,
--NHSO.sub.2C.sub.1-6alkyl,
--SO.sub.2C.sub.1-6alkyl,
--SR.sup.c,
##STR00027##
--C(.dbd.O)R.sub.e,
--P(.dbd.O)(OR.sup.f).sub.2,
--NH--P(.dbd.O)(OR.sup.f).sub.2,
##STR00028##
--C(.dbd.O)(C(R.sup.b).sup.2).sub.1-6SH,
--C(.dbd.O)C(.dbd.O)NHR.sup.b,
--C(.dbd.O)NHN(R.sup.b).sub.2,
--C(.dbd.O)NH(CH.sub.2).sub.1-3N(R.sup.b).sub.2,
##STR00029##
--S--(C.dbd.O)C.sub.1-6alkyl,
C.sub.3-10heterocycloalkyl optionally substituted with oxo
(.dbd.O), thioxo (.dbd.S), or both, aryl optionally substituted
with one or more of C.sub.1-6alkyl, --C(.dbd.O)R.sup.d, --NH.sub.2,
and --SH,
heteroaryl optionally substituted with --NH.sub.2, --SH, or
both,
--N(H)C(.dbd.O)SH,
--NHC(.dbd.O)NHR.sup.d,
--NHC(.dbd.O)CH.sub.2R.sup.d,
--NHC(.dbd.O)(CH.sub.2).sub.1-6SH,
--NHC(.dbd.O)CH.sub.2Hal,
--NHC(.dbd.S)NHR.sup.d,
--NHC(.dbd.S)CH.sub.2R.sup.d,
--C(.dbd.S)NHR.sup.d,
--C(.dbd.S)CH.sub.2R.sup.d,
--NHC(.dbd.S)CH.sub.2R.sup.d,
--NHC(.dbd.S)CH.sub.2Hal, and
--(C.dbd.O)C.sub.1-6alkyl;
R.sup.b, independently, is selected from the group consisting of
hydrogen, (C.dbd.O)CH.sub.3, C.sub.1-6alkyl, CF.sub.3, CH.sub.2F,
and aryl, or two R.sup.b groups are taken together with the carbon
to which they attached to form a C.sub.3-8cycloalkyl group;
R.sup.c is selected from hydrogen or (C.dbd.O)CH.sub.3;
R.sup.d is NH.sub.2 or OH;
R.sup.e is selected from the group consisting of OH,
N(R.sup.b).sup.2, NH(OCH.sub.3), N(CH.sub.3)OH, C.sub.1-6alkyl,
CF.sub.3, aryl, heteroaryl, C.sub.3-8cycloalkyl,
NHSO.sub.2CH.sub.3, NHSO.sub.2CF.sub.3, and C.sub.1-6haloalkyl;
R.sup.f independently is hydrogen, methyl, or ethyl; and
Hal is halo,
or a pharmaceutically acceptable salt, hydrate, or prodrug
thereof.
Compounds of the present invention inhibit HDAC and are useful in
the treatment of a variety of diseases and conditions. In
particular, the present HDACIs are used in methods of treating a
disease or condition wherein inhibition of HDAC provides a benefit,
for example, cancers, neurological diseases, neurodegenerative
conditions, peripheral neuropathies, autoimmune diseases,
inflammatory diseases and conditions, stroke, hypertension,
traumatic brain injury, autism, and malaria. The methods comprise
administering a therapeutically effective amount of a present HDACI
to an individual in need thereof.
The present methods also encompass administering a second
therapeutic agent to the individual in addition to a present HDACI.
The second therapeutic agent is selected from agents, such as drugs
and adjuvants, known as useful in treating the disease or condition
afflicting the individual, e.g., a chemotherapeutic agent and/or
radiation known as useful in treating a particular cancer.
As used herein, the term "alkyl" refers to straight chained and
branched saturated hydrocarbon groups, nonlimiting examples of
which include methyl, ethyl, and straight chain and branched
propyl, butyl, pentyl, hexyl, heptyl, and octyl groups containing
the indicated number of carbon atoms. The term C.sub.n means the
alkyl group has "n" carbon atoms.
The term "alkylene" refers to a bidentate moiety obtained by
removing two hydrogen atoms from an alkane. An "alkylene" is
positioned between two other chemical groups and serves to connect
them. An example of an alkylene group is --(CH.sub.2).sub.n--. An
alkyl, e.g., methyl, or alkylene, e.g., --CH.sub.2CH.sub.2--, group
can be substituted, independently, with one or more of halo,
trifluoromethyl, trifluoromethoxy, hydroxy, alkoxy, nitro, cyano,
alkylamino, and amino groups, for example.
The term "alkenyl" is defined identically as "alkyl," except for
containing a carbon-carbon double bond, e.g., ethenyl, propenyl,
and butenyl. The term "alkenylene" is defined identically to
"alkylene" except for containing a carbon-carbon double bond. The
term "alkdienylene" is defined identically as "alkenylene" except
the group contains two carbon-carbon double bonds, either
conjugated or non-conjugated.
The term "heteroalkyl" refers to an alkyl group having one or more,
and typically one to three, heteroatoms in the carbon chain of the
alkyl group. The heteroatoms, independently, are selected from O,
S, and NR, wherein R is hydrogen, alkyl, cycloalkyl,
heterocycloalkyl, aryl, and heteroaryl. A term such as
"C.sub.1-6heteroalkyl" means that the group contains 1 to 6 carbon
atoms in addition to the heteroatoms.
The term "perfluoroalkyl" is defined as an alkyl group wherein all
hydrogen atoms are replaced by fluorine atoms.
As used herein, the term "halo" and "Hal" are defined as fluoro,
chloro, bromo, and iodo.
The term "hydroxy" is defined as --OH.
The term "alkoxy" is defined as --OR, wherein R is alkyl. The term
"perfluoroalkoxy" is defined as an alkoxy group wherein all
hydrogen atoms are replaced by fluorine atoms.
The term "amino" is defined as --NR.sub.2, wherein each R group,
independently, is hydrogen, alkyl, cycloalkyl, heterocycloalkyl,
C.sub.1-3alkylenearyl, heteroaryl, or aryl, or both R groups are
taken together with the N to which they are attached to form a 4 to
8 membered ring.
The term "nitro" is defined as --NO.sub.2.
The term "cyano" is defined as --CN.
The term "trifluoromethyl" is defined as --CF.sub.3.
The term "trifluoromethoxy" is defined as --OCF.sub.3.
The term "Ac" is defined as --C(.dbd.O)CH.sub.3.
The term "tBu" is defined as tertiary butyl, i.e.
--C(CH.sub.3).sub.3.
The term "Boc" is defined as tert-butoxycarbonyl.
As used herein, compounds such as
##STR00030## is an abbreviation for
##STR00031## In addition, compounds such as
##STR00032## is an abbreviation for
##STR00033##
As used herein, groups such as C.sub.1-3alkylphenyl means a
C.sub.1-3alkyl group bonded to a phenyl ring, for example,
##STR00034## Groups such as C.sub.1-3alkylenephenyl means a phenyl
group bonded to a C.sub.1-3alkylene group, for example,
##STR00035##
As used herein, the term "aryl" refers to a monocyclic aromatic
group, e.g., phenyl. Unless otherwise indicated, an aryl group can
be unsubstituted or substituted with one or more, and in particular
one to five, groups independently selected from, for example, halo,
alkyl, alkenyl, --OCF.sub.3, --NO.sub.2, --CN, --NC, --OH, alkoxy,
amino, alkylamino, --CO.sub.2H, --CO.sub.2alkyl, alkynyl,
cycloalkyl, nitro, sulfhydryl, imino, amido, phosphonate,
phosphinate, silyl, alkylthio, sulfonyl, sulfonamide, aldehyde,
heterocycloalkyl, trifluoromethyl, aryl, and heteroaryl. Exemplary
aryl groups include, but are not limited to, phenyl, chlorophenyl,
methylphenyl, methoxyphenyl, trifluoromethylphenyl, nitrophenyl,
2,4-methoxychlorophenyl, and the like.
The term "arylene" refers to a bidentate aryl group that bonds to
two other groups and serves to connect these groups, e.g.,
##STR00036##
The term "C.sub.1-4alkylenearyleneC.sub.1-4alkylene" means
##STR00037## and serves to connect two other groups.
The term "C.sub.1-6alkylenearylene" means
##STR00038## and serves to connect two other groups.
The term "C.sub.2-6alkenylenearyleneC.sub.1-4alkylene" means
##STR00039## and serves to connect two other groups.
As used herein, the term "heteroaryl" refers to a monocyclic ring
system containing at least one nitrogen, oxygen, or sulfur atom in
an aromatic ring. Unless otherwise indicated, a heteroaryl group
can be unsubstituted or substituted with one or more, and in
particular one to four, substituents selected from, for example,
halo, alkyl, alkenyl, --OCF.sub.3, --NO.sub.2, --CN, --NC, --OH,
alkoxy, amino, alkylamino, --CO.sub.2H, --CO.sub.2alkyl, alkynyl,
cycloalkyl, nitro, sulfhydryl, imino, amido, phosphonate,
phosphinate, silyl, alkylthio, sulfonyl, sulfonamide, aldehyde,
heterocycloalkyl, trifluoromethyl, aryl, and heteroaryl. Examples
of heteroaryl groups include, but are not limited to, thienyl,
furyl, oxazolyl, thiophenyl, triazolyl, isothiazolyl, isoxazolyl,
imidazolyl, pyrimidinyl, thiazolyl, thiadiazolyl, pyridinyl,
pyridazinyl, pyrazolyl, pyrazinyl, tetrazolyl, oxazolyl, pyrrolyl,
and triazinyl.
As used herein, the term "C.sub.3-8cycloalkyl" means a monocyclic
aliphatic ring containing three to eight carbon atoms, either
saturated or unsaturated.
As used herein, the term "heterocycloalkyl" means a monocyclic or a
bicyclic aliphatic ring containing 3 to 10 total atoms, either
saturated or unsaturated, of which one to five of the atoms are
independently selected from nitrogen, oxygen, and sulfur and the
remaining atoms are carbon.
In accordance with the present invention, bicyclic ring system
##STR00040## can be, for example, a residue of:
##STR00041## ##STR00042##
The linker L can be attached to any atom D, E, F, G, W, X, Y, or Z,
and in some preferred embodiments the linker L is attached to a
nitrogen atom of the bicyclic ring system.
In accordance with the present invention, ring
##STR00043## is a five- or six-membered, aliphatic or aromatic
ring. For example, the ring can be
##STR00044## ##STR00045##
The linker L can be attached to any atom U, V, W, X, Y, or Z, and
preferably is attached to a nitrogen or carbon atom of the
ring.
In some embodiments, a bicyclic Cap group has a structure:
##STR00046## either unsubstituted or substituted with one or more
R.sup.1.
In some embodiments, a monocyclic Cap group has a structure:
##STR00047## either unsubstituted or substituted with one or more
R.sup.1 group.
In some embodiments, an alicyclic Cap group has a structure wherein
A is N, e.g.,
##STR00048##
In some embodiments, R.sup.1 substituents on the Cap group, if
present at all, preferably are null, hydrogen, OR.sup.a, halo,
C.sub.1-6alkyl, aryl, heterocycloalkyl,
--(CH.sub.2).sub.1-4heterocycloalkyl,
--(CH.sub.2).sub.1-4N(R.sup.a).sub.2,
##STR00049##
In some embodiments, R.sup.a is hydrogen, C.sub.1-6alkyl,
C.sub.1-6heteroalkyl, and heteroaryl.
In other preferred embodiments, L is null,
--(CH.sub.2).sub.1-6--,
##STR00050## optionally substituted with halo, CF.sub.3, or CN,
##STR00051## --CH.sub.2--CH.dbd.CH--CH.dbd.CH--,
##STR00052## --(CH.sub.2).sub.2--CH.dbd.CH--CH.dbd.CH.sub.2--,
--(CH.sub.2).sub.0-6--NH--,
##STR00053##
In still other preferred embodiments, M is
##STR00054## --C(.dbd.O)CH.sub.2SH, --CH.sub.2SH,
##STR00055## --SC(.dbd.O)tBu, --SC(.dbd.O)CF.sub.3,
--S(CH.sub.2).sub.1-3C(.dbd.O)CF.sub.3, --CH.sub.2SAc,
##STR00056## --NH--C(.dbd.O)CH.sub.2SH,
##STR00057## ##STR00058##
Additionally, salts, prodrugs, hydrates, isotopically labeled,
fluorescently labeled and any other therapeutically or
diagnostically relevant derivations of the present HDACIs also are
included in the present invention and can be used in the methods
disclosed herein. The present invention further includes all
possible stereoisomers and geometric isomers of the present
compounds. The present invention includes both racemic compounds
and optically active isomers. When a present HDACI is desired as a
single enantiomer, it can be obtained either by resolution of the
final product or by stereospecific synthesis from either
isomerically pure starting material or use of a chiral auxiliary
reagent, for example, see Z. Ma et al., Tetrahedron: Asymmetry,
8(6), pages 883-888 (1997). Resolution of the final product, an
intermediate, or a starting material can be achieved by any
suitable method known in the art. Additionally, in situations where
tautomers of a present compound is possible, the present invention
is intended to include all tautomeric forms of the compounds.
Prodrugs of the present compounds also are included in the present
invention. It is well established that a prodrug approach, wherein
a compound is derivatized into a form suitable for formulation
and/or administration, then released as a drug in vivo, has been
successfully employed to transiently (e.g., bioreversibly) alter
the physicochemical properties of the compound (see, H. Bundgaard,
Ed., "Design of Prodrugs," Elsevier, Amsterdam, (1985); R. B.
Silverman, "The Organic Chemistry of Drug Design and Drug Action,"
Academic Press, San Diego, chapter 8, (1992); K. M. Hillgren et
al., Med. Res. Rev., 15, 83 (1995)). Specific prodrugs of HDACIs
are discussed in WO 2008/055068, incorporated in its entirety
herein by reference.
Compounds of the present invention can contain one or more
functional groups. The functional groups, if desired or necessary,
can be modified to provide a prodrug. Suitable prodrugs include,
for example, acid derivatives, such as amides and esters. It also
is appreciated by those skilled in the art that N-oxides can be
used as a prodrug.
Compounds of the invention can exist as salts. Pharmaceutically
acceptable salts of the present HDACIs often are preferred in the
methods of the invention. As used herein, the term
"pharmaceutically acceptable salts" refers to salts or zwitterionic
forms of the present compounds. Salts of the present compounds can
be prepared during the final isolation and purification of the
compounds or separately by reacting the compound with an acid
having a suitable cation. The pharmaceutically acceptable salts of
the present compounds can be acid addition salts formed with
pharmaceutically acceptable acids. Examples of acids which can be
employed to form pharmaceutically acceptable salts include
inorganic acids such as nitric, boric, hydrochloric, hydrobromic,
sulfuric, and phosphoric, and organic acids such as oxalic, maleic,
succinic, tartaric, and citric. Nonlimiting examples of salts of
compounds of the invention include, but are not limited to, the
hydrochloride, hydrobromide, hydroiodide, sulfate, bisulfate,
2-hydroxyethansulfonate, phosphate, hydrogen phosphate, acetate,
adipate, alginate, aspartate, benzoate, bisulfate, butyrate,
camphorate, camphorsulfonate, digluconate, glycerolphosphate,
hemisulfate, heptanoate, hexanoate, formate, succinate, fumarate,
maleate, ascorbate, isethionate, salicylate, methanesulfonate,
mesitylenesulfonate, naphthylenesulfonate, nicotinate,
2-naphthalenesulfonate, oxalate, pamoate, pectinate, persulfate,
3-phenylproprionate, picrate, pivalate, propionate,
trichloroacetate, trifluoroacetate, phosphate, glutamate,
bicarbonate, paratoluenesulfonate, undecanoate, lactate, citrate,
tartrate, gluconate, methanesulfonate, ethanedisulfonate, benzene
sulphonate, and p-toluenesulfonate salts. In addition, available
amino groups present in the compounds of the invention can be
quaternized with methyl, ethyl, propyl, and butyl chlorides,
bromides, and iodides; dimethyl, diethyl, dibutyl, and diamyl
sulfates; decyl, lauryl, myristyl, and stearyl chlorides, bromides,
and iodides; and benzyl and phenethyl bromides. In light of the
foregoing, any reference to compounds of the present invention
appearing herein is intended to include the present compounds as
well as pharmaceutically acceptable salts, hydrates, or prodrugs
thereof.
The present compounds also can be conjugated or linked to auxiliary
moieties that promote a beneficial property of the compound in a
method of therapeutic use. Such conjugates can enhance delivery of
the compounds to a particular anatomical site or region of interest
(e.g., a tumor), enable sustained therapeutic concentrations of the
compounds in target cells, alter pharmacokinetic and
pharmacodynamic properties of the compounds, and/or improve the
therapeutic index or safety profile of the compounds. Suitable
auxiliary moieties include, for example, amino acids,
oligopeptides, or polypeptides, e.g., antibodies, such as
monoclonal antibodies and other engineered antibodies; and natural
or synthetic ligands to receptors in target cells or tissues. Other
suitable auxiliaries include fatty acid or lipid moieties that
promote biodistribution and/or uptake of the compound by target
cells (see, e.g., Bradley et al., Clin. Cancer Res. (2001)
7:3229).
Specific compounds of the present invention include, but are not
limited to,
##STR00059## ##STR00060## ##STR00061## ##STR00062##
Synthetic Methods
The following synthetic schemes are representative of the reactions
used to synthesize the present HDACIs. Modifications and alternate
schemes to prepare HDACIs of the invention are readily within the
capabilities of persons skilled in the art.
In the synthetic methods, the examples, and throughout the
specification, the abbreviations have the following meanings:
TABLE-US-00002 DMF dimethylformamide min minutes TLC thin layer
chromatography CH.sub.2Cl.sub.2 methylene chloride MeOH methanol
Na.sub.2SO.sub.4 sodium sulfate AcOH acetic acid MS mass
spectrometry Na.sub.2CO.sub.3 sodium carbonate HPLC high
performance liquid chromatography H or hrs hours NaHCO.sub.3 sodium
bicarbonate HCl hydrochloric acid g gram mol mole mmol millimole mL
milliliter H.sub.2SO.sub.4 sulfuric acid NaH sodium hydride TMS
tetramethylsilane TFA trifluoroacetic acid KOH potassium hydroxide
NH.sub.4Cl ammonium chloride NH.sub.2OH.cndot.HCl hydroxylamine
hydrochloride NaOMe sodium methoxide CD.sub.3OD deuterated methanol
M molar KOtBu potassium tert-butoxide DMSO dimethyl sulfoxide KOH
potassium hydroxide NaCNBH.sub.3 sodium cyanoborohydroxide N normal
KI potassium iodide SOCl.sub.2 thionyl chloride CD.sub.3CN
deuterated acetonitrile RT room temperature DME dimethyl ether
ZnCl.sub.2 zinc chloride CuI copper iodide NMR nuclear magnetic
resonance spectrometry EtOAc ethyl acetate THF tetrahydrofuran NaOH
sodium hydroxide PdCl.sub.2(PPh).sub.3
dichloro-triphenylphosphino-palladium (II) Et.sub.3N triethylamine
CDCl.sub.3 deuterated chloroform Hz Hertz
It should be understood that protecting groups can be utilized in
accordance with general principles of synthetic organic chemistry
to provide compounds of the present invention. Protecting
group-forming reagents are well known to persons skilled in the
art, for example, see T. W. Greene et al., "Protective Groups in
Organic Synthesis, Third Edition," John Wiley and Sons, Inc., NY,
N.Y. (1999). These protecting groups are removed when necessary by
appropriate basic, acidic, or hydrogenolytic conditions known to
persons skilled in the art. Accordingly, compounds of the present
invention not specifically exemplified herein can be prepared by
persons skilled in the art.
Synthetic Methods and Procedures
Procedures
General Information for Synthetic Procedures: .sup.1H NMR and
.sup.13C NMR spectra were obtained using a Bruker spectrometer with
TMS as an internal standard. The following standard abbreviations
indicating multiplicity were used: s=singlet, d=doublet, t=triplet,
q=quartet, quint=quintet, m=multiplet, dd=double doublet, dt=double
triplet, and br=broad. LRMS experiments were carried out using an
Agilent 1100 series LC/MSD instrument with MeCN and H.sub.2O spiked
with 0.1% formic acid as the mobile phase. HRMS experiments were
carried out using a Shimadzu IT-TOF instrument with MeCN and
H.sub.2O spiked with 0.1% formic acid as the mobile phase. Reaction
progress was monitored by TLC using precoated silica gel plates
(Merck silica gel 60 F254, 250 .mu.m thickness). Automated column
chromatography was performed using the CombiFlash Rf apparatus
available from Teledyne ISCO and prepacked cartridges (25 or 50 g)
loaded with Merck silica gel (40-60 mesh) along with the following
conditions: Method 1: 100% hexane, 5 min; 0-50% EtOAc/hexane, 25
min; 50% EtOAc/hexane, 5 min. Method 2: 100% DCM, 5 min; 0-10%
MeOH/DCM, 20 min, 10% MeOH/DCM, 5 min. Flow rate=30-40 mL/min
(depending on cartridge size) with wavelength monitoring at 254 and
280 nm. Preparatory HPLC was carried out using a Shimadzu
preparative liquid chromatograph with the following specifications:
Column: ACE 5 AQ (150.times.21.2 mm) with 5 .mu.m particle size.
Method 1: 25-100% MeOH/H.sub.2O, 30 min; 100% MeOH, 5 min; 100-25%
MeOH/H.sub.2O, 4 min; 25% MeOH/H.sub.2O, 1 min. Method 2: 8-100%
MeOH/H.sub.2O, 30 min; 100% MeOH, 5 min; 100-8% MeOH/H.sub.2O, 4
min; 8% MeOH/H.sub.2O, 1 min. Flow rate=17 mL/min with wavelength
monitoring at 254 and 280 nm. Both solvents were spiked with 0.05%
TFA. Where applicable (unless otherwise specified), resin bound
bicarbonate was used to neutralize the trifluoroacetic acid salts
obtained during preparatory HPLC purification. Analytical HPLC was
carried out using an Agilent 1100 series instrument with the
following specifications: Column: Luna 5 .mu. C18(2) 100A
(150.times.4.60 mm) with 5 .mu.m particle size. Flow rate=1.4
mL/min with wavelength monitoring at 254 nm. Gradient: 10-100%
MeOH/H.sub.2O, 18 min; 100% MeOH, 3 min; 100-10% MeOH/H.sub.2O, 3
min; 10% MeOH/H.sub.2O, 5 min. Both solvents were spiked with 0.05%
TFA.
A. General synthetic scheme for HDACI compounds containing a
bicyclic Cap group:
##STR00063##
B. General synthetic scheme for HDACI compound containing a
five-membered monocyclic Cap group:
##STR00064##
C. General synthetic scheme for HDACI compounds containing a
six-membered monocyclic Cap group:
##STR00065##
D. General synthetic scheme for HDACI compounds containing an
acyclic Cap group:
##STR00066##
General Procedure A: NaH (1 mol equiv) was dissolved in anhydrous
DMF (5 mL/mmol) under argon and cooled to 0.degree. C. To it was
added the appropriate bicyclic Cap group (1 mol equiv) dissolved in
anhydrous DMF (3 mL/mmol). The reaction was stirred for 15 min at
0.degree. C. followed by the addition of methyl
4-(bromomethyl)benzoate (1 mol equiv) in anhydrous DMF (2 mL/mmol).
The reaction was stirred for 2 h at 70.degree. C. and then quenched
by the addition of H.sub.2O (30 mL). The organic products were
extracted with EtOAc (3.times.30 mL), washed with H.sub.2O
(2.times.30 mL), brine (15 mL), dried over Na.sub.2SO.sub.4,
filtered and concentrated in vacuo.
General Procedure B: The appropriate ester was dissolved/suspended
in MeOH (3 mL/mmol) and added to a mixture of NH.sub.2OH.HCl (6 mol
equiv) in MeOH (3 mL/mmol) which was followed by the addition of
NaOMe (8 mol equiv of a 25% solution in MeOH). The mixture was
stirred for 2 h at 0.degree. C. followed by stirring for 22 h at
RT. When the reaction was complete as evidenced by TLC, the
reaction was quenched by the addition of trifluoroacetic acid (5
mol equiv of a 10% solution in DCM), filtered, and the filter cake
was washed with additional MeOH (5-15 mL). The combined filtrate
and wash were then concentrated in vacuo to yield the crude product
which was dissolved in DMF and purified by preparatory HPLC.
General Procedure C: To a round bottom flask charged with methyl
4-(bromomethyl)benzoate (1 mol equiv) in DMF (4 mL/mmol) was added
the appropriate 2-subsitituted benzimidazole (1 mol equiv) and
K.sub.2CO.sub.3 (2 mol equiv) in succession. The resulting mixture
was allowed to stir at 80.degree. C. for 2-16 hrs and the reaction
was monitored by TLC. Upon completion, the reaction was quenched
with H.sub.2O (20 mL) and extracted with EtOAc (3.times.20 mL). The
combined organic extracts were washed with H.sub.2O (2.times.15
mL), brine (15 mL), dried over Na.sub.2SO.sub.4, filtered and
concentrated in vacuo.
General Procedure D: Solid NaOH (8 mol equiv) was dissolved in a
50% aq. solution of NH.sub.2OH (.about.50 mol equiv) at 0.degree.
C. Then, a solution of the appropriate ester (1 mol equiv) in
THF/MeOH (9:9 mL/mmol) was added dropwise to the aforementioned,
vigorously stirred hydroxylamine solution. Upon complete addition,
the ice bath was removed and the reation was allowed to stir 15
min. The reaction was quenched with AcOH (10 mol equiv) and
concentrated in vacuo to yield the crude product which was
dissolved in DMF and purified by preparatory HPLC.
General Procedure E: The appropriate boronic acid (1 mol equiv),
methyl 4-(bromomethyl)benzoate (1.2 mol equiv),
tetrakis(triphenylphosphine)Pd(0) (0.02 mol equiv), and
K.sub.2CO.sub.3 (2.1 mol equiv) were placed in a dry, sealed tube
under Ar atmosphere. Diglyme (4 mL/mmol) and H.sub.2O (2 mL/mmol)
were added through a rubber septa which was immediately replaced by
the screw on cap. The reaction was heated to 100.degree. C. and
stirred for 4 h. Then, the reaction was diluted with H.sub.2O (20
mL) and the organic products were extracted with DCM (3.times.15
mL), washed with brine (15 mL), dried over Na.sub.2SO.sub.4,
filtered and concentrated in vacuo.
An example of a present HDACI having a bicyclic Cap group is
prepared as follows (see also General Synthetic Scheme A):
##STR00067##
Examples of bicyclic Cap groups include, but are not limited
to:
##STR00068##
HDACIs having a bicyclic Cap group:
Methyl 4-((1H-indol-1-yl)methyl)benzoate (1)
##STR00069##
The title compound was prepared from 1H-indole (0.500 g, 4.27 mmol)
according to General Procedure A and purified using automated
column chromatography method 1. The product was isolated as a white
solid (0.860 g, 76%). .sup.1H NMR (400 MHz, CDCl.sub.3): .delta.
8.05 (d, J=8.23 Hz, 2H), 7.79 (d, J=6.9 Hz, 1H), 7.24 (m, 6H), 6.70
(d, J=3.05 (d, J=3.05 Hz, 1H), 5.35 (s, 2H), 3.97 (s, 3H). .sup.13C
NMR (100 MHz, CDCl.sub.3): .delta. 166.7, 142.9, 136.3, 130.1,
129.6, 128.9, 128.4, 126.7, 122.0, 121.2, 119.9, 109.7, 102.2,
52.2, 49.8. ESI-HRMS: calc. for C.sub.17H.sub.15NO.sub.2:
[M+H].sup.+=266.1176 m/z, found: [M+H].sup.+=266.1182 m/z.
4-((1H-Indol-1-yl)methyl)-N-hydroxybenzamide (2)
##STR00070##
The title compound was synthesized from methyl
4-((1H-indol-1-yl)methyl)benzoate 1 (0.840 g, 3.17 mmol) according
to General Procedure B (prep. HPLC method 1) and isolated as a
white solid (0.443 g, 53%). .sup.1H NMR (400 MHz, DMSO-d.sub.6):
.delta. 11.14 (S, 1H), 9.01 (br, 1H), 7.67 (d, J=8.1 Hz, 2H), 7.56
(d, J=7.77 Hz, 1H), 7.52 (d, J=3.1 Hz, 1H), 7.42 (d, J=8.1 Hz, 1H),
7.23 (d, J=8.1 Hz, 2H), 7.09 (t, J=7.2 Hz, 1H), 7.02 (t, J=7.5 Hz,
1H), 6.50 (d, J=3.0 Hz, 1H), 5.48 (s, 2H). .sup.13C NMR (100 MHz,
DMSO-d.sub.6): .delta. 164.4, 141.9, 136.1, 132.3, 129.6, 128.7,
127.6, 127.3, 121.7, 120.9, 119.6, 110.5, 101.6, 49.2. ESI-HRMS:
calc. for C.sub.16H.sub.14N.sub.2O.sub.2: [M+H].sup.+=267.1128 m/z,
found: [M+H].sup.+=267.1137 m/z.
Methyl 4-((2-methyl-1H-indol-1-yl)methyl)benzoate (3)
##STR00071##
The title compound was prepared from 2-methyl-1H-indole (2.00 g,
15.3 mmol) according to General Procedure A and purified using
automated column chromatography method 1. The product was isolated
as a white solid (2.87 g, 67%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6): .delta. 7.89 (d, J=8.0 Hz, 2H), 7.48 (d, J=7.6 Hz,
1H), 7.33 (d, J=7.6 Hz, 1H), 7.09 (d, J=8.0 Hz, 2H), 7.01 (m, 2H),
6.32 (s, 1H), 5.49 (s, 2H), 3.81 (s, 3H), 2.34 (s, 3H). .sup.13C
NMR (100 MHz, DMSO-d.sub.6): .delta. 166.4, 144.5, 137.2, 137.1,
130.0, 128.9, 128.2, 126.8, 120.9, 119.8, 119.7, 109.9, 100.7,
52.5, 45.9, 12.8.
N-Hydroxy-4-((2-methyl-1H-indol-1-yl)methyl)benzamide (4)
##STR00072##
The title compound was synthesized from methyl
4-((2-methyl-1H-indol-1-yl)methyl)benzoate 3 (0.259 g, 0.937 mmol)
according to General Procedure B (prep. HPLC method 1) and isolated
as a light brown solid (86 mg, 33%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6): .delta. 11.13 (s, 1H), 8.96 (br, 1H), 7.65 (d, J=8.2
Hz, 2H), 7.46 (d, J=7.1 Hz, 1H), 7.33 (d, J=7.5 Hz, 1H), 7.00 (m,
4H), 6.31 (s, 1H), 5.45 (s, 2H), 2.36 (s, 3H). .sup.13C NMR (100
MHz, DMSO-d.sub.6): .delta. 166.0, 142.5, 136.6, 136.0, 128.9,
127.9, 127.0, 126.1, 120.6, 119.5, 119.4, 108.6, 100.5, 45.7, 12.3.
ESI-HRMS: calc. for C.sub.17H.sub.16N.sub.2O.sub.2:
[M+H].sup.+=281.1285 m/z, found: [M+H].sup.+=281.1283 m/z.
Methyl 4-((3-methyl-1H-indol-1-yl)methyl)benzoate (5)
##STR00073##
The title compound was prepared from 3-methyl-1H-indole (0.500 g,
3.81 mmol) according to General Procedure A and purified using
automated column chromatography method 1. The product was isolated
as a white solid (0.668 g, 63%). .sup.1H NMR (400 MHz, CDCl.sub.3):
.delta. 8.04 (d, J=7.9 Hz, 2H), 7.72 (d, J=7.4 Hz, 1H), 7.24 (m,
5H), 6.94 (s, 1H), 5.31 (s, 2H), 3.97 (s, 3H), 2.46 (s, 3H).
.sup.13C NMR (100 MHz, CDCl.sub.3): .delta. 166.7, 143.1, 136.5,
130.0, 129.4, 129.0, 126.6, 125.7, 1221.8, 119.1, 119.0, 109.3,
71.8, 52.1, 49.5, 9.6. ESI-HRMS: calc. for
C.sub.18H.sub.17NO.sub.2: [M+H].sup.+=280.1332 m/z, found:
[M+H].sup.+=280.1322 m/z.
N-Hydroxy-4-((3-methyl-1H-indol-1-yl)methyl)benzamide (6)
##STR00074##
The title compound was synthesized from methyl
4-((3-methyl-1H-indol-1-yl)methyl)benzoate 5 (0.200 g, 0.716 mmol)
according to General Procedure B (prep. HPLC method 1) and isolated
as a white solid (0.108 g, 54%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6): .delta. 11.13 (s, 1H), 8.99 (s, 1H), 7.66 (d, J=8.4
Hz, 2H), 7.50 (d, J=8.0 Hz, 1H), 7.37 (d, J=8.0 Hz, 1H), 7.23 (m,
3H), 7.06 (td, J=8.0, 1.2 Hz, 1H), 7.03 (td, J=7.6, 0.8 Hz, 1H),
5.39 (s, 2H), 2.27 (s, 3H). .sup.13C NMR (100 MHz, DMSO-d.sub.6):
.delta. 166.48, 142.50, 136.57, 131.08, 128.99, 126.97, 126.49,
125.78, 121.15, 118.39, 118.35, 110.36, 109.09, 48.62, 8.25.
ESI-HRMS: calc. for C.sub.17H.sub.16N.sub.2O.sub.2:
[M+H].sup.+=279.1139 m/z, found: [M+H].sup.+=279.1144 m/z.
Methyl 4-((2,3-dimethyl-1H-indol-1-yl)methyl)benzoate (7)
##STR00075##
The title compound was prepared from 2,3-dimethyl-1H-indole (0.500
g, 3.44 mmol) according to General Procedure A and purified using
automated column chromatography method 1. The product was isolated
as a viscous yellow oil (0.652 g, 65%)..sup.1H NMR (400 MHz,
CDCl.sub.3): .delta. 7.95 (d, J=8.3 Hz, 2H), 7.57 (dd, J=3.1 Hz,
2.2 Hz, 1H), 7.16 (m, 3H), 7.04 (d, J=8.2 Hz, 2H), 5.36 (s, 2H),
3.91 (s, 3H), 2.32 (s, 3H), 2.29 (s, 3H). .sup.13C NMR (100 MHz,
CDCl.sub.3): .delta. 166.8, 143.5, 136.3, 132.2, 130.1, 129.2,
128.7, 126.0, 121.0, 119.1, 118.1, 108.6, 107.4, 52.1, 46.4, 10.1,
8.9. ESI-HRMS: calc. for C.sub.19H.sub.19NO.sub.2:
[M+H].sup.+=294.1489 m/z, found: [M+H].sup.+=294.1503 m/z.
4-((2,3-Dimethyl-1H-indol-1-yl)methyl)-N-hydroxybenzamide (8)
##STR00076##
The title compound was synthesized from methyl
4-((2,3dimethyl-1H-indol-1-yl)methyl)benzoate 7 (0.632 g, 2.15
mmol) according to General Procedure B (prep. HPLC method 1) and
isolated as a white solid (0.487 g, 77%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6): .delta. 11.13 (s, 1H), 9.00 (br, 1H), 7.66 (d, J=8.2
Hz, 2H), 7.45 (d, J=6.9 Hz, 1H), 7.32 (d, J=7.3 Hz, 1H), 7.01 (m,
4H), 5.43 (s, 2H), 2.27 (s, 3H), 2.22 (s, 3H). .sup.13C NMR (100
MHz, DMSO-d.sub.6): .delta. 164.4, 142.4, 136.4, 133.0, 132.1,
128.6, 127.6, 126.5, 120.9, 119.0, 118.1, 109.6, 106.5, 45.9, 10.3,
9.2. ESI-HRMS: calc. for C.sub.18H.sub.18N.sub.2O.sub.2:
[M+H].sup.+=295.1441 m/z, found: [M+H].sup.+=295.1453 m/z.
Methyl 4-((3-benzyl-1H-indol-1-yl)methyl)benzoate (9)
##STR00077##
To a solution of methyl 4-((1H-indol-1-yl)methyl)benzoate 1 (0.250
g, 0.94 mmol) and benzaldehyde (96 .mu.L, 0.94 mmol) in DCM (10 mL)
at 0.degree. C. was added SiEt.sub.3H (0.45 mL, 2.83 mmol) followed
by trifluoroacetic acid (0.14 mL, 1.88 mmol). The reaction was
stirred at 0.degree. C. for 1 h. The reaction was then adjusted to
pH 10 with 2 N NaOH and the organic products were extracted with
DCM (3.times.15 mL). The combined organic fractions were washed
with brine (15 mL), dried with Na.sub.2SO.sub.4, filtered and
concentrated in vacuo. The title compound was isolated using
automated column chromatography method 1 (0.262 g, 78%). .sup.1H
NMR (400 MHz, CDCl.sub.3): .delta. 7.97 (d, J=8.0 Hz, 2H), 7.56 (d,
J=8.0 Hz, 1H), 7.32-7.27 (m, 4H), 7.23-7.10 (m, 6H), 6.87 (s, 1H),
5.33 (s, 2H), 4.15 (s, 2H), 3.91 (s, 3H). .sup.13C NMR (100 MHz,
CDCl.sub.3): .delta. 166.8, 143.0, 141.2, 136.8, 130.1, 129.5,
128.7, 128.4, 128.3, 126.6, 126.5, 126.0, 122.1, 119.5, 119.3,
115.4, 109.6, 52.2, 49.7, 31.6.
4-((3-Benzyl-1H-indol-1-yl)methyl)-N-hydroxybenzamide (10)
##STR00078##
The title compound was synthesized from methyl
4-(3-benzyl-1H-indol-1-yl)methyl)benzoate 9 (0.200 g, 0.563 mmol)
according to General Procedure B (prep. HPLC method 1) and isolated
as a white solid (80 mg, 40%). .sup.1H NMR (400 MHz, DMSO-d.sub.6):
.delta. 11.20 (br, 1H), 7.67 (d, J=8.2 Hz, 2H), 7.45 (d, J=7.9 Hz,
1H), 7.36 (d, J=8.2 Hz, 1H), 7.27 (m, 6H), 7.15 (d, J=6.7 Hz, 1H),
7.07 (t, J=7.9 Hz, 1H), 6.96 (t, J=7.8 Hz, 1H), 5.40 (s, 2H), 4.05
(s, 2H). .sup.13C NMR (100 MHz, MeOD): .delta. 164.48, 142.06,
141.91, 136.63, 132.18, 128.83, 128.68, 128.06, 127.57, 127.41,
127.29, 126.17, 121.83, 119.44, 119.19, 114.53, 110.45, 49.01,
31.30. ESI-HRMS: calc. for C.sub.23H.sub.20N.sub.2O.sub.2:
[M+H].sup.+=357.1598 m/z, found: [M+H].sup.+=357.1597 m/z.
Methyl 4-((9H-purin-9-yl)methyl)benzoate (11)
##STR00079##
The title compound was prepared from 9H-purine (0.500 g, 3.81 mmol)
according to General Procedure A (substituting KO.sup.tBu for NaH)
and purified using automated column chromatography method 2. The pH
was adjusted to 10 with 1 N NaOH prior to extraction with EtOAc.
The product was isolated as a white solid (0.287 g, 64%). .sup.1H
NMR (400 MHz, DMSO-d.sub.6): .delta. 9.20 (s, 1H), 8.94 (s, 1H),
8.78 (s, 1H), 7.92 (d, J=8.3 Hz, 2H), 7.44 (d, J=8.3 Hz, 2H), 5.62
(s, 2H), 3.82 (s, 3H). .sup.13C NMR (100 MHz, DMSO-d.sub.6):
.delta. 165.9, 152.3, 151.2, 148.0, 147.2, 141.7, 133.7, 129.7,
129.2, 127.8, 52.2, 46.1. ESI-HRMS: calc. for
C.sub.14H.sub.12N.sub.4O.sub.2: [M+H].sup.+=269.1033 m/z, found:
[M+H].sup.+=269.1032 m/z.
4-((9H-Purin-9-yl)methyl)-N-hydroxybenzamide (12)
##STR00080##
The title compound was synthesized from methyl
4-((9H-purin-9-yl)methyl)benzoate 11 (0.287 g, 1.07 mmol) according
to General Procedure B (prep. HPLC method 2) and isolated as a
white solid (288 mg, 45%). .sup.1H NMR (400 MHz, DMSO-d.sub.6):
.delta. 11.18 (br, 1H), 9.20 (s, 1H), 8.95 (s, 1H), 8.78 (s, 1H),
7.71 (d, J=8.2 Hz, 2H), 7.40 (d, J=8.2 Hz, 2H), 5.58 (s, 2H), 3.73
(br, 1H). .sup.13C NMR (100 MHz, DMSO-d.sub.6): .delta. 164.2,
152.6, 151.5, 148.4, 147.5, 139.8, 134.0, 132.9, 128.0, 127.8,
46.5. ESI-HRMS: calc. for C.sub.13H.sub.11N.sub.5O.sub.2:
[M+H].sup.+=270.0986 m/z, found: [M+H].sup.+=270.0992 m/z.
Methyl 4-((7H-purin-7-yl)methyl)benzoate (13)
##STR00081##
The title compound was prepared from 9H-purine (0.500 g, 3.81 mmol)
according to General Procedure A (substituting KO.sup.tBu for NaH)
and purified using automated column chromatography method 2. The pH
was adjusted to 10 with 1 N NaOH prior to extraction with EtOAc.
The product was isolated as a viscous yellow oil (0.108 g, 24%).
.sup.1H NMR (400 MHz, DMSO-d.sub.6): .delta. 9.16 (s, 1H), 9.02 (s,
1H), 8.97 (s, 1H), 7.94 (d, J=8.2 Hz, 2H), 7.51 (d, J=8.2 Hz, 2H),
5.74 (s, 2H), 3.83 (s, 3H). .sup.13C NMR (100 MHz, DMSO-d.sub.6):
.delta. 165.9, 160.4, 152.0, 150.2, 141.2, 140.6, 129.8, 129.5,
128.0, 125.0, 52.2, 48.4. ESI-HRMS: calc. for
C.sub.14H.sub.12N.sub.4O.sub.2: [M+H].sup.+=269.1033 m/z, found:
[M+H].sup.+=269.1036 m/z.
4-((7H-Purin-7-yl)methyl)-N-hydroxybenzamide (14)
##STR00082##
The title compound was synthesized from methyl
4-((7H-purin-7-yl)methyl)benzoate 13 (0.108 g, 0.40 mmol) according
to General Procedure B (prep. HPLC method 2) and isolated as a
white solid (50 mg, 46%). .sup.1H NMR (400 MHz, DMSO-d.sub.6):
.delta. 11.21 (br, 1H), 9.16 (s, 1H), 9.00 (s, 1H), 8.95 (s, 1H),
7.73 (d, J=8.2 Hz, 2H), 7.47 (d, J=8.2 Hz, 2H), 6.75 (br, 1H), 5.68
(s, 2H). .sup.13C NMR (100 MHz, DMSO-d.sub.6): .delta. 164.1,
160.7, 152.5, 150.3, 141.2, 139.3, 133.1, 128.2, 127.9, 125.3,
48.8. ESI-HRMS: calc. for C.sub.13H.sub.11N.sub.5O.sub.2:
[M+H].sup.+=270.0986 m/z, found: [M+H].sup.+=270.0984 m/z.
3-[2-(4-Methyl-piperazin-1-yl)-ethyl]-1H-indole (15)
##STR00083##
Tryptophol (0.500 g, 3.10 mmol) and Et.sub.3N (1 mL) were dissolved
in DCM (2 mL) and then mesyl chloride (0.24 mL, 3.102 mmol) was
added dropwise at RT. The reaction was stirred for 3 h and then
volatiles were removed in vacuo. The crude mesylate was taken up in
DCM (3 mL) and to it was added 1-methylpiperazine (1.72 mL, 15.5
mmol) and Et.sub.3N (1 mL). The reaction was heated to 40.degree.
C. and stirred overnight. After completion, the reaction mixture
was poured into cold water, the pH was adjusted to 10 with 1 N NaOH
and the organic products were extracted with DCM (3.times.15 mL).
The combined organic extracts were washed with brine (15 mL), dried
with Na.sub.2SO.sub.4, filtered and concentrated in vacuo.
Automated column chromatography method 2 was used to isolate the
title compound (0.42 g, 56%). .sup.1H NMR (400 MHz, CDCl.sub.3):
.delta. 7.54 (d, J=7.8 Hz, 1H), 7.34 (d, J=8.1 Hz, 1H), 7.10 (t,
J=7.1 Hz, 1H), 7.03 (t, J=7.3 Hz, 1H), 6.98 (s, 1H), 2.87 (t, J=7.8
Hz, 2H), 2.58 (t, J=5.9 Hz, 2H), 2.38 (br, 8H), 2.17 (s, 3H).
.sup.13C NMR (100 MHz, CDCl.sub.3): .delta. 136.7, 127.4, 121.8,
121.0, 118.3, 118.0, 112.4, 111.0, 58.9, 54.1, 52.2, 44.6, 22.2.
ESI-HRMS: calc. for C.sub.15H.sub.21N.sub.3: [M+H].sup.+=244.1808
m/z, found: [M+H].sup.+=244.1797 m/z.
4-{3-[2-(4-Methyl-piperazin-1-yl)-ethyl]-indol-1-ylmethyl}-benzoic
acid methyl ester (16)
##STR00084##
The title compound was prepared from
3-[2-(4-methylpiperazin-1-yl)-ethyl]-1H-indole 15 (0.400 g, 1.64
mmol) according to General Procedure A (substituting KO.sup.tBu for
NaH). The pH was adjusted to 10 with 1 N NaOH prior to extraction
with EtOAc. The product was purified via automated column
chromatography method 2 (0.412 g, 64%). .sup.1H NMR (400 MHz,
CDCl.sub.3): .delta. 7.79 (d, J=8.3 Hz, 2H), 7.56 (d, J=7.0 Hz,
1H), 7.04 (m, 6H), 5.14 (s, 2H), 3.76 (s, 3H), 2.90 (t, J=7.6 Hz,
2H), 2.63 (t, J=8.6 Hz, 2H), 2.46 (br, 8H), 2.23 (s, 3H). .sup.13C
NMR (100 MHz, CDCl.sub.3): .delta. 166.2, 143.3, 136.2, 129.1,
128.6, 127.8, 126.1, 125.4, 121.2, 118.4, 118.2, 112.6, 109.1,
58.4, 53.8, 51.9, 50.8, 48.4, 44.3, 21.8. ESI-HRMS: calc. for
C.sub.24H.sub.29N.sub.3O.sub.2: [M+H].sup.+=392.2333 m/z, found:
[M+H].sup.+=392.2343 m/z.
N-Hydroxy-4-{3-[2-(4-methyl-piperazin-1-yl)ethyl]-indol-1-ylmethyl}-benzam-
ide (17)
##STR00085##
The title compound was synthesized from
4-{3-[2-(4-Methyl-piperazin-1-yl)-ethyl]-indol-1-ylmethyl}-benzoic
acid methyl ester 16 (0.412 g, 1.05 mmol) according to General
Procedure B (prep. HPLC method 2) and isolated as a white solid
(0.265 g, 64%). .sup.1H NMR (400 MHz, DMSO-d.sub.6): .delta. 7.67
(d, J=7.8 Hz, 3H), 7.42 (m, 2H), 7.24 (d, J=7.9 Hz, 2H), 7.12 (t,
J=7.2 Hz, 1H), 7.04 (t, J=7.4 Hz, 1H), 5.43 (s, 2H), 3.81 (m, 4H),
3.53 (br, 4H), 3.40 (br, 2H), 3.25 (br, 2H), 2.84 (s, 3H). .sup.13C
NMR (100 MHz, DMSO-d.sub.6): .delta. 163.9, 141.4, 136.1, 131.9,
127.3, 127.2, 127.1, 127.0, 121.8, 119.0, 118.9, 110.3, 109.2,
55.6, 49.6, 48.7, 48.0, 42.2, 19.4. ESI-HRMS: calc. for
C.sub.23H.sub.28N.sub.4O.sub.2: [M+H].sup.+=393.2285 m/z, found:
[M+H].sup.+=393.2299 m/z.
2-(1H-Indol-3-yl)-N,N-dimethylethanamine
##STR00086##
.sup.1H NMR (400 MHz, CDCl.sub.3): .delta. 8.68 (s, 1H), 7.63 (d,
J=7.7 Hz, 1H), 7.35 (d, J=8.0 Hz, 1H), 7.21 (t, J=7.1 Hz, 1H), 7.14
(t, J=7.5 Hz, 1H), 6.97 (s, 1H), 3.00 (t, J=7.5 Hz, 2H), 2.72 (t,
J=8.4 Hz, 2H), 2.40 (s, 6H).
Methyl 4-((3-(2-(dimethylamino)ethyl)-1H-indol-1-yl)methyl)benzoate
(18)
##STR00087##
The title compound was prepared from
2-(1H-indol-3-yl)-N,N-dimethylethanamine (0.130 g, 0.69 mmol)
according to General Procedure A. The pH was adjusted to 10 with 1
N NaOH prior to extraction with EtOAc. The product was purified via
automated column chromatography method 2 (0.174 g, 61%). .sup.1H
NMR (400 MHz, CDCl.sub.3): .delta. 7.97 (d, J=8.3 Hz, 2H), 7.65 (d,
J=7.5 Hz, 1H), 7.17 (m, 5H), 6.99 (s, 1H), 5.34 (s, 2H), 3.91 (s,
3H), 3.04 (t, J=7.6 Hz, 2H), 2.75 (m, 2H), 2.43 (s, 6H).
4-((3-(2-(Dimethylamino)ethyl)-1H-indol-1-yl)methyl)-N-hydroxybenzamide
(19)
##STR00088##
The title compound was synthesized from methyl
4-((3-(2-(dimethylamino)ethyl)-1H-indol-1-yl)methyl)benzoate 18
(0.412 g, 1.05 mmol) according to General Procedure B (prep. HPLC
method 2) and isolated as a white solid (18 mg, 11%). .sup.1H NMR
(400 MHz, MeOD): .delta. 7.66 (m, 3H), 7.29 (m, 2H), 7.31 (d, J=8.0
Hz, 2H), 7.13 (m, 2H), 5.41 (s, 2H), 3.48 (t, J=7.4 Hz, 2H), 3.24
(t, J=8.3 Hz, 2H), 2.97 (s, 6H). .sup.13C NMR (100 MHz, MeOD):
.delta. 166.33, 141.99, 136.78, 131.29, 127.48, 127.05, 126.74,
126.65, 121.88, 119.23, 118.09, 109.73, 109.73, 108.70, 57.65,
48.90, 42.12, 20.35. ESI-HRMS: calc. for
C.sub.20H.sub.23N.sub.3O.sub.2: [M+H].sup.+=338.1863 m/z, found:
[M+H].sup.+=338.1865 m/z.
Methyl
4-((3-((dimethylamino)methyl)-2-methyl-1H-indol-1-yl)methyl)benzoat-
e (20)
##STR00089##
Methyl 4-((2-methyl-1H-indol-1-yl)methyl)benzoate 3 (0.500 g, 1.90
mmol) and dimethylformiminium chloride (0.200 g, 2.15 mmol) were
heated to reflux in a solution of DCM (5 mL) and DMF (2 mL) for 20
h. The reaction was then cooled to RT and DCM (50 mL) was added
along with sat. NaHCO.sub.3 (20 mL). The organic fraction was
isolated and the aqueous solution was further extracted with DCM
(2.times.20 mL). The combined organic fractions were washed with
H.sub.2O (20 mL), brine (20 mL), dried with Na.sub.2SO.sub.4 and
concentrated in vacuo. The desired product was purified via
automated column chromatography method 2 (0.221 g, 37%). .sup.1H
NMR (400 MHz, CDCl.sub.3): .delta. 7.94 (d, J=8.1 Hz, 2H), 7.69 (m,
1H), 7.14 (m, 3H), 7.02 (d, J=8.0 Hz, 2H), 5.38 (s, 2H), 3.90 (s,
3H), 3.62 (s, 2H), 2.35 (s, 3H), 2.30 (s, 6H).
4-((3-((Dimethylamino)methyl)-2-methyl-1
H-indol-1-yl)methyl)-N-hydroxybenzamide (21)
##STR00090##
The title compound was synthesized from methyl
4-((3-((dimethylamino)methyl)-2-methyl-1H-indol-1-yl)methyl)benzoate
20 (0.124 g, 1.78 mmol) according to General Procedure B (prep.
HPLC method 2) and isolated as a white solid (0.100 g, 80%).
.sup.1H NMR (400 MHz, MeOD): .delta. 7.72 (br, 1H), 7.68 (d, J=7.8
Hz, 2H), 7.38 (br, 1H), 7.21 (br, 2H), 7.08 (d, J=7.9 Hz, 2H), 5.56
(s, 2H), 4.57 (s, 2H), 2.91 (s, 6H), 2.48 (s, 3H).
4-((1H-Indol-1-yl)methyl)benzamide (22)
##STR00091##
Methyl 4-((1H-indol-1-yl)methyl)benzoate 1 was dissolved in MeOH
(10 mL) and to it was added a 30% solution of ammonium hydroxide (5
mL). The reaction was heated to reflux for 16 h after which it was
cooled to RT and H.sub.2O (30 mL) was added. The organic products
were extracted with EtOAc (3.times.15 mL), washed with brine (15
mL), dried with Na.sub.2SO.sub.4 and concentrated in vacuo. The
crude product was dissolved in DMF and purified by preparatory HPLC
method 1. The title compound was isolated as a white solid (37 mg,
13%). .sup.1H NMR (400 MHz, MeOD): .delta. 7.79 (d, J=7.9 Hz, 2H),
7.58 (d, J=7.8 Hz, 1H), 7.29 (m, 2H), 7.19 (d, J=8.0 Hz, 2H), 7.11
(t, J=7.2 Hz, 1H), 7.04 (t, J=7.5 Hz, 1H), 6.53 (d, J=2.8 Hz, 1H),
5.47 (s, 2H). .sup.13C NMR (100 MHz, MeOD): .delta. 170.59, 142.41,
136.24, 132.64, 128.93, 128.14, 127.57, 126.36, 121.13, 120.32,
118.99, 109.29, 101.14, 48.93. ESI-LRMS: [M+H].sup.+=251.1 m/z.
ESI-HRMS: calc. for C.sub.16H.sub.14N.sub.2O: [M+H].sup.+=251.1179
m/z, found: [M+H].sup.+=251.1179 m/z.
1-(4-((1H-Indol-1-yl)methyl)phenyl)ethanone (23)
##STR00092##
The title compound was prepared from 1H-indole (0.55 g, 4.69 mmol)
and 1-(4-(bromomethyl)phenyl)ethanone (1.00 g, 4.69 mmol) following
a procedure similar to General Procedure A. The title compound was
purified using automated column chromatography method 1 (0.410 g,
35%). .sup.1H NMR (400 MHz, CDCl.sub.3): .delta. 7.90 (d, J=8.2 Hz,
2H), 7.69 (d, J=7.5 Hz, 1H), 7.21 (m, 6H), 6.61 (d, J=3.1 Hz, 1H),
5.41 (s, 2H), 2.58 (s, 3H). .sup.13C NMR (100 MHz, MeOD): .delta.
198.58, 143.95, 136.22, 136.04, 128.94, 128.39, 128.17, 126.51,
121.19, 120.37, 119.06, 109.29, 101.24, 48.95, 25.23. ESI-HRMS:
calc. for C.sub.17H.sub.15NO: [M+H].sup.+=250.1226 m/z, found:
[M+H].sup.+=250.1232 m/z.
3-(1H-Indol-1-yl)propanamide (24)
##STR00093##
The title compound was prepared from 1H-indole (0.500 g, 4.27 mmol)
and 3-bromopropanamide (0.650 g, 4.27 mmol) following a procedure
similar to General Procedure A. The title compound was purified by
preparatory HPLC method 1 and isolated as a white solid (0.339 g,
42%). .sup.1H NMR (400 MHz, MeOD): .delta. 7.53 (d, J=7.9 Hz, 1H),
7.44 (d, J=8.3 Hz, 1H), 7.19 (d, J=2.9 Hz, 1H), 7.16 (t, J=7.5 Hz,
1H), 7.03 (t, J=7.4 Hz, 1H), 6.42 (d, J=2.8 Hz, 1H), 4.47 (t, J=6.8
Hz, 2H), 2.70 (t, J=6.8 Hz, 2H). .sup.13C NMR (100 MHz,
DMSO-d.sub.6): .delta. 172.4, 135.9, 129.0, 128.6, 121.4, 120.8,
119.4, 110.2, 101.0, 42.3, 36.2. ESI-LRMS: [M+H].sup.+=189.1 m/z.
ESI-HRMS: calc. for C.sub.11H.sub.12N.sub.2O.sub.1:
[M+H].sup.+=189.1022 m/z, found: [M+H].sup.+=189.1030 m/z.
N-Hydroxy-3-(1H-indol-1-yl)propanamide (25)
##STR00094##
NaH (0.444 g, 11.1 mmol) was dissolved in anhydrous DMF (5 mL)
under argon and cooled to 0.degree. C. To it was added 1H-indole
(1.00 g, 8.54 mmol) dissolved in anhydrous DMF (5 mL). The reaction
was stirred for 15 min at 0.degree. C. followed by the addition of
ethyl 3-bromopropanoate (1.10 mL, 8.54 mmol). The reaction was then
stirred for 3 h at 70.degree. C. and then quenched by the addition
of H.sub.2O (30 mL). The organic products were extracted with EtOAc
(3.times.30 mL), washed with H.sub.2O (2.times.30 mL), brine (15
mL), dried with Na.sub.2SO.sub.4, filtered and concentrated in
vacuo. The crude product was purified using automated column
chromatography method 1 to yield ethyl 3-(1H-indol-1-yl)propanoate
(0.759 g, 3.49 mmol). The title compound was then prepared from
ethyl 3-(1H-indol-1yl)propanoate (0.486 g, 5.52 mmol) according to
General Procedure B (prep. HPLC method 1) and isolated as a white
solid (0.104 g, 23%). .sup.1H NMR (400 MHz, MeOD): .delta. 7.53 (d,
J=7.83 Hz, 1H), 7.42 (d, J=8.1 Hz, 1H), 7.16 (m, 2H), 7.02 (t,
J=7.4 Hz, 1H), 6.41 (d, J=2.8 Hz, 1H), 4.48 (t, J=6.5 Hz, 2H), 2.56
(t, J=6.6 Hz, 2H). ESI-LRMS: [M+H].sup.+=205.1 m/z. ESI-HRMS: calc.
for C.sub.11H.sub.12N.sub.2O.sub.2: [M+H].sup.+=205.0972 m/z,
found: [M+H].sup.+=205.0971 m/z.
Methyl 4-((1H-benzo[d]imidazol-1-yl)methyl)benzoate (26)
##STR00095##
The title compound was prepared from 1H-benzo[d]imidazole (236 mg,
2.0 mmol) according to General Procedure C and purified using
automated column chromatography method 2. The product was isolated
as an off-white waxy solid (501 mg, 94%). .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 8.02-7.98 (m, 3H), 7.85 (d, J=7.6 Hz, 1H),
7.31-7.28 (m, 2H), 7.26-7.22 (m, 3H), 5.43 (s, 2H), 3.91 (s, 3H).
.sup.13C NMR (100 MHz, CDCl.sub.3) .delta. 166.41, 143.95, 143.13,
140.46, 133.72, 130.28, 130.14, 126.81, 123.26, 122.43, 120.54,
109.83, 52.19, 48.45 ESI-LRMS: [M+H].sup.+=267 m/z.
4-((1H-Benzo[d]imidazol-1-yl)methyl)-N-hydroxybenzamide.TFA
(27)
##STR00096##
The title compound was synthesized from methyl
4-((1H-benzo[d]imidazol-1-yl)methyl)benzoate 26 (93 mg, 0.349 mmol)
according to General Procedure D (prep. HPLC method 2) and isolated
as a white solid (81 mg, 87%). .sup.1H NMR (400 MHz, DMSO-d.sub.6)
.delta. 11.21 (br s, 1H), 9.35 (s, 1H), 7.83 (m, 1H), 7.74 (m, 3H),
7.47 (m, 4H). .sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 163.59,
142.78, 138.12, 135.10, 131.65, 127.80, 127.41, 125.03, 116.58,
112.53, 48.64. ESI-LRMS: [M+H].sup.+=268 m/z. ESI-HRMS: calc. for
C.sub.15H.sub.13N.sub.3O.sub.2: [M+H].sup.+=268.1081 m/z, found:
[M+H].sup.+=268.1080 m/z.
N-(2-(1H-Indol-3-yl)ethyl)methanesulfonamide (28)
##STR00097##
A round bottom flask charged with tryptamine (2.0 g, 12.5 mmol) in
DCM (20 mL) was added Et.sub.3N (3.5 mL, 25 mmol) under an
atmosphere of Ar and cooled to 0.degree. C. The solution was then
added mesyl chloride (1.5 mL, 18.7 mmol) and the resulting reaction
mixture was stir to RT for 2 h. The reaction was quenched with
water (20 mL) and extracted with DCM (3.times.20 mL). The combined
organic extracts were washed with brine (30 mL), dried over
Na.sub.2SO.sub.4 and concentrated in vacuo. The desired product was
purified via automated column chromatography method 2 (2.1 g, 70%)
and isolated as a brown oil. .sup.1H NMR (400 MHz, MeOD) .delta.
7.55 (d, J=7.6 Hz, 1H), 7.33 (d, J=8.0 Hz, 1H), 7.08 (m, 2H), 7.01
(td, J=8.0, 0.8 Hz, 1H), 3.35 (t, J=7.2 Hz, 2H), 2.99 (t, J=7.2 Hz,
2H), 2.76 (s, 3H). .sup.13C NMR (100 MHz, MeOD) .delta. 138.28,
128.77, 123.91, 122.54, 119.85, 119.34, 112.93, 112.43, 45.09,
40.02, 27.60.
Methyl
4-((3-(2-(methylsulfonamido)ethyl)-1H-indol-1-yl)methyl)benzoate
(29)
##STR00098##
The title compound was prepared from
N-(2-(1H-indol-3-yl)ethyl)methanesulfonamide 28 (300 mg, 1.26 mmol)
according to General Procedure A (substituting KO.sup.tBu for NaH)
and purified using automated column chromatography method 2. The
product was isolated as a brown waxy solid (0.399 g, 82%). .sup.1H
NMR (400 MHz, CDCl.sub.3) .delta. 8.20 (br s, 1 H), 8.01 (d, J=8.0
Hz, 2 H), 7.42 (m, 3 H), 7.33 (d, J=8.4 Hz, 1 H), 7.17 (t, J=7.6
Hz, 1 H), 7.08 (t, J=7.2 Hz, 1 H), 6.95 (d, J=1.6 Hz, 1 H), 4.45
(s, 2 H), 3.92 (s, 3 H), 3.49 (t, J=7.6 Hz, 2 H), 2.96 (t, J=7.6
Hz, 2 H), 2.74 (s, 3 H). .sup.13C NMR (100 MHz, CDCl.sub.3) .delta.
166.69, 141.53, 136.17, 129.95, 129.80, 126.96, 122.14, 122.13,
119.49, 118.40, 112.09, 111.27, 52.13, 51.23, 48.35, 38.90, 24.82.
ESI-LRMS: [M+H].sup.+=386 m/z.
N-Hydroxy-4-((3-(2-(methylsulfonamido)ethyl)-1H-indol-1-yl)methyl)benzamid-
e (30)
##STR00099##
The title compound was synthesized from methyl
4-((3-(2-(methylsulfonamido)ethyl)-1H-indol-1-yl)methyl)benzoate 29
(155 mg, 0.401 mmol) according to General Procedure D (prep. HPLC
method 2) and isolated as a beige solid (55 mg, 35%). .sup.1H NMR
(400 MHz, DMSO-d.sub.6) .delta. 11.22 (br s, 1H), 10.82 (s, 1H),
7.87 (d, J=8.4 Hz, 2H), 7.48 (d, J=8.4 Hz, 2H), 7.31 (d, J=8.0 Hz,
1H), 7.11 (d, J=2.4 Hz, 1H), 7.05 (td, J=7.6, 0.8 Hz, 1H), 6.94
(td, J=7.6, 0.8 Hz, 1H), 4.48 (s, 2H), 3.35 (m, 2H), 2.97 (s, 3H),
2.84 (m, 2H). .sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 164.00,
140.73, 136.15, 131.99, 128.01, 127.09, 126.84, 122.98, 121.00,
118.32, 118.07, 111.42, 110.70, 50.53, 48.42, 38.02, 24.48. LR-ESI
MS (m/z): 388 [M+H].sup.+. ESI-HRMS: calc. for
C.sub.19H.sub.21N.sub.3O.sub.4S: [M+H].sup.+=388.1326 m/z, found:
[M+H].sup.+=388.1319 m/z.
Methyl 4-((2-methyl-1H-benzo[d]imidazol-1-yl)methyl)benzoate
(31)
##STR00100##
The title compound was prepared from 2-methyl-1H-benzo[d]imidazole
(264 mg, 2.0 mmol) according to General Procedure C and purified
using automated column chromatography method 2. The product was
isolated as an off-white waxy solid (298 mg, 53%). .sup.1H NMR (400
MHz, CDCl.sub.3) .delta. 7.95 (d, J=8.4 Hz, 2H), 7.72 (d, J=8.0 Hz,
1H), 7.21 (m, 3H), 7.08 (d, J=8.4 Hz, 2H), 5.32 (s, 2H), 3.87 (s,
3H), 2.53 (s, 2H). .sup.13C NMR (100 MHz, CDCl.sub.3) .delta.
166.32, 151.60, 142.56, 140.77, 135.18, 130.18, 129.78, 126.04,
122.33, 122.08, 119.14, 109.06, 71.82, 52.06, 46.69, 13.81.
ESI-LRMS: [M+H].sup.+=281 m/z.
N-hydroxy-4-((2-methyl-1H-benzo[d]imidazol-1-yl)methyl)benzamide
(32)
##STR00101##
The title compound was synthesized from methyl
4-((2-methyl-1H-benzo[d]imidazol-1-yl)methyl)benzoate 31 (157 mg,
0.560) according to General Procedure D (prep. HPLC method 2) and
isolated as an off-white solid (55 mg, 35%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 11.27 (br s, 1H), 7.74 (m, 3H), 7.69 (d,
J=8.4 Hz, 1H), 7.44 (m, 2H), 7.34 (d, J=8.4 Hz, 2H), 5.72 (s, 2H),
2.80 (s, 3H). .sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 164.05,
152.38, 138.48, 133.78, 133.10, 132.92, 127.90, 127.55, 125.24,
125.01, 115.64, 112.54, 47.33, 12.72. LR-ESI MS (m/z): 282
[M+H].sup.+. ESI-HRMS: calc. for C.sub.16H.sub.15N.sub.3O.sub.2:
[M+H].sup.+=282.1237 m/z, found: [M+H].sup.+=282.1244 m/z.
tert-Butyl (3-(1H-benzo[d]imidazol-2-yl)propyl)carbamate (33)
##STR00102##
To a round bottom flask charged with
4-((tert-butoxycarbonyl)amino)butanoic acid (0.5 g, 2.46 mmol) in
DCM (10 mL) was added Et.sub.3N (1.03 mL, 7.36 mmol), EDCI (706 mg,
2.46 mmol), DMAP (30 mg, 0.246 mmol) and allowed to stir overnight
at RT. The reaction was quenched with water (10 mL) and extracted
with DCM (2.times.10 mL). Combined organics washed with 10% citric
acid (10 mL), brine (10 mL), dried over Na.sub.2SO.sub.4 and
concentrated in vacuo. The crude material was dissolved in AcOH (7
mL) and stirred at 65.degree. C. for 1 h. The mixture was diluted
with saturated bicarbonate (15 mL) and extracted with DCM
(2.times.15 mL), and again worked up in the above manner. The title
compound was obtained as a white solid (433 mg, 64%) and used
without further purification. .sup.1H NMR (400 MHz, CDCl.sub.3)
.delta. 7.57 (m, 2H), 7.20 (m, 2H), (4.98 (s, 2H), 3.24 (m, 2H),
2.95 (m, 2H), 1.92 (m, 2H), 1.47 (s, 9H). ESI-LRMS: [M+H].sup.+=276
m/z.
Methyl
4-((2-(3-aminopropyl)-1H-benzo[d]imidazol-1-yl)methyl)benzoate
(34)
##STR00103##
The title compound was prepared from tert-Butyl
(3-(1H-benzo[d]imidazol-2-yl)propyl)carbamate 33 (433 mg, 1.57
mmol) using General Procedure C. The crude material was taken up
into acetone (7 mL), added conc. HCl (3 equiv) and allowed to stir
at 50.degree. C. for 4 h. The reaction was quenched with saturated
bicarbonate and worked up in the usual manner, affording the title
compound as a viscous brown oil (458 (mg, 90%) used without further
purification. .sup.1H NMR (400 MHz, MeOD) .delta. 7.95 (m, 3H),
7.63 (d, J=7.6 Hz, 1H), 7.44 (d, J=8.0 Hz, 1H), 7.33 (m, 1H), 7.23
(m, 2H), 7.18 (m, 2H), 5.58 (s, 2H), 3.86 (m, 3H), 2.93 (t, J=7.6
Hz, 2H), 2.06 (t, J=7.6 Hz, 2H), 1.98 (m, 2H). ESI-LRMS:
[M+H].sup.+=324 m/z.
Methyl
4-((2-(3-acetamidopropyl)-1H-benzo[d]imidazol-1-yl)methyl)benzoate
(35)
##STR00104##
To a round bottom flask charged with methyl
4-((2-(3-aminopropyl)-1H-benzo[d]imidazol-1-yl)methyl)benzoate 34
(135 mg, 0.417 mmol) in DCM (5 mL) was added Et.sub.3N (0.076 mL,
0.543 mmol) and catalytic DMAP (5 mg, 0.042 mmol). The reaction was
cooled to 0.degree. C. and then Ac.sub.2O (0.051 mL, 0.543 mmol)
was added dropwise. The resulting solution was allowed to warm to
RT and stirred for 18 h after which the reaction was quenched with
H.sub.2O (10 mL) and extracted with chloroform (3.times.10 mL).
Combined extracts were washed with brine (20 mL), dried over
Na.sub.2SO.sub.4 and concentrated in vacuo. The crude product was
purified using automated column chromatography method 2 and
isolated as a white, waxy solid (53 mg, 35%). .sup.1H NMR (400 MHz,
CDCl3) .delta. 7.96 (d, J=8.0 Hz, 2H), 7.73 (d, J=7.6 Hz, 1H), 7.24
(m, 3H), 7.07 (d, J=8.0 Hz, 2H), 6.73 (br s, 1H), 5.38 (s, 2H),
3.88 (s, 3H), 3.33 (m, 2H), 2.89 (t, J=7.2H, 2H), 2.05 (m, 2H),
1.86 (s, 3H). .sup.13C NMR (100 MHz, CDCl.sub.3) .delta. 170.44,
166.32, 154.48, 141.76, 140.60, 134.98, 130.26, 129.91,
126.00122.81, 122.49, 118.89, 109.42, 52.12, 46.60, 38.91, 26.42,
24.92, 22.97. ESI-LRMS: [M+H].sup.+=366 m/z.
4-((2-(3-Acetamidopropyl)-1H-benzo[d]imidazol-1-yl)methyl)-N-hydroxybenzam-
ide.TFA (36)
##STR00105##
The title compound was synthesized from methyl
4-((2-(3-acetamidopropyl)-1H-benzo[d]imidazol-1-yl)methyl)benzoate
35 (53 mg, 0.145 mmol) according to General Procedure D (prep. HPLC
method 2) and isolated as a white solid (36 mg, 52%). .sup.1H NMR
(400 MHz, DMSO-d.sub.6) .delta. 8.01 (m, 1H), 7.78 (m, 4H), 7.52
(m, 2H), 7.33 (d, J=8.0 Hz, 2H), 5.78 (s, 2H), 3.18 (m, 4H), 1.92
(m, 2H), 1.78 (s, 3H). .sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta.
169.12, 163.16, 157.65, 154.11, 137.32, 132.19, 131.80, 127.09,
126.62, 125.30, 124.98, 114.39, 112.34, 46.74, 37.29, 26.03, 22.62,
22.19. ESI-LRMS: [M+H].sup.+=367 m/z. ESI-HRMS: calc. for
C.sub.20H.sub.22N.sub.4O.sub.3: [M+H].sup.+=367.1765 m/z, found:
[M+H].sup.+=367.1757 m/z.
Methyl
4-((2-(2-(dimethylamino)ethyl)-1H-benzo[d]imidazol-1-yl)methyl)benz-
oate (37)
##STR00106##
tert-Butyl 2-(1H-benzo[d]imidazol-2-yl)ethylcarbamate (390 mg, 1.70
mmol) was subjected to general procedure C to produce the ester
intermediate methyl
4-((2-(2-(tert-butoxycarbonylamino)ethyl)-1H-benzo[d]imidazol-1-yl-
)methyl)benzoate. The crude material was dissolved in acetone (2
mL/mmol ester), conc. HCl (3 mol equiv) was added and then the
reaction was stirred for 16 h. The precipitate was filtered, washed
with acetone, dried, and used without further purification. The
dihydrochloride intermediate (131 mg, 0.343 mmol) was dissolved in
MeOH (3 mL) and cooled to 0.degree. C. AcOH (0.100 .mu.M) was added
followed by NaCNBH.sub.3 (43 mg, 0.686 mmol) under an atmosphere of
Ar. Lastly, a solution of CH.sub.2O (0.1 mL, 37% wt soln) in MeOH
(1 mL) was added dropwise. The resulting reaction mixture was
allowed to stir 4.5 h to RT. The reaction was quenched with 1N HCl
(10 mL) and extracted with EtOAc (3.times.10 mL). The aqueous layer
was made basic with 2 N NaOH and again extracted with EtOAc
(3.times.10 mL). The basic extraction was washed with brine (20
mL), dried over Na.sub.2SO.sub.4 and concentrated in vacuo.
Material was purified via automated column chromatography method 2
to yield a white waxy solid (88 mg, 76%). .sup.1H NMR (400 MHz,
CDCl.sub.3) .delta. 7.98 (d, J=8.4 Hz, 2H), 7.76 (J=8.0 Hz, 1H),
7.23 (m, 3H), 7.11 (d, J=8.4 Hz, 2H), 5.42 (s, 2H), 3.89 (s, 3H),
3.00 (t, J=7.6 Hz, 2H), 2.83 (t, J=7.6 Hz, 2H), 2.26 (s, 6H).
.sup.13C NMR (100 MHz, CDCl.sub.3) .delta. 166.44, 153.45, 142.61,
140.99, 135.17, 130.26, 129.86, 126.10, 122.58, 122.25, 119.42,
109.27, 57.13, 52.15, 46.68, 45.29, 26.07. ESI-LRMS:
[M+H].sup.+=338 m/z, [M+Na].sup.+=360 m/z.
4-((2-(2-(Dimethylamino)ethyl)-1H-benzo[d]imidazol-1-yl)methyl)-N-hydroxyb-
enzamide (38)
##STR00107##
The title compound was synthesized from methyl
4-(2-(2-(dimethylamino)ethyl)-1H-benzo[d]imidazol-1-yl)methyl)benzoate
37 (88 mg, 0.261 mmol) according to General Procedure D (prep. HPLC
method 2) and isolated as a white solid (53 mg, 60%). .sup.1H NMR
(400 MHz, DMSO-d.sub.6) .delta. 7.68 (d, J=8.4 Hz, 2H), 7.58 (m,
1H), 7.42 (m, 1H), 7.16 (m, 4H), 5.55 (s, 2H), 2.96 (t, J=7.6 Hz,
2H), 2.64 (t, J=7.6 Hz, 2H), 2.13 (s, 6H). .sup.13C NMR (100 MHz,
DMSO-d.sub.6) .delta. 163.65, 153.91, 142.34, 139.87, 135.25,
132.50, 127.17, 126.34, 121.77, 121.42, 118.47, 110.10, 56.75,
45.80, 44.98, 25.15. ESI-LRMS: [M+H].sup.+=339 m/z. ESI-HRMS: calc.
for C.sub.19H.sub.22N.sub.4O.sub.2: [M+H].sup.+=339.1816 m/z,
found: [M+H].sup.+=339.1811 m/z.
N-Hydroxy-4-((2-isopropyl-1H-benzo[d]imidazol-1-yl)methyl)benzamide
(39)
##STR00108##
The title compound was synthesized by subjecting
2-isopropyl-1H-benzo[d]imidazole (209 mg, 1.30 mmol) to General
Procedure C (automated column chromatography method 2) followed by
General Procedure D (prep. HPLC method 2). The desired product was
isolated as a white solid (23 mg, 25%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6) .delta. 11.95 (s, 1H), 9.01 (s, 1H), 7.68 (d, J=8.4
Hz, 2H), 7.60 (m, 1H), 7.40 (m, 1H), 7.16 (m, 2H), 7.10 (d, J=8.4
Hz, 2H), 5.57 (s, 2H), 3.25 (quint, J=6.8 Hz, 1H), 1.26 (s, 3H),
1.24 (s, 3H). .sup.13C NMR (100 MHz, DMSO-d.sub.6) .delta. 163.84,
159.85, 142.26, 140.43, 135.12, 131.98, 127.32, 126.20, 121.77,
121.44, 118.63, 110.20, 45.54, 25.71, 21.75 ESI-LRMS:
[M+H].sup.+=310 m/z. ESI-HRMS: calc. for
C.sub.18H.sub.19N.sub.3O.sub.2: [M+H].sup.+=310.1550 m/z, found:
[M+H].sup.+=310.1563 m/z.
Two examples of present HDACIs having a monocyclic Cap group are
prepared as follows (see also General Synthetic Schemes B and
C):
##STR00109##
##STR00110##
Examples of monocyclic Cap groups include, but are not limited
to:
##STR00111##
HDACIs having a five-membered or six-membered monocylic Cap
group:
Methyl 4-((1H-pyrrol-1-yl)methyl)benzoate (40)
##STR00112##
The title compound was prepared from 1H-pyrrole (0.150 g, 2.24
mmol) according to General Procedure A (substituting KO.sup.tBu for
NaH) and purified using automated column chromatography method 1.
The product was isolated as a clear viscous oil (0.432 g, 90%).
.sup.1H NMR (400 MHz, CDCl.sub.3): .delta. 8.01 (d, J=8.3 Hz, 2H),
7.17 (d, J=8.3 Hz, 2H), 6.72 (t, J=2.0 Hz, 2H), 6.24 (t, J=2.1 Hz,
2H), 5.15 (s, 2H), 3.93 (s, 3H). .sup.13C NMR (100 MHz,
CDCl.sub.3): .delta. 166.7, 143.4, 130.0, 129.5, 126.7, 121.2,
108.9, 53.0, 52.1.
4-((1H-Pyrrol-1-yl)methyl)-N-hydroxybenzamide (41)
##STR00113##
The title compound was synthesized from methyl
4-((1H-pyrrol-1-yl)methyl)benzoate 40 (0.400 g, 1.86 mmol)
according to General Procedure B (prep. HPLC method 1) and isolated
as a white solid (0.225 g, 56%). .sup.1H NMR (400 MHz, MeOD):
.delta. 7.70 (d, J=8.3 Hz, 2H), 7.20 (d, J=8.3 Hz, 2H), 6.73 (t,
J=2.0 Hz, 2H), 6.11 (t, J=2.1 Hz, 2H), 5.17 (s, 2H). .sup.13C NMR
(100 MHz, MeOD): .delta. 166.4, 143.0, 131.2, 127.0, 126.6, 120.7,
108.0, 52.0. ESI-HRMS: calc. for C.sub.12H.sub.12N.sub.2O.sub.2:
[M+H].sup.+=217.0972 m/z, found: [M+H].sup.+=217.0975 m/z.
Methyl 4-((1H-pyrazol-1-yl)methyl)benzoate (42)
##STR00114##
The title compound was prepared from 1H-pyrazole (0.150 g, 2.20
mmol) according to General Procedure A (substituting KO.sup.tBu for
NaH) and purified using automated column chromatography method 1.
The product was isolated as a viscous yellow oil (0.387 g, 81%).
.sup.1H NMR (400 MHz, CDCl.sub.3): .delta. 7.99 (d, J=8.2 Hz, 2H),
7.56 (s, 1H), 7.41 (d, J=2.0 Hz, 1H), 7.20 (d, J=8.1 Hz, 2H), 6.30
(d, J=1.9 Hz, 1H), 5.36 (s, 2H), 3.88 (s, 3H). .sup.13C NMR (100
MHz, CDCl.sub.3): .delta. 166.2, 141.5, 139.5, 129.7, 129.4, 129.1,
126.9, 105.9, 55.0, 51.7. ESI-HRMS: calc. for
C.sub.12H.sub.12N.sub.2O.sub.2: [M+H].sup.+=217.0972 m/z, found:
[M+H].sup.+=217.0969 m/z.
4-((1H-Pyrazol-1-yl)methyl)-N-hydroxybenzamide (43)
##STR00115##
The title compound was synthesized from methyl
44(1H-pyrazol-1-yl)methyl)benzoate 42 (0.387 g, 1.79 mmol)
according to General Procedure B (prep. HPLC method 1) and isolated
as a white solid (0.251 g, 65%). .sup.1H NMR (400 MHz, MeOD):
.delta. 7.74 (m, 3H), 7.55 (d, J=1.5 Hz, 1H), 7.27 (d, J=8.4 Hz,
2H), 6.37 (t, J=2.1 Hz, 1H), 5.43 (s, 2H). .sup.13C NMR (100 MHz,
MeOD): .delta. 166.3, 140.9, 139.3, 131.7, 130.5, 127.1, 127.0,
105.7, 54.3. ESI-HRMS: calc. for C.sub.11H.sub.11N.sub.3O.sub.2:
[M+H].sup.+=218.0924 m/z, found: [M+H].sup.+=218.0917 m/z.
Methyl 4-(pyridin-4-ylmethyl)benzoate (44)
##STR00116##
The title compound was prepared from pyridin-4-ylboronic acid
(0.123 g, 1.00 mmol) according to General Procedure E and purified
using automated column chromatography method 2 (20 mg, 9%). .sup.1H
NMR (400 MHz, MeOD): .delta. 8.59 (d, J=6.6 Hz, 2H), 7.86 (d, J=8.3
Hz, 2H), 7.78 (d, J=6.6 Hz, 2H), 7.28 (d, J=8.3 Hz, 2H), 4.26 (s,
2H), 3.74 (s, 3H). .sup.13C NMR (100 MHz, CDCl.sub.3): .delta.
166.7, 162.3, 142.4, 141.2, 129.9, 129.3, 129.1, 127.2, 51.3, 40.8.
ESI-LRMS: [M+H].sup.+=228 m/z.
N-Hydroxy-4-(pyridin-4-ylmethyl)benzamide (45)
##STR00117##
The title compound was synthesized from methyl
4-(pyridin-4-ylmethyl)benzoate 44 (0.020 g, 0.09 mmol) according to
General Procedure B (prep. HPLC method 2) and isolated as a white
solid (5 mg, 25%). .sup.1H NMR (400 MHz, MeOD): .delta. 8.74 (d,
J=6.4 Hz, 2H), 7.93 (d, J=6.3 Hz, 2H), 7.78 (d, J=8.2 Hz, 2H), 7.43
(d, J=8.1 Hz, 2H), 4.41 (s, 2H). .sup.13C NMR (100 MHz, MeOD):
.delta. 166.1, 162.4, 141.4, 140.8, 131.3, 129.3, 127.5, 127.1,
40.7. ESI-HRMS: calc. for C.sub.13H.sub.12N.sub.2O.sub.2:
[M+H].sup.+=229.0972 m/z, found: [M+H].sup.+=229.0966 m/z.
Methyl 4-(4-(dimethylamino)benzyl)benzoate (46)
##STR00118##
The title compound was prepared from 4-(dimethylamino)phenylboronic
acid (0.165 g, 1.00 mmol) according to General Procedure E and
purified using automated column chromatography method 2. The
product was isolated as an orange oil (0.211 g, 78%). .sup.1H NMR
(400 MHz, MeOD): .delta. 7.93 (d, J=8.4 Hz, 2H), 7.59 (d, J=8.7 Hz,
2H), 7.45 (d, J=8.7 Hz, 2H), 7.33 (d, J=8.4 Hz, 2H), 4.10 (s, 2H),
3.87 (s, 3H), 3.28 (s, 6H). .sup.13C NMR (100 MHz, CDCl.sub.3):
.delta. 166.9, 146.0, 142.8, 141.3, 130.6, 129.5, 128.7, 128.1,
120.3, 51.2, 45.6, 40.5. ESI-LRMS: [M+H].sup.+=270 m/z.
4-(4-(Dimethylamino)benzyl)-N-hydroxybenzamide (47)
##STR00119##
The title compound was synthesized from methyl
4-(4-(dimethylamino)benzyl)benzoate 46 (0.211 g, 0.78 mmol)
according to General Procedure B (prep. HPLC method 2) and isolated
as a white solid (0.140 g, 66%). .sup.1H NMR (400 MHz, MeOD):
.delta. 7.69 (d, J=8.2 Hz, 2H), 7.53 (d, J=8.6 Hz, 2H), 7.42 (d,
J=8.56, 2H), 7.31 (d, J=8.1 Hz, 2H), 4.09 (s, 2H), 3.25 (s, 6H).
.sup.13C NMR (100 MHz, MeOD): .delta. 166.6, 144.5, 142.3, 141.7,
130.5, 130.2, 128.8, 127.1, 119.8, 45.3, 40.4. ESI-HRMS: calc. for
C.sub.16H.sub.18N.sub.2O.sub.2: [M+H].sup.+=271.1441 m/z, found:
[M+H].sup.+=271.1448 m/z.
Methyl 4-(3-(dimethylamino)benzyl)benzoate (48)
##STR00120##
The title compound was prepared from 3-(dimethylamino)phenylboronic
acid (0.165 g, 1.00 mmol) according to General Procedure E and
purified using automated column chromatography method 2. The
product was isolated as a clear oil (0.254 g, 94%). .sup.1H NMR
(400 MHz, CDCl.sub.3): .delta. 7.99 (d, J=8.1 Hz, 2H), 7.30 (m,
6H), 4.08 (s, 2H), 3.91 (s, 3H), 3.15 (s, 6H). .sup.13C NMR (100
MHz, CDCl.sub.3): .delta. 166.9, 144.9, 144.5, 143.1, 130.6, 130.0,
128.9, 128.8, 128.6, 119.9, 117.5, 52.1, 45.6, 41.6. ESI-LRMS:
[M+H].sup.+=270 m/z.
4-(3-(Dimethylamino)benzyl)-N-hydroxybenzamide (49)
##STR00121##
The title compound was synthesized from methyl
4-(3-(dimethylamino)benzyl)benzoate 48 (0.254 g, 0.94 mmol)
according to General Procedure B (prep. HPLC method 2) and isolated
as a white solid (0.117 g, 46%). .sup.1H NMR (400 MHZ, CDCl.sub.3):
.delta. 7.70 (d, J=8.2 Hz, 2H), 7.50 (m, 3H), 7.35 (m, 3H), 4.12
(s, 2H), 3.27 (s, 6H). .sup.13C NMR (100 MHZ, CDCl.sub.3): .delta.
166.6, 144.3, 143.7, 143.6, 130.4, 130.2, 129.7, 128.8, 127.1,
120.2, 117.5, 45.4, 40.7. ESI-HRMS: calc. for
C.sub.16H.sub.18N.sub.2O.sub.2: [M+H].sup.+=271.1450 m/z, found:
[M+H].sup.+=271.1450 m/z.
An example of a present HDACI having an acyclic Cap group is
prepared as follows (see also General Synthetic Scheme D):
##STR00122##
Examples of acyclic Cap groups include, but are not limited to:
##STR00123## HDACI compounds having an acyclic Cap group:
Methyl 4-((diethylamino)methyl)benzoate (50)
##STR00124##
The title compound was prepared from diethylamine (0.350 g, 4.79
mmol) according to General Procedure A (substituting KO.sup.tBu for
NaH). The pH was adjusted to 10 with 2 N NaOH prior to extraction
with EtOAc. The product was isolated as a yellow oil and did not
require further purification (0.790 g, 75%). .sup.1H NMR (400 MHz,
MeOD): .delta. 8.14 (d, J=8.2 Hz, 2H), 7.66 (d, J=8.2 Hz, 2H), 4.44
(s, 2H), 3.95 (s, 3H), 3.25 (m, 4H), 1.37 (t, J=7.3 Hz, 6H).
.sup.13C NMR (100 MHz, DMSO-d.sub.6): .delta. 166.2, 135.9, 131.8,
130.9, 130.0, 54.8, 52.8, 46.6, 8.8. ESI-HRMS: calc. for
C.sub.13H.sub.19NO.sub.2: [M+H].sup.+=222.1489 m/z, found:
[M+H].sup.+=222.1485 m/z.
4-((Diethylamino)methyl)-N-hydroxybenzamide (51)
##STR00125##
The title compound was synthesized from methyl
4-((diethylamino)methyl)benzoate 50 (0.518 g, 2.34 mmol) according
to General Procedure B (prep. HPLC method 2) and isolated as a
viscous, yellow oil (0.448 g, 56%). .sup.1H NMR (400 MHz, MeOD):
.delta. 7.72 (d, J=8.2 Hz, 2H), 7.42 (d, J=8.1 Hz, 2H), 3.65 (s,
2H), 2.55 (q, J=7.2 Hz, 4H), 1.08 (t, J=5.0 Hz, 6H). .sup.13C NMR
(100 MHz, MeOD): .delta. 166.3, 139.5, 131.9, 129.7, 127.0, 56.1,
46.4, 9.3. ESI-HRMS: calc. for C.sub.12H.sub.18N.sub.2O.sub.2:
[M+H].sup.+=223.1441 m/z, found: [M+H].sup.+=223.1441 m/z.
Methyl 4-((diisopropylamino)methyl)benzoate (52)
##STR00126##
The title compound was prepared from diisopropylamine (0.500 g,
4.94 mmol) according to General Procedure A (substituting
KO.sup.tBu for NaH). The pH was adjusted to 10 with 2 N NaOH prior
to extraction with EtOAc. The product was isolated as a yellow oil
and did not require further purification (1.10 g, 89%). .sup.1H NMR
(400 MHz, MeOD): .delta. 8.10 (d, J=8.4 Hz, 2H), 7.71 (d, J=8.4 Hz,
2H), 4.51 (s, 2H), 3.93 (s, 3H), 3.85 (m, 2H), 1.46 (dd, J=6.1 Hz,
J=12.5 Hz, 12H). .sup.13C NMR (100 MHz, MeOD): .delta. 165.9,
135.9, 130.6, 130.1, 129.4, 54.8, 51.1, 49.4, 17.2, 16.3. ESI-HRMS:
calc. for C.sub.15H.sub.23NO.sub.2: [M+H].sup.+=250.1802 m/z,
found: [M+H].sup.+=250.1800 m/z.
4-((Diisopropylamino)methyl)-N-hydroxybenzamide (53)
##STR00127##
The title compound was synthesized from methyl
4-((diisopropylamino)methyl)benzoate 52 (0.750 g, 3.0 mmol)
according to General Procedure B (prep. HPLC method 2) and isolated
as a viscous, light-yellow oil (0.324 g, 42%). .sup.1H NMR (400
MHz, MeOD): .delta. 7.84 (d, J=8.2 Hz, 2H), 7.67 (d, J=8.2 Hz, 2H),
4.48 (s, 2H), 3.84 (m, 2H), 1.47 (br, 12H). .sup.13C NMR (100 MHz,
MeOD): .delta. 165.6, 134.7, 133.2, 130.5, 127.5, 55.1, 49.7, 17.5,
16.8. ESI-HRMS: calc. for C.sub.14H.sub.22N.sub.2O.sub.2:
[M+H].sup.+=251.1754 m/z, found: [M+H].sup.+=251.1744 m/z.
Methyl 4-((diphenylamino)methyl)benzoate (54)
##STR00128##
NaH (0.220 g, 5.54 mmol) was dissolved in anhydrous DMF (2 mL)
under argon and cooled to 0.degree. C. To it was added
diphenylamine (0.0.750 g, 4.43 mmol) dissolved in anhydrous DMF (2
mL). The reaction was stirred for 15 min at 0.degree. C. followed
by the addition of methyl 4-(bromomethyl)benzoate (1.02 g, 4.43
mmol) in anhydrous DMF (2 mL). The reaction was stirred for 2 h at
60.degree. C. and then quenched by the addition of H.sub.2O (20
mL). The organic products were extracted with EtOAc (3.times.30
mL), washed with H.sub.2O (2.times.30 mL), brine (15 mL), dried
with Na.sub.2SO.sub.4, filtered and concentrated in vacuo. The
title compound was purified using automated column chromatography
method 1 (0.746 g, 53%). .sup.1H NMR (400 MHz, CDCl.sub.3): .delta.
8.00 (d, J=8.2 Hz, 2H), 7.45 (d, J=7.7 Hz, 2H), 7.27 (m, 4H), 7.07
(d, J=7.9 Hz, 4H), 6.98 (t, J=7.3 Hz, 2H), 5.06 (s, 2H), 3.92 (s,
3H). .sup.13C NMR (100 MHz, CDCl.sub.3): .delta. 166.9, 147.8,
144.8, 130.0, 129.4, 128.9, 126.5, 121.7, 120.7, 56.3, 52.0.
ESI-LRMS: [M+H].sup.+=318 m/z.
4-((Diphenylamino)methyl)-N-hydroxybenzamide (55)
##STR00129##
The title compound was synthesized from methyl
4-((diphenylamino)methyl)benzoate 54 (0.200 g, 0.33 mmol) according
to General Procedure B (prep. HPLC method 2) and isolated as a
white solid (27 mg, 13%). .sup.1H NMR (400 MHz, MeOD): .delta. 7.69
(d, J=8.0 Hz, 2H), 7.47 (d, J=8.0 Hz, 2H), 7.23 (t, J=7.6 Hz, 4H),
7.05 (d, J=8.0 Hz, 4H), 6.93 (t, J=7.5 Hz, 2H), 5.07 (s, 2H).
.sup.13C NMR (100 MHz, DMSO-d.sub.6): .delta. 164.5, 147.8, 143.0,
131.8, 129.8, 127.5, 126.9, 121.7, 120.7, 55.5. ESI-HRMS: calc. for
C.sub.20H.sub.18N.sub.2O.sub.2: [M+H].sup.+=319.1441 m/z, found:
[M+H].sup.+=319.1447 m/z.
Methyl 4-(pyrrolidin-1-ylmethyl)benzoate (56)
##STR00130##
Pyrrolidine (0.12 mL, 1.52 mmol), methyl 4-formylbenzoate (0.250 g,
1.52 mmol), NaBH(OAc).sub.3 (0.52 g, 2.4 mmol) and 5 .ANG.
molecular sieves were dissolved in 1,2-dichloroethane (5 mL) under
Ar atmosphere and stirred for 24 h at RT. Then, the reaction was
diluted with 2 N NaOH (30 mL) and extracted with EtOAc (3.times.20
mL). The combined organic fractions were washed with brine (15 mL),
dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo.
The crude product was purified via automated column chromatography
(50-100% EtOAc/hexane, 25 g cartridge) to yield the title compound
as a clear oil (0.166 g, 50%). .sup.1H NMR (400 MHz, CDCl.sub.3):
.delta. 8.00 (d, J=8.0 Hz, 2H), 7.43 (d, J=8.0 Hz, 2H), 3.92 (s,
3H), 3.69 (s, 2H), 2.54 (br, 4H), 1.81 (t, J=3.2 Hz, 4H).
N-Hydroxy-4-(pyrrolidin-1-ylmethyl)benzamide.TFA (57)
##STR00131##
The title compound was synthesized from methyl
4-(pyrrolidin-1-ylmethyl)benzoate 56 (0.120 g, 0.55 mmol) according
to General Procedure B (prep. HPLC method 2) and isolated as the
trifluoroacetic acid salt (38 mg, 21%). .sup.1H NMR (400 MHz,
DMSO-d.sub.6): .delta. 11.33 (s, 1H), 10.69 (br, 1H), 9.15 (br,
1H), 7.81 (d, J=8.0 Hz, 2H), 7.58 (d, J=8.0 Hz, 2H), 4.39 (s, 2H),
3.22 (br, 4H), 1.94 (br, 4H). .sup.13C NMR (100 MHz, MeOD): .delta.
165.61, 134.17, 133.62, 130.29, 127.62, 57.21, 53.60, 22.37.
ESI-HRMS: calc. for C.sub.12H.sub.16N.sub.2O.sub.2:
[M+H].sup.+=221.1285 m/z, found: [M+H].sup.+=221.1286 m/z.
(S)-Methyl 4-((2-(methoxymethyl)pyrrolidin-1-yl)methyl)benzoate
(58)
##STR00132##
The title compound was synthesized from
(S)-2-(methoxymethyl)pyrrolidine (0.175 g, 1.52 mmol) according to
a procedure similar to that used for compound 56. .sup.1H NMR (400
MHz, CDCl.sub.3): .delta. 7.99 (d, J=8.4 Hz, 2H), 7.42 (d, J=8.0
Hz, 2H), 4.17 (d, J=13.6 Hz, 1H), 3.92 (s, 3H), 3.45 (m, 2H), 3.35
(m, 4H), 2.92 (m, 1H), 2.74 (m, 1H), 2.21 (m, 1H), 1.95 (m, 1H),
1.69 (m, 3H).
(S)-N-Hydroxy-4-((2-(methoxymethyl)pyrrolidin-1-yl)methyl)benzamide.TFA
(59)
##STR00133##
The title compound was synthesized from (S)-methyl
4-((2-(methoxymethyl)pyrrolidin-1-yl)methyl)benzoate 58 (0.120 g,
0.46 mmol) according to General Procedure B (prep. HPLC method 2)
and isolated as the trifluoroacetic acid salt (23 mg, 13%). .sup.1H
NMR (400 MHz, MeOD): .delta. 7.87 (d, J=8.0 Hz, 2H), 7.64 (d, J=8.0
Hz, 2H), 4.67 (d, J=12.8 Hz, 1H), 4.35 (d, J=12.8 Hz, 1H), 3.85
(br, 1 H), 3.60 (d, J=4.8 Hz, 2H), 3.42 (br, 4H), 3.32 (m, 1H),
2.30 (m, 1H), 2.15 (m, 1H), 1.95 (m, 2H). .sup.13C NMR (100 MHz,
MeOD): .delta. 165.55, 133.74, 130.81, 127.55, 69.86, 67.00, 58.08,
57.75, 54.48, 26.03, 21.81. ESI-HRMS: calc. for
C.sub.14H.sub.20N.sub.2O.sub.3: [M+H].sup.+=265.1547 m/z, found:
[M+H].sup.+=265.1550 m/z.
(R)-Methyl 4-((2-(methoxymethyl)pyrrolidin-1-yl)methyl)benzoate
(60)
##STR00134##
The title compound was synthesized from
(R)-2-(methoxymethyl)pyrrolidine (0.175 g, 1.52 mmol) according to
a procedure similar to that used for compound 56. .sup.1H NMR (400
MHz, CDCl.sub.3): .delta. 7.98 (d, J=8.0 Hz, 2H), 7.41 (d, J=8.0
Hz, 2H), 4.16 (d, J=13.6 Hz, 1H), 3.91 (s, 3H), 3.42 (m, 2H), 3.33
(m, 4H), 2.92 (m, 1H), 2.73 (m, 1H), 2.19 (m, 1H), 1.93 (m, 1H),
1.70 (m, 3H). .sup.13C NMR (100 MHz, CDCl.sub.3): .delta. 166.7,
144.7, 129.1, 128.4, 128.3, 76.0, 62.8, 58.9, 58.7, 54.2, 51.6,
28.0, 22.4. ESI-LRMS: [M+H].sup.+=264 m/z.
(R)--N-Hydroxy-4-(2-(methoxymethyl)pyrrolidin-1-yl)methyl)benzamide.TFA
(61)
##STR00135##
The title compound was synthesized from (R)-methyl
4-((2-(methoxymethyl)pyrrolidin-1-yl)methyl)benzoate 60 (0.120 g,
0.46 mmol) according to General Procedure B (prep. HPLC method 2)
and isolated as the trifluoroacetic acid salt (21 mg, 12%). .sup.1H
NMR (400 MHz, DMSO-d.sub.6): .delta. 11.23 (s, 1H), 9.87 (br, 1H),
9.02 (br, 1H), 7.71 (d, J=8.4 Hz, 2H), 7.49 (d, J=8.0 Hz, 2H), 4.45
(d, J=12.8 Hz, 1H), 4.20 (m, 1H), 3.62 (br, 1H), 3.47 (m, 1H), 3.39
(m, 1H), 3.17 (s, 4H), 3.08 (br, 1H), 2.05 (m, 1H), 1.87 (m, 1H),
1.736 (m, 1H), 1.63 (m, 1H). .sup.13C NMR (100 MHz, DMSO-d.sub.6):
.delta. 163.85, 134.05, 131.35, 127.68, 70.70, 66.37, 58.91, 57.36,
54.37, 26.59, 22.16. ESI-HRMS: calc. for
C.sub.14H.sub.20N.sub.2O.sub.3: [M+H].sup.+=265.1547 m/z, found:
[M+H].sup.+=265.1551 m/z.
The effectiveness, or potency, of a present HDACI with respect to
inhibiting the activity of an HDAC is measured by an IC.sub.50
value. The quantitative IC.sub.50 value indicates the concentration
of a particular compound that is needed to inhibit the activity of
an enzyme by 50% in vitro. Stated alternatively, the IC.sub.50
value is the half maximal (50%) inhibitory concentration of a
compound tested using a specific enzyme, e.g., HDAC, of interest.
The smaller the IC.sub.50 value, the more potent the inhibiting
action of the compound because a lower concentration of the
compound is needed to inhibit enzyme activity by 50%.
In preferred embodiments, a present HDACI inhibits HDAC enzymatic
activity by about at least 50%, preferably at least about 75%, at
least 90%, at least 95%, or at least 99%.
Compounds of the present invention were tested for IC.sub.50 values
against both HDAC6 and HDAC1. In some embodiments, a present
compound also was tested against HDAC1, 2, 3, 4, 5, 8, 10, and 11.
The tested compounds showed a range of IC.sub.50 values vs. HDAC6
of about 1 nm to greater than 30 .mu.m, and a range of IC.sub.50
values vs. HDAC1 of about 91 nm to greater than 30 .mu.m.
Therefore, in some embodiments, a present HDACI is a selective
HDAC6 inhibitor which, because of a low affinity for other HDAC
isozymes, e.g., HDAC1, give rise to fewer side effects than
compounds that are non-selective HDAC inhibitors.
In some embodiments, the present HDACIs interact with and reduce
the activity of all histone deacetylases in a cell. In some
preferred embodiments, the present HDACIs interact with and reduce
the activity of fewer than all histone deacetylases in the cell. In
certain preferred embodiments, the present HDACIs interact with and
reduce the activity of one histone deacetylase (e.g., HDAC-6), but
do not substantially interact with or reduce the activities of
other histone deacetylases (e.g., HDAC-1, HDAC-2, HDAC-3, HDAC-4,
HDAC-5, HDAC-7, HDAC-8, HDAC-9, HDAC-10, and HDAC-11).
The present invention therefore provides HDACIs for the treatment
of a variety of diseases and conditions wherein inhibition of HDAC
has a beneficial effect. Preferably, a present HDACI is selective
for HDAC6 over the other HDAC isozymes by a factor of at least 2,
at least 5, at least 10, at least 20, at least 50, at least 100, at
least 500, at least 1000, at least 2000, at least 3000, and
preferably up to about 4000. For example, in various embodiments, a
present HDACI exhibits an IC.sub.50 value versus HDAC6 that is
about 350 or about 1000 times less than the IC.sub.50 value vs.
HDAC1, i.e., a selectivity ratio (HDAC1 IC.sub.50/HDAC6 IC.sub.50)
of about 350 or about 1000.
Other assays also showed a selectivity of a present HDACI for HDAC6
over HDAC1, 2, 3, 4, 5, 8, 10, and 11 of about 1000.
The IC.sub.50 values for compounds of structural formula (I) vs.
HDAC1 and HDAC6 were determined as follows:
The HDAC1, 2, 4, 5, 6, 7, 8, 9, 10, and 11 assays used isolated
recombinant human protein; HDAC3/NcoR2 complex was used for the
HDAC3 assay. Substrate for HDAC1, 2, 3, 6, 10, and 11 assays is a
fluorogenic peptide from p53 residues 379-382 (RHKKAc); substrate
for HDAC8 is fluorogenic diacyl peptide based on residues 379-382
of p53 (RHK.sub.AcK.sub.Ac). Acetyl-Lys(trifluoroacetyl)-AMC
substrate was used for HDAC4, 5, 7, and 9 assays. Compounds were
dissolved in DMSO and tested in 10-dose IC.sub.50 mode with 3-fold
serial dilution starting at 30 .mu.M. Control Compound Trichostatin
A (TSA) was tested in a 10-dose IC.sub.50 with 3-fold serial
dilution starting at 5 .mu.M. IC.sub.50 values were extracted by
curve-fitting the dose/response slopes. Assays were performed in
duplicate and IC.sub.50 values are an average of data from both
experiments.
Materials
Human HDAC1 (GenBank Accession No. NM_004964): Full length with
C-terminal GST tag, MW=79.9 kDa, expressed by baculovirus
expression system in Sf9 cells. Enzyme is in 50 mM Tris-HCl, pH
8.0, 138 mM NaCl, 20 mM glutathione, and 10% glycerol, and stable
for >6 months at -80.degree. C. Purity is >10% by SDS-PAGE.
Specific Activity is 20 U/.mu.g, where one U=1 pmol/min under assay
condition of 25 mM Tris/Cl, pH8.0, 137 mM NaCl, 2.7 mM KCl, 1 mM
MgCl.sub.2, 0.1 mg/ml BSA, 100 .mu.M HDAC substrate, and 13.2
ng/.mu.l HDAC1, incubation for 30 min at 30.degree. C.
Human HDAC6 (GenBank Accession No. BC069243): Full length with
N-terminal GST tag, MW=159 kDa, expressed by baculovirus expression
system in Sf9 cells. Enzyme is in 50 mM Tris-HCl, pH 8.0, 138 mM
NaCl, 20 mM glutathione, and 10% glycerol, and stable for >6
months at -80.degree. C. Purity is >90% by SDS-PAGE. Specific
Activity is 50 U/.mu.g, where one U=1 pmol/min under assay
condition of 25 mM Tris/Cl, pH8.0, 137 mM NaCl, 2.7 mM KCl, 1 mM
MgCl.sub.2, and 0.1 mg/ml BSA, 30 .mu.M HDAC substrate, and 5
ng/.mu.l HDAC6, incubation for 60 min at 30.degree. C.
Substrate for HDAC1 and HDAC6: Acetylated peptide substrate for
HDAC, based on residues 379-382 of p53 (Arg-His-Lys-Lys(Ac)), a
site of regulatory acetylation by the p300 and CBP
acetyltransferases (lysines 381, 382)1-6, is the best for HDAC from
among a panel of substrates patterned on p53, histone H3 and
histone H4 acetylation sites 7.
References: W. Gu et al., Cell (1997) 90 595; K. Sakaguchi et al.,
Genes Dev., (1998) 12 2831; L. Liu et al., Mol. Cell. Biol., (1999)
19 1202; A. Ito et al., EMBO J., (2001) 20 1331; N. A. Barley et
al., Mol. Cell, (2001) 8 1243; and A. Ito et al., EMBO J., (2002)
21 6236.
Reaction Buffer: 50 mM Tris-HCl, pH 8.0, 137 mM NaCl, 2.7 mM KCl, 1
mM MgCl.sub.2, 1 mg/ml BSA.
Assay Conditions
HDAC1: 75 nM HDAC1 and 50 .mu.M HDAC substrate are in the reaction
buffer and 1% DMSO final. Incubate for 2 hours at 30.degree. C.
HDAC6: 12.6 nM HDAC6 and 50 .mu.M HDAC substrate are in the
reaction buffer and 1% DMSO final. Incubate for 2 hours at
30.degree. C.
IC.sub.50 Calculations
All IC.sub.50 values are automatically calculated using the
GraphPad Prism version 5 and Equation of Sigmoidal dose-response
(variable slope): Y=Bottom+(Top-Bottom)/(1+10^((Log
EC50-X)*HillSlope)), where X is the logarithm of concentration, Y
is the response, Y starts at Bottom and goes to Top with a sigmoid
shape. In most cases, "Bottom" is set 0, and "Top" is set "less
than 120%". This is identical to the "four parameter logistic
equation". IC.sub.50 curves also are drawn using the GraphPad
Prism, and IC.sub.50 values and Hill slopes are provided.
HDAC Activity Assays: HDAC assay is performed using
fluorescently-labeled acetylated substrate, which comprises an
acetylated lysine side chain. After incubation with HDAC,
deacetylation of the substrate sensitizes the substrate such that,
in a second step, treatment with the detection enzyme produces a
fluorophore. HDACs 1 and 6 were expressed as full length fusion
proteins. Purified proteins were incubated with 50 .mu.M
fluorescently-labeled acetylated peptide substrate and test
compound for 2 hours at RT in HDAC assay buffer containing 50 mM
Tris-HCl (pH 8.0), 137 mM NaCl, 2.7 mM KCl, 1 mM MgCl.sub.2, 1%
DMSO, and 1% BSA.
Reactions were terminated by the addition of the Developer after 2
hours, and the development of fluorescence signal, which was
relative to the amount of deacetylated peptide, was monitored by
time-course measurement of En Vision (PerkinElmer). The HDAC
activity was estimated from the slope of time-course measurement of
the fluorescence intensity. The slope of no-enzyme control
(substrate alone) was served as background, and % Enzyme activity
was calculated using background-subtracted slope of no inhibitor
control (DMSO) as 100% activity.
To date, HDACIs have demonstrated a relatively non-specific
inhibition of various HDAC isozymes. Most HDACI so far identified
primarily inhibit HDAC 1, 2, 3, and 8, producing an
antiproliferative phenotype which is useful for oncology
applications, but not for the many non-oncology applications of
HDACIs. (K. B. Glaser et al, Biochemical and biophysical research
communications 2003, 310, 529-36.) The potential toxicities
associated with the inhibition of certain HDAC isozymes can lead to
additional difficulties for the clinical development of pan-HDAC,
i.e., nonselective HDAC, inhibitors. Because the network of
cellular effects mediated by acetylation is so vast and because
inhibition of some HDAC isozymes may lead to undesirable side
effects, HDAC isozyme selective inhibitors hold a greater
therapeutic promise than their nonselective counterparts.
As illustrated below, many HDACIs of the present invention exhibit
selective inhibition of HDAC6 compared to other HDAC isozymes.
TABLE-US-00003 TABLE 1 ##STR00136## HDAC Isoform IC.sub.50 (.mu.M)
HDAC6 Selectivity Cap 1 2 3 4 5 6 7 8 9 10 11 (HDAC1/HDAC6)
##STR00137## 5.83 9.19 0.00975 0.417 8.16 598 ##STR00138## 5.34
8.14 0.00251 0.397 18.3 2127 ##STR00139## 6.79 11.9 0.00511 0.911
14.5 1329 ##STR00140## 3.09 0.00146 2116 ##STR00141## 13.2 0.0293
0.324 451 ##STR00142## 5.86 0.0125 0.422 469 ##STR00143## 1.23
0.00294 0.174 418 ##STR00144## 0.940 0.00348 270 ##STR00145## 3.56
0.0142 251 ##STR00146## 6.47 0.0358 181 ##STR00147## 6.87 0.0119
577 ##STR00148## 5.73 0.0313 183 ##STR00149## 1.47 0.0109 135
##STR00150## 6.66 4.50 2.38 1.48 ##STR00151## 29.0 0.540 1.20 54
##STR00152## 27.9 0.150 1.82 186 ##STR00153## >30 1.25 5.59
>24 ##STR00154## >30 6.28 9.43 >5 ##STR00155## 23.8 0.124
1.92 192
TABLE-US-00004 TABLE 2 HDAC inhibition data for inventive HDACIs
and comparative HDAC inhibitors. ##STR00156## ##STR00157## HDAC
Isoform IC.sub.50 (.mu.M) Compound 1 2 3 4 5 6 7 8 9 10 11
Tubastatin A 16.4 >30 >30 >30 >30 0.015 >30 0.854
>30 >30 >30 Tubacin 1.40 6.27 1.27 17.3 3.35 0.004 9.70
1.27 4.31 3.71 3.79 Bicyclic 2 5.83 9.19 0.00975 0.417 8.16 4 5.34
8.14 0.00251 0.397 18.3 6 6.79 11.9 0.00511 0.911 14.5 8 12.2
0.00408 10 13.2 0.0293 0.324 12 0.940 0.00348 14 3.56 0.0142 17
1.23 0.00294 0.174 19 5.86 0.0125 0.422 21 3.09 0.00146 22 >30
>30 23 >30 >30 27 8.54 0.0294 30 0.623 0.000170 32 8.79
0.0092 36 14.7 0.0470 38 52.4 0.126 39 27.7 0.0470 Monocyclic 41
5.73 0.0313 43 1.47 0.0109 45 6.47 0.0358 47 6.87 0.0119 49 12.2
0.0701 Acyclic 51 >30 6.28 9.43 53 23.8 0.124 1.92 55 6.66 4.50
2.38 57 29.0 0.540 1.20 59 27.9 0.150 1.82 61 >30 1.25 5.59
Assays values are an average of two experiments. ISOX was
previously found to have a low picomolar IC.sub.50 at HDAC6. When
ISOX was tested in these assays, an HDAC6 IC.sub.50 value of 2.4 nM
was observed. After investigating the source of this discrepancy,
it was found that lack of a detergent (Triton X100) in the original
assay caused the anomalously high activity.
The present HDACIs demonstrate excellent HDAC6 potency and
selectivity, often exhibiting an IC.sub.50 of 1 to 5 nM at HDAC6
and 1000- to 2000-fold selectivity against HDAC1. For example, 4
demonstrated an IC.sub.50 of 2.5 nM at HDAC6 and 2125-fold
selectivity against HDAC1.
Ligand efficiency is an important metric to judge the value of
potential drugs. Ligand efficiency was proposed after finding that
the average molecular weight of successful drugs was lower than
those at an early clinical phase, thereby establishing an
association between lower molecular weight and a higher chance of
clinical success. (C. Abad-Zapatero, et al., Drug discovery today.
10:464-9, 2005; M. C. Wenlock, et al., J Med. Chem. 46:1250-6,
2003). This observation led to the use of ligand efficiency values
as an important value for determining lead and clinical candidate
selection. Ligand efficiency relates binding free energy to the
number of non-hydrogen atoms in the equation: .DELTA.g=AG/N, where
N is the number of non-hydrogen atoms, and .DELTA.g is ligand
efficiency. A closely related value used in the analysis of
deacetylase inhibitors is binding efficiency index, defined as
BEI=pIC50/MW, MW (molecular weight) in kDa.
With the aim of improving ligand efficiency, cap group size was
reduced while maintaining potency and selectivity. Table 3 shows
that indole, 2-methylindole, and 3-methylindole cap groups had a
potency at the target under 10 nM, while maintaining high HDAC6
selectivity. BEI was higher for the indole capped compounds
compared to Tubastatin. Substitution at the indole 3' position with
ethyldimethylamine or benzyl did not enhance potency at HDAC6
relative to the 2'-unsubstituted indole. Notably, the benzoyl
substituted compound had very low activity at HDAC1, below the
cutoff value for the assay. The importance of rigidity in the cap
group also was investigated through synthesis of compound 55. The
diphenylamine cap group of compound 55 is not locked in a planar
conformation. The diphenylamine capped compound 55 in Table 3 was
more potent and selective compared to an analogous carbazole
compound.
Table 3 also shows that the indole and the diphenylamine cap had
reduced HDAC8 activity relative to Tubastatin. For compounds to be
especially useful as molecular probes of HDAC6 activity, the
activity at HDAC8 should be minimized. The diphenylamine capped
compound was greatly enhanced in this respect, with over 500-fold
selectivity for HDAC6 vs HDAC8 Likewise, the 2-methyl and
3-methylindole cap groups had greater selectivity for HDAC6 over
HDAC8 compared to Tubastatin.
TABLE-US-00005 TABLE 3 HDAC6 Inhibitors Featuring an Indole
Scaffold. HDAC1 HDAC6 HDAC8 IC.sub.50 (.mu.M) IC.sub.50 (.mu.M)
IC.sub.50 (.mu.M) BEI Tubastatin 16.4 .+-. 2.6 0.015 .+-. 0.001
0.854 .+-. 0.040 23.3 ##STR00158## 5.83 .+-. 0.70 0.0097 .+-.
0.0003 0.417 .+-. 0.040 30.1 ##STR00159## 5.34 .+-. 0.16 0.0025
.+-. 0.0004 0.397 .+-. 0.099 30.7 ##STR00160## 6.79 .+-. 0.29
0.0051 .+-. 0.0008 0.911 .+-. 0.042 29.6 ##STR00161## 5.86 .+-.
1.19 0.013 .+-. 0.002 0.422 .+-. 0.044 23.4 ##STR00162## >30
0.035 .+-. 0.003 NT 20.9 ##STR00163## 6.66 .+-. 0.51 0.0045 .+-.
0.0004 2.38 .+-. 0.25 26.3
Values are the mean of two experiments. Data are shown as IC.sub.50
values in .mu.M.+-.standard deviation. Compounds were tested in
duplicate in 10-dose IC.sub.50 mode with 3-fold serial dilution
starting from 30 .mu.M solutions. IC.sub.50 values were extracted
by curve-fitting the dose/response slopes. Assays performed by
Reaction Biology Corp.
The present compounds have been evaluated for their activity at
HDAC6 and their selectivity for HDAC6 compared to HDAC1. It
previously was shown that selective HDAC6 inhibitors are implicated
in a variety of disease states including, but not limited to,
arthritis, autoimmune disorders, inflammatory disorders, cancer,
neurological diseases such as Rett syndrome, peripheral
neuropathies such as CMT, stroke, hypertension, and diseases in
which oxidative stress is a causative factor or a result thereof.
It also was shown that selective HDAC6 inhibitors, when
administered in combination with rapamycin, prolonged the lifespan
of mice with kidney xenografts. This model was used to evaluate the
immunosuppressant properties of the present compounds and serve as
a model of transplant rejection. Furthermore, it was previously
shown that selective HDAC6 inhibitors confer neuroprotection in rat
primary cortical neuron models of oxidative stress. These studies
identified selective HDAC6 inhibitors as non-toxic neuroprotective
agents. The present compounds behave in a similar manner because
they also are selective HDAC6 agents. The present compounds
demonstrate a ligand efficiency that renders them more drug-like in
their physiochemical properties. In addition, the present compounds
maintain the potency and selectivity observed in prior HDACIs. The
present compounds therefore are pharmaceutical candidates and
research tools to identify the specific functions of HDAC6.
In one embodiment, the present invention relates to a method of
treating an individual suffering from a disease or condition
wherein inhibition of HDACs provides a benefit comprising
administering a therapeutically effective amount of a present HDACI
compound to an individual in need thereof.
The methods described herein relate to the use of a present HDACI
and an optional second therapeutic agent useful in the treatment of
diseases and conditions wherein inhibition of HDAC provides a
benefit. The methods of the present invention can be accomplished
by administering a present HDACI as the neat compound or as a
pharmaceutical composition. Administration of a pharmaceutical
composition, or a neat HDACI of the present invention, can be
performed during or after the onset of the disease or condition of
interest. Typically, the pharmaceutical compositions are sterile,
and contain no toxic, carcinogenic, or mutagenic compounds that
would cause an adverse reaction when administered.
In many embodiments, a present HDACI is administered in conjunction
with a second therapeutic agent useful in the treatment of a
disease or condition wherein inhibition of HDAC provides a benefit.
The second therapeutic agent is different from the present HDACI. A
present HDACI and the second therapeutic agent can be administered
simultaneously or sequentially. In addition, a present HDACI and
second therapeutic agent can be administered from a single
composition or two separate compositions. A present HDACI and the
second therapeutic agent can be administered simultaneously or
sequentially to achieve the desired effect.
The second therapeutic agent is administered in an amount to
provide its desired therapeutic effect. The effective dosage range
for each second therapeutic agent is known in the art, and the
second therapeutic agent is administered to an individual in need
thereof within such established ranges.
The present invention therefore is directed to compositions and
methods of treating diseases or conditions wherein inhibition of
HDAC provides a benefit. The present invention also is directed to
pharmaceutical compositions comprising a present HDACI and an
optional second therapeutic agent useful in the treatment of
diseases and conditions wherein inhibition of HDAC provides a
benefit. Further provided are kits comprising a present HDACI and,
optionally, a second therapeutic agent useful in the treatment of
diseases and conditions wherein inhibition of HDAC provides a
benefit, packaged separately or together, and an insert having
instructions for using these active agents.
A present HDACI and the second therapeutic agent can be
administered together as a single-unit dose or separately as
multi-unit doses, wherein the present HDACI is administered before
the second therapeutic agent or vice versa. One or more dose of a
present HDACI and/or one or more dose of the second therapeutic
agent can be administered. The present HDACIs therefore can be used
in conjunction with one or more second therapeutic agents, for
example, but not limited to, anticancer agents.
Within the meaning of the present invention, the term "disease" or
"condition" denotes disturbances and/or anomalies that as a rule
are regarded as being pathological conditions or functions, and
that can manifest themselves in the form of particular signs,
symptoms, and/or malfunctions. As demonstrated below, a present
HDACI is a potent inhibitor of HDAC and can be used in treating
diseases and conditions wherein inhibition of HDAC provides a
benefit, for example, cancer, a neurological disease, a
neurodegenerative condition, traumatic brain injury, stroke, an
inflammation, an autoimmune disease, autism, and malaria.
In one preferred embodiment, the present invention provides methods
for treating cancer, including but not limited to killing a cancer
cell or neoplastic cell; inhibiting the growth of a cancer cell or
neoplastic cell; inhibiting the replication of a cancer cell or
neoplastic cell; or ameliorating a symptom thereof, said methods
comprising administering to a subject in need thereof a
therapeutically effective amount of a present HDACI.
In one embodiment, the invention provides a method for treating
cancer comprising administering to a subject in need thereof an
amount of a present HDACI or a pharmaceutically acceptable salt
thereof sufficient to treat the cancer. A present HDACI can be used
as the sole anticancer agent, or in combination with another
anticancer treatment, e.g., radiation, chemotherapy, and
surgery.
In another embodiment, the invention provides a method for
increasing the sensitivity of a cancer cell to the cytotoxic
effects of radiotherapy and/or chemotherapy comprising contacting
the cell with a present HDACI or a pharmaceutically acceptable salt
thereof in an amount sufficient to increase the sensitivity of the
cell to the cytotoxic effects of radiotherapy and/or
chemotherapy.
In a further embodiment, the present invention provides a method
for treating cancer comprising: (a) administering to an individual
in need thereof an amount of a present HDACI compound; and (b)
administering to the individual an amount of radiotherapy,
chemotherapy, or both. The amounts administered are each effective
to treat cancer. In another embodiment, the amounts are together
effective to treat cancer.
In another embodiment, the invention provides a method for treating
cancer, said method comprising administering to a subject in need
thereof a pharmaceutical composition comprising an amount of a
present HDACI effective to treat cancer.
This combination therapy of the invention can be used accordingly
in a variety of settings for the treatment of various cancers. In a
specific embodiment, the individual in need of treatment has
previously undergone treatment for cancer. Such previous treatments
include, but are not limited to, prior chemotherapy, radiotherapy,
surgery, or immunotherapy, such as cancer vaccines.
In another embodiment, the cancer being treated is a cancer which
has demonstrated sensitivity to radiotherapy and/or chemotherapy or
is known to be responsive to radiotherapy and/or chemotherapy. Such
cancers include, but are not limited to, non-Hodgkin's lymphoma,
Hodgkin's disease, Ewing's sarcoma, testicular cancer, prostate
cancer, ovarian cancer, bladder cancer, larynx cancer, cervical
cancer, nasopharynx cancer, breast cancer, colon cancer, pancreatic
cancer, head and neck cancer, esophageal cancer, rectal cancer,
small-cell lung cancer, non-small cell lung cancer, brain tumors,
or other CNS neoplasms.
In still another embodiment, the cancer being treated has
demonstrated resistance to radiotherapy and/or chemotherapy or is
known to be refractory to radiotherapy and/or chemotherapy. A
cancer is refractory to a therapy when at least some significant
portion of the cancer cells are not killed or their cell division
is not arrested in response to therapy. Such a determination can be
made either in vivo or in vitro by any method known in the art for
assaying the effectiveness of treatment on cancer cells, using the
art-accepted meanings of "refractory" in such a context. In a
specific embodiment, a cancer is refractory where the number of
cancer cells has not been significantly reduced or has
increased.
Other cancers that can be treated with the compounds and methods of
the invention include, but are not limited to, cancers and
metastases selected from the group consisting of solid tumors,
including but not limited to: fibrosarcoma, myxosarcoma,
liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma,
angiosarcoma, endotheliosarcoma, lymphangiosarcoma,
lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's
tumor, leiornyosarcoma, rhabdomyosarcoma, colon cancer, colorectal
cancer, kidney cancer, pancreatic cancer, bone cancer, breast
cancer, ovarian cancer, prostate cancer, esophageal cancer, stomach
cancer, oral cancer, nasal cancer, throat cancer, squamous cell
carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland
carcinoma, sebaceous gland carcinoma, papillary carcinoma,
papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma,
bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct
carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms'
tumor, cervical cancer, uterine cancer, testicular cancer, small
cell lung carcinoma, bladder carcinoma, lung cancer, epithelial
carcinoma, glioma, glioblastoma multiforma, astrocytoma,
medulloblastoma, craniopharyngioma, ependymoma, pinealoma,
hemangioblastoma, acoustic neuroma, oligodendroglioma, meningioma,
skin cancer, melanoma, neuroblastoma, and retinoblastoma;
blood-borne cancers, including but not limited to: acute
lymphoblastic leukemia, acute lymphoblastic B-cell leukemia, acute
lymphoblastic T-cell leukemia, acute myeloblastic leukemia, acute
promyelocytic leukemia, acute monoblastic leukemia, acute
erythroleukemic leukemia, acute megakaryoblastic leukemia, acute
myclomonocytic leukemia, acute nonlymphocyctic leukemia, acute
undifferentiated leukemia, chronic myclocytic leukemia, chronic
lymphocytic leukemia, hairy cell leukemia, and multiple myeloma;
acute and chronic leukemias: lymphoblastic, myelogenous
lymphocytic, and myelocytic leukemias; lymphomas: Hodgkin's disease
and non-Hodgkin's lymphoma; multiple myeloma; Waldenstrom's
macroglobulinemia; heavy chain disease; and polycythemia vera.
The present HDACIs can also be administered to prevent progression
to a neoplastic or malignant state, including but not limited to
the cancers listed above. Such prophylactic use is indicated in
conditions known or suspected of preceding progression to neoplasia
or cancer, in particular, where non-neoplastic cell growth
consisting of hyperplasia, metaplasia, or most particularly,
dysplasia has occurred (for review of such abnormal growth
conditions, see Robbins and Angell, 1976, Basic Pathology, 2d Ed.,
W.B. Saunders Co., Philadelphia, pp. 68-79). Hyperplasia is a form
of controlled cell proliferation involving an increase in cell
number in a tissue or organ, without significant alteration in
structure or function. For example, endometrial hyperplasia often
precedes endometrial cancer and precancerous colon polyps often
transform into cancerous lesions. Metaplasia is a form of
controlled cell growth in which one type of adult or fully
differentiated cell substitutes for another type of adult cell.
Metaplasia can occur in epithelial or connective tissue cells. A
typical metaplasia involves a somewhat disorderly metaplastic
epithelium. Dysplasia is frequently a forerunner of cancer, and is
found mainly in the epithelia; it is the most disorderly form of
non-neoplastic cell growth, involving a loss in individual cell
uniformity and in the architectural orientation of cells.
Dysplastic cells often have abnormally large, deeply stained
nuclei, and exhibit pleomorphism. Dysplasia characteristically
occurs where chronic irritation or inflammation exists, and often
is found in the cervix, respiratory passages, oral cavity, and gall
bladder.
Alternatively or in addition to the presence of abnormal cell
growth characterized as hyperplasia, metaplasia, or dysplasia, the
presence of one or more characteristics of a transformed phenotype,
or of a malignant phenotype, displayed in vivo or displayed in
vitro by a cell sample from a subject, can indicate the
desirability of prophylactic/therapeutic administration of the
composition of the invention. Such characteristics of a transformed
phenotype include, for example, morphology changes, looser
substratum attachment, loss of contact inhibition, loss of
anchorage dependence, protease release, increased sugar transport,
decreased serum requirement, expression of fetal antigens,
disappearance of the 250,000 dalton cell surface protein.
In a specific embodiment, leukoplakia, a benign-appearing
hyperplastic or dysplastic lesion of the epithelium, or Bowen's
disease, a carcinoma in situ, are pre-neoplastic lesions indicative
of the desirability of prophylactic intervention.
In another embodiment, fibrocystic disease (cystic hyperplasia,
mammary dysplasia, particularly adenosis (benign epithelial
hyperplasia)) is indicative of the desirability of prophylactic
intervention.
The prophylactic use of the compounds and methods of the present
invention are also indicated in some viral infections that may lead
to cancer. For example, human papilloma virus can lead to cervical
cancer (see, e.g., Hernandez-Avila et al., Archives of Medical
Research (1997) 28:265-271), Epstein-Ban virus (EBV) can lead to
lymphoma (see, e.g., Herrmann et al., J Pathol (2003)
199(2):140-5), hepatitis B or C virus can lead to liver carcinoma
(see, e.g., El-Serag, J Clin Gastroenterol (2002) 35(5 Suppl
2):572-8), human T cell leukemia virus (HTLV)-I can lead to T-cell
leukemia (see e.g., Mortreux et al., Leukemia (2003) 17(1):26-38),
human herpesvirus-8 infection can lead to Kaposi's sarcoma (see,
e.g., Kadow et al., Curr Opin Investig Drugs (2002) 3(11):1574-9),
and Human Immune deficiency Virus (HIV) infection contribute to
cancer development as a consequence of immunodeficiency (see, e.g.,
Dal Maso et al., Lancet Oncol (2003) 4(2):110-9).
In other embodiments, a subject exhibiting one or more of the
following predisposing factors for malignancy can be treated by
administration of the present HDACIs and methods of the invention:
a chromosomal translocation associated with a malignancy (e.g., the
Philadelphia chromosome for chronic myelogenous leukemia, t(14;18)
for follicular lymphoma, etc.), familial polyposis or Gardner's
syndrome (possible forerunners of colon cancer), benign monoclonal
gammopathy (a possible forerunner of multiple myeloma), a first
degree kinship with persons having a cancer or procancerous disease
showing a Mendelian (genetic) inheritance pattern (e.g., familial
polyposis of the colon, Gardner's syndrome, hereditary exostosis,
polyendocrine adenomatosis, medullary thyroid carcinoma with
amyloid production and pheochromocytoma, Peutz-Jeghers syndrome,
neurofibromatosis of Von Recklinghausen, retinoblastoma, carotid
body tumor, cutaneous melanocarcinoma, intraocular melanocarcinoma,
xeroderma pigmentosum, ataxia telangiectasia, Chediak-Higashi
syndrome, albinism, Fanconi's aplastic anemia, and Bloom's
syndrome; see Robbins and Angell, 1976, Basic Pathology, 2d Ed.,
W.B. Saunders Co., Philadelphia, pp. 112-113) etc.), and exposure
to carcinogens (e.g., smoking, and inhalation of or contacting with
certain chemicals).
In another specific embodiment, the present HDACIs and methods of
the invention are administered to a human subject to prevent
progression of breast, colon, ovarian, or cervical cancer.
In one embodiment, the invention provides methods for treating
cancer comprising (a) administering to an individual in need
thereof an amount of a present HDACI; and (b) administering to the
individual one or more additional anti-cancer treatment modality
including, but not limited to, radiotherapy, chemotherapy, surgery
or immunotherapy, such as a cancer vaccine. In one embodiment, the
administering of step (a) is prior to the administering of step
(b). In another embodiment, the administering of step (a) is
subsequent to the administering of step (b). In still another
embodiment, the administering of step (a) is concurrent with the
administering of step (b).
In one embodiment, the additional anticancer treatment modality is
radiotherapy and/or chemotherapy. In another embodiment, the
additional anticancer treatment modality is surgery.
In still another embodiment, the additional anticancer treatment
modality is immunotherapy, such as cancer vaccines.
In one embodiment, a present HDACI or a pharmaceutically acceptable
salt thereof is administered adjunctively with the additional
anticancer treatment modality.
In a preferred embodiment, the additional anticancer treatment
modality is radiotherapy. In the methods of the present invention,
any radiotherapy protocol can be used depending upon the type of
cancer to be treated. Embodiments of the present invention employ
electromagnetic radiation of: gamma-radiation (10.sup.-20 to
10.sup.-13 m), X-ray radiation (10.sup.-12 to 10.sup.-9 m),
ultraviolet light (10 nm to 400 nm), visible light (400 nm to 700
nm), infrared radiation (700 nm to 1 mm), and microwave radiation
(1 mm to 30 cm).
For example, but not by way of limitation, X-ray radiation can be
administered; in particular, high-energy megavoltage (radiation of
greater that 1 MeV energy) can be used for deep tumors, and
electron beam and orthovoltage X-ray radiation can be used for skin
cancers. Gamma-ray emitting radioisotopes, such as radioactive
isotopes of radium, cobalt and other elements, can also be
administered. Illustrative radiotherapy protocols useful in the
present invention include, but are not limited to, stereotactic
methods where multiple sources of low dose radiation are
simultaneously focused into a tissue volume from multiple angles;
"internal radiotherapy," such as brachytherapy, interstitial
irradiation, and intracavitary irradiation, which involves the
placement of radioactive implants directly in a tumor or other
target tissue; intraoperative irradiation, in which a large dose of
external radiation is directed at the target tissue which is
exposed during surgery; and particle beam radiotherapy, which
involves the use of fast-moving subatomic particles to treat
localized cancers.
Many cancer treatment protocols currently employ radiosensitizers
activated by electromagnetic radiation, e.g., X-rays. Examples of
X-ray-activated radiosensitizers include, but are not limited to,
metronidazole, misonidazole, desmethylmisonidazole, pimonidazole,
etanidazole, nimorazole, mitomycin C, RSU 1069, SR 4233, EO9, RB
6145, nicotinamide, 5-bromodeoxyuridine (BUdR), 5-iododeoxyuridine
(IUdR), bromodeoxycytidine, fluorodeoxyuridine (FUdR), hydroxyurea,
cis-platin, and therapeutically effective analogs and derivatives
of the same.
Photodynamic therapy (PDT) of cancers employs visible light as the
radiation activator of the sensitizing agent. Examples of
photodynamic radiosensitizers include the following, but are not
limited to: hematoporphyrin derivatives, PHOTOFRIN.RTM.,
benzoporphyrin derivatives, NPe6, tin etioporphyrin (SnET2),
pheoborbide-a, bacteriochlorophylla, naphthalocyanines,
phthalocyanines, zinc phthalocyanine, and therapeutically effective
analogs and derivatives of the same.
Radiosensitizers can be administered in conjunction with a
therapeutically effective amount of one or more compounds in
addition to a present HDACI, such compounds including, but not
limited to, compounds that promote the incorporation of
radiosensitizers to the target cells, compounds that control the
flow of therapeutics, nutrients, and/or oxygen to the target cells,
chemotherapeutic agents that act on the tumor with or without
additional radiation, or other therapeutically effective compounds
for treating cancer or other disease. Examples of additional
therapeutic agents that can be used in conjunction with
radiosensitizers include, but are not limited to, 5-fluorouracil
(5-FU), leucovorin, oxygen, carbogen, red cell transfusions,
perfluorocarbons (e.g., FLUOSOLW.RTM.-DA), 2,3-DPG, BW12C, calcium
channel blockers, pentoxifylline, antiangiogenesis compounds,
hydralazine, and L-BSO.
In a preferred embodiment, a present HDACI or a pharmaceutically
acceptable salt thereof is administered prior to the administration
of radiotherapy and/or chemotherapy.
In another preferred embodiment, a present HDACI or a
pharmaceutically acceptable salt thereof is administered
adjunctively with radiotherapy and/or chemotherapy.
A present HDACI and additional treatment modalities can act
additively or synergistically (i.e., the combination of a present
HDACI or a pharmaceutically acceptable salt thereof, and an
additional anticancer treatment modality is more effective than
their additive effects when each are administered alone). A
synergistic combination permits the use of lower dosages of a
present HDACI and/or the additional treatment modality and/or less
frequent administration of a present HDACI and/or additional
treatment modality to a subject with cancer. The ability to utilize
lower dosages of a present HDACI and/or an additional treatment
modality and/or to administer a compound of the invention and the
additional treatment modality less frequently can reduce the
toxicity associated with the administration without reducing the
efficacy of a present HDACI and/or the additional treatment
modality in the treatment of cancer. In addition, a synergistic
effect can result in the improved efficacy of the treatment of
cancer and/or the reduction of adverse or unwanted side effects
associated with the administration of a present HDACI and/or an
additional anticancer treatment modality as monotherapy.
In one embodiment, the present HDACIs may act synergistically with
radiotherapy when administered in doses typically employed when
such HDACIs are used alone for the treatment of cancer. In another
embodiment, the present HDACIs may act synergistically with
radiotherapy when administered in doses that are less than doses
typically employed when such HDACIs are used as monotherapy for the
treatment of cancer.
In one embodiment, radiotherapy may act synergistically with a
present HDACI when administered in doses typically employed when
radiotherapy is used as monotherapy for the treatment of cancer. In
another embodiment, radiotherapy may act synergistically with a
compound of the invention when administered in doses that are less
than doses typically employed when radiotherapy is used as
monotherapy for the treatment of cancer.
The effectiveness of the HDACIs as HDAC inhibitors for sensitizing
cancer cells to the effect of radiotherapy can be determined by the
in vitro and/or in vivo determination of post-treatment survival
using techniques known in the art. In one embodiment, for in vitro
determinations, exponentially growing cells can be exposed to known
doses of radiation, and the survival of the cells monitored.
Irradiated cells are plated and cultured for about 14- about 21
days, and the colonies are stained. The surviving fraction is the
number of colonies divided by the plating efficiency of
unirradiated cells. Graphing the surviving fraction on a log scale
versus the absorbed dose on a linear scale generates a survival
curve. Survival curves generally show an exponential decrease in
the fraction of surviving cells at higher radiation doses after an
initial shoulder region in which the dose is sublethal. A similar
protocol can be used for chemical agents when used in the
combination therapies of the invention.
Inherent radiosensitivity of tumor cells and environmental
influences, such as hypoxia and host immunity, can be further
assessed by in vivo studies. The growth delay assay is commonly
used. This assay measures the time interval required for a tumor
exposed to radiation to regrow to a specified volume. The dose
required to control about 50% of tumors is determined by the
TCD.sub.50 assay.
In vivo assay systems typically use transplantable solid tumor
systems in experimental subjects. Radiation survival parameters for
normal tissues as well as for tumors can be assayed using in vivo
methods known in the art.
The present invention provides methods of treating cancers
comprising the administration of an effective amount of a present
HDACI in conjunction with recognized methods of surgery,
radiotherapy, and chemotherapies, including, for example,
chemical-based mimics of radiotherapy whereby a synergistic
enhancement of the effectiveness of the recognized therapy is
achieved. The effectiveness of a treatment can be measured in
clinical studies or in model systems, such as a tumor model in
mice, or cell culture sensitivity assays.
The present invention provides combination therapies that result in
improved effectiveness and/or reduced toxicity. Accordingly, in one
aspect, the invention relates to the use of the present HDACIs as
radiosensitizers in conjunction with radiotherapy.
When the combination therapy of the invention comprises
administering a present HDACI with one or more additional
anticancer agents, the present HDACI and the additional anticancer
agents can be administered concurrently or sequentially to an
individual. The agents can also be cyclically administered. Cycling
therapy involves the administration of one or more anticancer
agents for a period of time, followed by the administration of one
or more different anticancer agents for a period of time and
repeating this sequential administration, i.e., the cycle, in order
to reduce the development of resistance to one or more of the
anticancer agents of being administered, to avoid or reduce the
side effects of one or more of the anticancer agents being
administered, and/or to improve the efficacy of the treatment.
An additional anticancer agent may be administered over a series of
sessions; anyone or a combination of the additional anticancer
agents listed below may be administered.
The present invention includes methods for treating cancer
comprising administering to an individual in need thereof a present
HDACI and one or more additional anticancer agents or
pharmaceutically acceptable salts thereof. A present HDACI and the
additional anticancer agent can act additively or synergistically.
Suitable anticancer agents include, but are not limited to,
gemcitabine, capecitabine, methotrexate, taxol, taxotere,
mereaptopurine, thioguanine, hydroxyurea, cyclophosphamide,
ifosfamide, nitrosoureas, mitomycin, dacarbazine, procarbizine,
etoposide, teniposide, campatheeins, bleomycin, doxorubicin,
idarubicin, daunorubicin, dactinomycin, plicamycin, mitoxantrone,
L-asparaginase, doxorubicin, epirubicin, 5-fluorouracil (5-FU),
taxanes (such as docetaxel and paclitaxel), leucovorin, levamisole,
irinotecan, estramustine, etoposide, nitrogen mustards, BCNU,
nitrosoureas (such as carmustine and lomustine), platinum complexes
(such as cisplatin, carboplatin and oxaliplatin), imatinib
mesylate, hexamethylmelamine, topotecan, tyrosine kinase
inhibitors, tyrphostins herbimycin A, genistein, erbstatin, and
lavendustin A.
In one embodiment, the anti-cancer agent can be, but is not limited
to, a drug selected from the group consisting of alkylating agents,
nitrogen mustards, cyclophosphamide, trofosfamide, chlorambucil,
nitrosoureas, carmustine (BCNU), lomustine (CCNU),
alkylsulphonates, busulfan, treosulfan, triazenes, plant alkaloids,
vinca alkaloids (vineristine, vinblastine, vindesine, vinorelbine),
taxoids, DNA topoisomcrase inhibitors, epipodophyllins,
9-aminocamptothecin, camptothecin, crisnatol, mitomycins, mitomycin
C, anti-metabolites, anti-folates, DHFR inhibitors, trimetrexate,
IMP dehydrogenase inhibitors, mycophenolic acid, tiazofurin,
ribavirin, EICAR, ribonucleotide reductase inhibitors, hydroxyurea,
deferoxamine, pyrimidine analogs, uracil analogs, floxuridine,
doxifluridine, ratitrexed, cytosine analogs, cytarabine (ara C),
cytosine arabinoside, fludarabine, purine analogs, mercaptopurine,
thioguanine, DNA antimetabolites, 3-HP, 2'-deoxy-5-fluorouridine,
5-HP, alpha-TGDR, aphidicolin glycinate, ara-C,
5-aza-2'-deoxycytidine, beta-TGDR, cyclocytidine, guanazole
(inosine glycodialdehyde), macebecin II, pyrazoloimidazole,
hormonal therapies, receptor antagonists, anti-estrogen, tamoxifen,
raloxifene, megestrol, LHRH agonists, goserelin, leuprolide
acetate, anti-androgens, flutamide, bicalutamide,
retinoids/deltoids, cis-retinoic acid, vitamin A derivative,
all-trans retinoic acid (ATRA-IV), vitamin D3 analogs, E1) 1089, CB
1093, ICH 1060, photodynamic therapies, vertoporfin, BPD-MA,
phthalocyanine, photosensitizer Pc4, demethoxy-hypocrellin A
(2BA-2DMHA), cytokines, interferon-.alpha., interferon-I3,
interferon-.gamma., tumor necrosis factor, angiogenesis inhibitors,
angiostatin (plasminogen fragment), antiangiogenic antithrombin UI,
angiozyme, ABT-627, Bay 12-9566, benefin, bevacizumab, BMS-275291,
cartilage-derived inhibitor (CDI), CAI, CD59 complement fragment,
CEP-7055, Col 3, combretastatinA-4, endostatin (collagen XVIII
fragment), fibronectin fragment, Gro-beta, halofuginone,
heparinases, heparin hexasaccharide fragment, HMV833, human
chorionic gonadotropin (hCG), IM-862, interferon inducible protein
(IP-10), interleukin-12, kringle 5 (plasminogen fragment),
marimastat, metalloproteinase inhibitors (UMPs),
2-methoxyestradiol, MMI 270 (CGS 27023A), MoAb IMC-I C11,
neovastat, NM-3, panzem, P1-88, placental ribonuclease inhibitor,
plasminogen activator inhibitor, platelet factor-4 (PF4),
prinomastat, prolactin 161(D fragment, proliferin-related protein
(PRP), PTK 787/ZK 222594, retinoids, solimastat, squalamine, SS
3304, SU 5416, SU 6668, SU 11248, tetrahydrocortisol-S,
tetrathiomolybdate, thalidomide, thrombospondin-1 (TSP-1), TNP-470,
transforming growth factor-beta (TGF-11), vasculostatin, vasostatin
(calreticulin fragment), ZD 6126, ZD 6474, farnesyl transferase
inhibitors (FTI), bisphosphonates, antimitotic agents,
allocolchicine, halichondrin B, colchicine, colchicine derivative,
dolstatin 10, maytansine, rhizoxin, thiocolchicine, trityl
cysteine, isoprenylation inhibitors, dopaminergic neurotoxins,
1-methyl-4-phenylpyridinium ion, cell cycle inhibitors,
staurosporine, actinomycins, actinomycin D, dactinomycin,
bleomycins, bleomycin A2, bleomycin B2, peplomycin, anthracycline,
adriamycin, epirubicin, pirarnbicin, zorubicin, mitoxantrone, MDR
inhibitors, verapamil, Ca.sup.2, ATPase inhibitors, and
thapsigargin.
Other anti-cancer agents that may be used in the present invention
include, but are not limited to, acivicin; aclarubicin; acodazole
hydrochloride; acronine; adozelesin; aldesleukin; altretamine;
arnbomycin; ametantrone acetate; aminoglutethimide; amsacrine;
anastrozole; anthramycin; asparaginase; asperlin; azacitidine;
azetepa; azotomycin; batimastat; benzodepa; bicalutamide;
bisantrene hydrochloride; bisnafide dimesylate; bizelcsin;
bleomycin sulfate; brequinar sodium; bropirimine; busul fan;
cactinomycin; calusterone; caracemide; carbetimer; carmustine;
carubicin hydrochloride; carzelesin; cedefingol; chlorambucil;
cirolemycin; cisplatin; cladribine; crisnatol mesylate;
cyclophosphamide; cytarabine; dacarbazine; dactinomycin;
daunorubicin hydrochloride; decitabine; dexorrnaplatin;
dezaguanine; dezaguanine mesylate; diaziquone; docetaxel;
doxorubicin hydrochloride; droloxifene; droloxifene citrate;
dromostanolone propionate; duazomycin; edatrexate; eflomithine
hydrochloride; elsamitrucin; enloplatin; enpromate; epipropidine;
epirubicin hydrochloride; erbulozole; esorubicin hydrochloride;
estramustine; estramustine phosphate sodium; etanidazole; etoposide
phosphate; etoprine; fadrozole hydrochloride; fazarabine;
fenretinide; floxuridine; fludarabine phosphate; fluorouracil;
fluorocitabine; fosquidone; fostriecin sodium; gemcitabine
hydrochloride; hydroxyurea; idarubicin hydrochloride; ifosfamide;
ilmofosine; interleukin II (including recombinant interleukin II,
or rIL2), interferon alfa-2a; interferon alfa-2b; interferon
alfa-n1; interferon alfa-n3; interferon beta-Ia; interferon
gamma-Ib; iproplatin; irinotecan hydrochloride; lanreotide acetate;
letrozole; leuprolide acetate; liarozole hydrochloride; lometrexol
sodium; lomustine; losoxantrone hydrochloride; masoprocol;
maytansine; mecchlorethamine hydrochloride; megestrol acetate;
melengestrol acetate; melphalan; menogaril; mercaptopurine;
methotrexate sodium; metoprine; meturedepa; mitindomide;
mitocarcin; mitocromin; mitogillin; mitomalcin; mitomycin;
mitusper; mitotane; mitoxantrone hydrochloride; mycophenolic acid;
nocodazole; nogalamycin; ormaplatin; oxisuran; pegaspargase;
peliomycin; pentamustine; peplomycin sulfate; perfosfamide;
pipobroman; piposulfan; piroxantrone hydrochloride; plicamycin;
plomestane; porfimer sodium; porfiromycin; prednimustine;
procarbazine hydrochloride; puromycin; puromycin hydrochloride;
pyrazofurin; riboprine; rogletimide; safingol; safingol
hydrochloride; semustine; simtrazene; sparfosate sodium;
sparsornycin; spirogermanium hydrochloride; spiromustine;
spiroplatin; streptonigrin; streptozocin; sulofenur; talisomycin;
tecogalan sodium; tegafur; teloxantrone hydrochloride; temoporfin;
teroxirone; testolactone; thiamiprine; thioguanine; thiotepa;
tiazofurin; tirapazamine; toremifene citrate; trestolone acetate;
triciribine phosphate; trimetrexate; trimetrexate glucuronate;
triptorelin; tubulozole hydrochloride; uracit mustard; uredepa;
vapreotide; verteporfln; vinblastine sulfate; vincristine sulfate;
vindesine; vindesine sulfate; vinepidine sulfate; vinglycinate
sulfate; vinleurosine sulfate; vinorelbine tartrate; vinrosidine
sulfate; vinzolidine sulfate; vorozolc; zeniplatin; zinostatin;
zorubicin hydrochloride.
Further anti-cancer drugs that can be used in the present invention
include, but are not limited to: 17-AAG; 20-epi-1,
25-dihydroxyvitamin D3; 5-ethynyluracil; abiraterone; aclarubicin;
acylfulvene; adecypenol; adozelesin; aldesleukin; ALL TK
antagonists; altretamine; ambamustine; amidox; amifostine;
aminolevulinic acid; amrubicin; amsacrine; anagrelide; anastrozole;
andrographolide; angiogenesis inhibitors; antagonist D; antagonist
G; antarelix; anti-dorsalizing morphogenetic protein 1;
antiandrogen, prostatic carcinoma; antiestrogen; antineoplaston;
antisense oligonucleotides; aphidicolin glycinate; apoptosis gene
modulators; apoptosis regulators; apurinic acid; ara CDP DL PTBA;
arginine deaminase; asulacrine; atamestane; atrimustine;
axinastatin 1; axinastatin 2; axinastatin 3; azasetron; azatoxin;
azatyrosine; baccatin III derivatives; balanol; batimastat; BCR-ABL
antagonists; benzochlorins; benzoylstaurosporine; beta lactam
derivatives; beta alethine; betaclarnycin B; betulinic acid; bFGF
inhibitor; bicalutamide; bisantrene; bisaziridinylsperrnine;
bisnafide; bistratene A; bizelesin; bortezomib; breflate;
bropirimine; budotitane; buthionine sulfoximine; calcipotriol;
calphostin C; camptothecin derivatives; canarypox IL-2; carboxamide
amino triazole; carboxyamidotriazole; CaRest M3; CARN 700;
cartilage derived inhibitor; carzelesin; casein kinase inhibitors;
castanospermine; cecropin B; cetrorelix; chlorins;
chloroquinoxaline sulfonamide; cicaprost; cis porphyrin;
cladribine; clomifene analogues; clotrimazole; collismycin A;
collismycin B; combretastatin A4; combretastatin analogue;
conagenin; crambescidin 816; crisnatol; cryptophycin 8;
cryptophycin A derivatives; curacin A; cyclopentanthraquinones;
cycloplatam; cypemycin; cytarabine ocfosfate; cytolytic factor;
cytostatin; dacliximab; decitabine; dehydrodidemnin B; deslorelin;
dexamethasone; dexifosfamide; dexrazoxane; dexveraparnil;
diaziquone; didemnin B; didox; diethylnorspermine; dihydro 5
azacytidine; dihydrotaxol, 9; dioxamycin; diphenyl spiromustine;
docetaxel; docosanol; dolasetron; doxifluridine; droloxifene;
dronabinol; duocarmycin SA; ebselen; ecomustine; edelfosine;
edrecolomab; eflomithine; elemene; emitefur; epirubicin;
epristeride; estramustine analogue; estrogen agonists; estrogen
antagonists; etanidazole; etoposide phosphate; exemestane;
fadrozole; fazarabine; fenretinide; filgrastim; finasteride;
flavopiridol; flezelastine; fluasterone; fltidarabine;
fluorodaunoruniein hydrochloride; forfenimex; formestane;
fostriecin; fotemustine; gadolinium texaphyrin; gallium nitrate;
galocitabine; ganirelix; gelatinase inhibitors; glutathione
inhibitors; hepsulfam; heregulin; hexamethylene bisacetamide;
hypericin; ibandronic acid; idarubicin; idoxifene; idramantone;
ilmofosine; ilomastat; imidazoacridones; imiquimod; immunostimulant
peptides; insulin like growth factor 1 receptor inhibitor;
interferon agonists; interferons; interleukins; iobenguane;
iododoxorubiein; ipomeanol, 4; iroplact; irsogladine; isobengazole;
isohomohalicondrin B; itasetron; jasplakinolide; kahalalide F;
larnellarin N triacetate; lanreotide; leinamycin; lenograstim;
lentinan sulfate; leptolstatin; letrozole; leukemia inhibiting
factor; leukocyte alpha interferon;
leuprolide+estrogen+progesterone; leuprorelin; levamisole;
liarozole; linear polyamine analogue; lipophilic disaccharide
peptide; lipophilic platinum complexes; lissoclinamide 7;
lobaplatin; lombricine; lometrexol; lonidamine; losoxantrone;
lovastatin; loxoribine; lurtotecan; lutetium texaphyrin;
lysofylline; lytic peptides; maitansine; mannostatin A; marimastat;
masoprocol; maspin; matrilysin inhibitors; matrix metalloproteinase
inhibitors; menogaril; merbarone; meterelin; methioninase;
metoclopramide; MIF inhibitor; mifepristone; miltefosine;
mirimostim; mismatched double stranded RNA; mitoguazone;
mitolactol; mitomycin analogues; mitonafide; mitotoxin fibroblast
growth factor saporin; mitoxantrone; mofarotene; molgramostim;
monoclonal antibody, human chorionic gonadotrophin; monophosphoryl
lipid A+myobacterium cell wall sk; mopidamol; multiple drug
resistance gene inhibitor; multiple tumor suppressor 1 based
therapy; mustard anti-cancer agent; mycaperoxide B; mycobacterial
cell wall extract; myriaporone; N acetyldinaline; N substituted
benzamides; nafarelin; nagrestip; naloxone+pentazocine; napavin;
naphterpin; nartograstim; nedaplatin; nemorubicin; neridronic acid;
neutral endopeptidase; nilutamide; nisamycin; nitric oxide
modulators; nitroxide antioxidant; nitrullyn; 06 benzylguanine;
octreotide; okicenone; oligonucleotides; onapristone; ondansetron;
ondansetron; oracin; oral cytokine inducer; ormaplatin; osaterone;
oxaliplatin; oxaunomycin; paclitaxel; paclitaxel analogues;
paclitaxel derivatives; palauamine; palmitoylrhizoxin; pamidronic
acid; panaxytriol; panomifene; parabactin; pazelliptine;
pegaspargase; peldesine; pentosan polysulfate sodium; pentostatin;
pentrozole; perflubron; perfosfamide; perillyl alcohol;
phenazinomycin; phenylacetate; phosphatase inhibitors; picibanil;
pilocarpine hydrochloride; pirarubicin; piritrexim; placetin A;
placetin B; plasminogen activator inhibitor; platinum complex;
platinum complexes; platinum triamine complex; porfimer sodium;
porfiromycin; prednisone; acridones; prostaglandin J2; proteasome
inhibitors; protein A based immune modulator; protein kinase C
inhibitor; protein kinase C inhibitors, microalgal; protein
tyrosine phosphatase inhibitors; purine nucleoside phosphorylase
inhibitors; purpurins; pyrazoloaeridine; pyridoxylated hemoglobin
polyoxyethylene conjugate; raf antagonists; raltitrexed;
ramosetron; ras farnesyl protein transferase inhibitors; ras
inhibitors; ras GAP inhibitor; retelliptine demethylated; rhenium
Re 186 etidronate; rhizoxin; ribozymes; RH retinamide; rogletimide;
rohitukine; romurtide; roquinimex; rubiginone BI; ruboxyl;
safingol; saintopin; SarCNU; sarcophytol A; sargramostim; Sdi 1
mimetics; semustine; senescence derived inhibitor 1; sense
oligonucleotides; signal transduction inhibitors; signal
transduction modulators; single chain antigen binding protein;
sizofiran; sobuzoxane; sodium borocaptate; sodium phenylacetate;
solverol; somatomedin binding protein; sonermin; sparfosic acid;
spicamycin D; spiromustine; splenopentin; spongistatin 1;
squalamine; stem cell inhibitor; stem cell division inhibitors;
stipiamide; stromelysin inhibitors; sulfinosine; superactive
vasoactive intestinal peptide antagonist; suradista; suramin;
swainsonine; synthetic glycosaminoglycans; tallimustine; tamoxifen
methiodide; tauromustine; tazarotene; tecogalan sodium; tegafur;
tellurapyrylium; telomerase inhibitors; temoporfin; temozolomide;
teniposide; tetrachlorodecaoxide; tetrazomine; thaliblastine;
thiocoraline; thrombopoietin; thrombopoietin mimetic; thymalfasin;
thymopoietin receptor agonist; thymotrinan; thyroid stimulating
hormone; tin ethyl etiopurpurin; tirapazamine; titanocene
bichloride; topsentin; toremifene; totipotent stem cell factor;
translation inhibitors; tretinoin; triacetyluridine; triciribine;
trimetrexate; triptorelin; tropisetron; turosteride; tyrosine
kinase inhibitors; tyrphostins; UBC inhibitors; ubenimex;
urogenital sinus derived growth inhibitory factor; urokinase
receptor antagonists; vapreotide; variolin B; vector system,
erythrocyte gene therapy; velaresol; veramine; verdins;
verteporfin; vinorelbine; vinxaltine; vitaxin; vorozole;
zanoterone; zeniplatin; zilascorb; and zinostatin stimalamer.
It is a further aspect of the invention that the present HDACIs can
be administered in conjunction with chemical agents that are
understood to mimic the effects of radiotherapy and/or that
function by direct contact with DNA. Preferred agents for use in
combination with the present HDACIs for treating cancer include,
but are not limited to cis-diamminedichloro platinum (II)
(cisplatin), doxorubicin, 5-fluorouracil, taxol, and topoisomerase
inhibitors such as etoposide, teniposide, irinotecan and
topotecan.
Additionally, the invention provides methods of treatment of cancer
using the present HDACIs as an alternative to chemotherapy alone or
radiotherapy alone where the chemotherapy or the radiotherapy has
proven or can prove too toxic, e.g., results in unacceptable or
unbearable side effects, for the subject being treated. The
individual being treated can, optionally, be treated with another
anticancer treatment modality such as chemotherapy, surgery, or
immunotherapy, depending on which treatment is found to be
acceptable or bearable.
The present HDACIs can also be used in an in vitro or ex vivo
fashion, such as for the treatment of certain cancers, including,
but not limited to leukemias and lymphomas, such treatment
involving autologous stem cell transplants. This can involve a
multi-step process in which the subject's autologous hematopoietic
stem cells are harvested and purged of all cancer cells, the
subject is then administered an amount of a present HDACI effective
to eradicate the subject's remaining bone-marrow cell population,
then the stem cell graft is infused back into the subject.
Supportive care then is provided while bone marrow function is
restored and the subject recovers.
The present methods for treating cancer can further comprise the
administration of a present HDACI and an additional therapeutic
agent or pharmaceutically acceptable salts or hydrates thereof. In
one embodiment, a composition comprising a present HDACI is
administered concurrently with the administration of one or more
additional therapeutic agent(s), which may be part of the same
composition or in a different composition from that comprising the
present HDACI. In another embodiment, a present HDACI is
administered prior to or subsequent to administration of another
therapeutic agent(s).
In the present methods for treating cancer the other therapeutic
agent may be an antiemetic agent. Suitable antiemetic agents
include, but are not limited to, metoclopromide, domperidone,
prochlorperazine, promethazine, chlorpromazine, trimethobenzamide,
ondansetron, granisetron, hydroxyzine, acethylleucine
monoethanolamine, alizapride, azasetron, benzquinamide,
bietanautine, bromopride, buclizine, clebopride, cyclizine,
dimenhydrinate, diphenidol, dolasetron, meclizine, methallatal,
metopimazine, nabilone, oxyperndyl, pipamazine, scopolamine,
sulpiride, tetrahydrocannabinols, thiethylperazine,
thioproperazine, and tropisetron.
In a preferred embodiment, the antiemetic agent is granisetron or
ondansetron. In another embodiment, the other therapeutic agent may
be an hematopoietic colony stimulating factor. Suitable
hematopoietic colony stimulating factors include, but are not
limited to, filgrastim, sargramostim, molgramostim, and epoietin
alfa.
In still another embodiment, the other therapeutic agent may be an
opioid or non-opioid analgesic agent. Suitable opioid analgesic
agents include, but are not limited to, morphine, heroin,
hydromorphone, hydrocodone, oxymorphone, oxycodone, metopon,
apomorphine, normorphine, etorphine, buprenorphine, meperidine,
lopermide, anileridine, ethoheptazine, piminidine, betaprodine,
diphenoxylate, fentanil, sufentanil, alfentanil, remifentanil,
levorphanol, dextromethorphan, phenazocine, pentazocine,
cyclazocine, methadone, isomethadone, and propoxyphene. Suitable
non-opioid analgesic agents include, but are not limited to,
aspirin, celecoxib, rofecoxib, diclofinac, diflusinal, etodolac,
fenoprofen, flurbiprofen, ibuprofen, ketoprofen, indomethacin,
ketorolac, meclofenamate, mefanamic acid, nabumetone, naproxen,
piroxicam, and sulindac.
In still another embodiment, the other therapeutic agent may be an
anxiolytic agent. Suitable anxiolytic agents include, but are not
limited to, buspirene, and benzodiazepines such as diazepam,
lorazepam, oxazapam, chlorazepate, clonazepam, chlordiazepoxide and
alprazolam.
In addition to treating cancers and sensitizing a cancer cell to
the cytotoxic effects of radiotherapy and chemotherapy, the present
HDACIs are used in methods of treating diseases, conditions, and
injuries to the central nervous system, such as neurological
diseases, neurodegenerative disorders, and traumatic brain injuries
(TBIs). In preferred embodiments, a present HDACI is capable of
crossing the blood brain barrier to inhibit HDAC in the brain of
the individual.
It has been shown that HDAC6 inhibition protects against neuronal
degeneration and stimulates neurite outgrowth in dorsal root
ganglion neurons, therefore indicating methods of treating CNS
diseases. Accordingly, present HDACI compounds were examined in a
model of oxidative stress induced by homocysteic acid (HCA). This
model leads to depletion of glutathione, the major intracellular
antioxidant. HDAC6 inhibition rescues neuronal death in this model,
possibly by causing hyperacetylation of peroxiredoxins. Previous
work reported that nonselective, hydroxamic acid HDACIs displayed
considerable toxicity to the primary cortical neurons. (A. P.
Kozikowski et al., J. Med. Chem. 2007, 50, 3054-61.)
In the HCA-induced neurodegeneration assays, TSA was moderately
neuroprotective at 0.5 .mu.M, although protection declined at
higher concentrations due to dose-dependant neurotoxicity (FIG. 1).
Compounds of the present invention displayed dose-dependent
protection against HCA-induced neuronal cell death starting at 10
.mu.M with near complete protection at 10 .mu.M (FIG. 2). This
compares well with published results showing that Tubacin induces
.alpha.-tubulin acetylation at 5 .mu.M and protects prostate cancer
(LNCaP) cells from hydrogen peroxide-induced death at 8 .mu.M via
peroxiredoxin acetylation. (R. B. Parmigiani et al., Proc. Natl.
Acad. Sci. USA 2008, 105, 9633-8.) Importantly, when tested at all
of the concentrations shown, the present HDACI compounds exhibited
no toxicity, indicating that neurotoxicity is likely a product of
class I HDAC inhibition, and not a property inherent to hydroxamic
acids. These results demonstrate that HDAC6 inhibition provides a
method for treating neurodegenerative conditions.
FIGS. 1 and 2 contain neuroprotection bar graphs of the HCA
oxidative stress test assay. Neurons were treated with TSA (FIG. 1)
or a present HDACI compound (FIG. 2), alone or with the addition of
HCA (homocysteic acid).
The data summarized in FIGS. 1 and 2 was obtained according to the
following neuroprotective assay. Primary cortical neuron cultures
were obtained from the cerebral cortex of fetal Sprague-Dawley rats
(embryonic day 17). All experiments were initiated 24 hours after
plating. Under these conditions, the cells are not susceptible to
glutamate-mediated excitotoxicity. For cytotoxicity studies, cells
were rinsed with warm PBS, then placed in minimum essential medium
(Invitrogen) containing 5.5 g/liter glucose, 10% fetal calf serum,
2 mM L-glutamine, and 100 .mu.M cystine. Oxidative stress was
induced by the addition of the glutamate analog homocysteate (HCA;
5 mM) to the media. HCA was diluted from 100-fold concentrated
solutions that were adjusted to pH 7.5. In combination with HCA,
neurons were treated with either TSA or a present HDACI compound at
the indicated concentrations. Viability was assessed after 24 hours
by the MTT assay
(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)
method.
The present HDACI compounds also provide a therapeutic benefit in
models of peripheral neuropathies, such as CMT. HDAC6 inhibitors
have been found to cross the blood nerve barrier and rescue the
phenotype observed in transgenic mice exhibiting symptoms of distal
hereditary motor neuropathy. Administration of HDAC6 inhibitors to
symptomatic mice increased acetylated .alpha.-tubulin levels,
restored proper mitochondrial motility and axonal transport, and
increased muscle re-innervation. Other peripheral neuropathies
include, but are not limited to, giant axonal neuropathy and
various forms of mononeuropathies, polyneuropathies, autonomic
neuropathies, and neuritis.
The present HDACI compounds also ameliorate associative memory loss
following A.beta. elevation. In this test, mice were infused with
A.beta.42 via cannulas implanted into dorsal hippocampus 15 minutes
prior to training. The test compounds are dosed ip (25 mg/kg) 2
hours before training. Fear learning was assessed 24 hours
later.
Contextual fear conditioning performed 24 hours after training
shows a reduction of freezing in A.beta.-infused mice compared to
vehicle-infused mice. Treatment with a present compound ameliorates
deficit in freezing responses in A.beta.-infused mice, and has no
effect in vehicle-infused mice. A test compound alone does not
affect the memory performance of the mice. In addition, treatment
had no effects on motor, sensorial, or motivational skills assessed
using the visible platform test in which the compounds are injected
twice a day for two days. During these experiments, no signs of
overt toxicity, including changes in food and liquid intake, weight
loss, or changes in locomotion and exploratory behavior, are
observed.
These results demonstrate that the HDACIs of the present invention
are beneficial against impairment of associative memory following
A.beta. elevation.
The present HDACIs therefore are useful for treating a neurological
disease by administration of amounts of a present HDACI effective
to treat the neurological disease or by administration of a
pharmaceutical composition comprising amounts of a present HDACI
effective to treat the neurological disease. The neurological
diseases that can be treated include, but are not limited to,
Huntington's disease, lupus, schizophrenia, multiple sclerosis,
muscular dystrophy, dentatorubralpallidoluysian atrophy (DRRLA),
spinal and bulbar muscular atrophy (SBMA), and fine spinocerebellar
ataxias (SCA1, SCA2, SCA3/MJD (Machado-Joseph Disease), SCA6, and
SCA7), drug-induced movement disorders, Creutzfeldt-Jakob disease,
amyotrophic lateral sclerosis, Pick's disease, Alzheimer's disease,
Lewy body dementia, cortico basal degeneration, dystonia,
myoclonus, Tourette's syndrome, tremor, chorea, restless leg
syndrome, Parkinson's disease, Parkinsonian syndromes, anxiety,
depression, psychosis, manic depression, Friedreich's ataxia,
Fragile X syndrome, spinal muscular dystrophy, Rett syndrome,
Rubinstein-Taybi syndrome, Wilson's disease, multi-infarct state,
CMT, GAN and other peripheral neuropathies.
In a preferred embodiment, the neurological disease treated is
Huntington's disease, Parkinson's disease, Alzheimer's disease,
spinal muscular atrophy, lupus, or schizophrenia.
A present HDACI also can be used with a second therapeutic agent in
methods of treating conditions, diseases, and injuries to the CNS.
Such second therapeutic agents are those drugs known in the art to
treat a particular condition, diseases, or injury, for example, but
not limited to, lithium in the treatment of mood disorders,
estradiol benzoate, and nicotinamide in the treatment of
Huntington's disease.
The present HDACIs also are useful in the treatment of TBIs.
Traumatic brain injury (TBI) is a serious and complex injury that
occurs in approximately 1.4 million people each year in the United
States. TBI is associated with a broad spectrum of symptoms and
disabilities, including a risk factor for developing
neurodegenerative disorders, such as Alzheimer's disease.
TBI produces a number of pathologies including axonal injury, cell
death, contusions, and inflammation. The inflammatory cascade is
characterized by proinflammatory cytokines and activation of
microglia which can exacerbate other pathologies. Although the role
of inflammation in TBI is well established, no efficacious
anti-inflammatory therapies are currently available for the
treatment of TBI.
Several known HDAC inhibitors have been found to be protective in
different cellular and animal models of acute and chronic
neurodegenerative injury and disease, for example, Alzheimer's
disease, ischemic stroke, multiple sclerosis (MS), Huntington's
disease (HD), amyotrophic lateral sclerosis (ALS), spinal muscular
atrophy (SMA), and spinal and bulbar muscular atrophy (SBMA). A
recent study in experimental pediatric TBI reported a decrease in
hippocampal CA3 histone H3 acetylation lasting hours to days after
injury. These changes were attributed to documented upstream
excitotoxic and stress cascades associated with TBI. HDACIs also
have been reported to have anti-inflammatory actions acting through
acetylation of non-histone proteins. The HDAC6 selective inhibitor,
4-dimethylamino-N-[5-(2-mercaptoacetylamino)pentyl]benzamide
(DMA-PB), was found to be able to increase histone H3 acetylation
and reduce microglia inflammatory response following traumatic
brain injury in rats, which demonstrates the utility of HDACIs as
therapeutics for inhibiting neuroinflammation associated with
TBI.
The present HDACIs therefore also are useful in the treatment of
inflammation and strokes, and in the treatment of autism and autism
spectrum disorders. The present HDACIs further can be used to treat
parasitic infections, (e.g., malaria, toxoplasmosis,
trypanosomiasis, helminthiasis, protozoal infections (see Andrews
et al. Int. J. Parasitol. 2000, 30(6), 761-768).
In certain embodiments, the compound of the invention can be used
to treat malaria. A present HDACI can be co-administered with an
antimalarial compound selected from the group consisting of aryl
amino alcohols, cinchona alkaloids, 4-aminoquinolines, type 1 or
type 2 folate synthesis inhibitors, 8-aminoquinolines,
antimicrobials, peroxides, naphthoquinones, and iron chelating
agents. The antimalarial compound can be, but is not limited to,
quinine, quinidine, mefloquine, halfantrine, chloroquine,
amodiaquine, proguanil, chloroproquanil, pyrimethamine, primaquine,
8-[(4-amino-1-methylbutyl)amino]-2,6-dimethoxy-4-methyl-5-[(3-trifluorome-
thyl)phenoxy]quinoline succinate (WR238, 605), tetracycline,
doxycycline, clindamycin, azithromycin, fluoroquinolones,
artemether, areether, artesunate, artelinic acid, atovaquone, and
deferrioxamine. In a preferred embodiment, the antimalarial
compound is chloroquine.
The present HDACIs also can be used as imaging agents. In
particular, by providing a radiolabeled, isotopically labeled, or
fluorescently-labeled HDACI, the labeled compound can image HDACs,
tissues expressing HDACs, and tumors. Labeled HDACIs of the present
invention also can image patients suffering from a cancer, or other
HDAC-mediated diseases, e.g., stroke, by administration of an
effective amount of the labeled compound or a composition
containing the labeled compound. In preferred embodiments, the
labeled HDACI is capable of emitting positron radiation and is
suitable for use in positron emission tomography (PET). Typically,
a labeled HDACI of the present invention is used to identify areas
of tissues or targets that express high concentrations of HDACs.
The extent of accumulation of labeled HDACI can be quantified using
known methods for quantifying radioactive emissions. In addition,
the labeled HDACI can contain a fluorophore or similar reporter
capable of tracking the movement of particular HDAC isoforms or
organelles in vitro.
The present HDACIs useful in the imaging methods contain one or
more radioisotopes capable of emitting one or more forms of
radiation suitable for detection by any standard radiology
equipment, such as PET, SPECT, gamma cameras, MRI, and similar
apparatus. Preferred isotopes including tritium (.sup.3H) and
carbon (.sup.11C). Substituted HDACIs of the present invention also
can contain isotopes of fluorine (.sup.18F) and iodine (.sup.123I)
for imaging methods. Typically, a labeled HDACI of the present
invention contains an alkyl group having a .sup.11C label, i.e., a
.sup.11C-methyl group, or an alkyl group substituted with .sup.18F,
.sup.123I, .sup.125I, .sup.131I, or a combination thereof.
Fluorescently-labeled HDACIs of the present invention also can be
used in the imaging method of the present invention. Such compounds
have an FITC, carbocyamine moiety or other fluorophore which will
allow visualization of the HDAC proteins in vitro.
The labeled HDACIs and methods of use can be in vivo, and
particularly on humans, and for in vitro applications, such as
diagnostic and research applications, using body fluids and cell
samples. Imaging methods using a labeled HDACI of the present
invention are discussed in WO 03/060523, designating the U.S. and
incorporated in its entirety herein. Typically, the method
comprises contacting cells or tissues with a radiolabeled,
isotopically labeled, fluorescently labeled, or tagged (such as
biotin tagged) compound of the invention, and making a
radiographic, fluorescent, or similar type of image depending on
the visualization method employed, i.e., in regard to radiographic
images, a sufficient amount to provide about 1 to about 30 mCi of
the radiolabeled compound.
Preferred imaging methods include the use of labeled HDACIs of the
present invention which are capable of generating at least a 2:1
target to background ratio of radiation intensity, or more
preferably about a 5:1, about 10:1, or about 15:1 ratio of
radiation intensity between target and background.
In preferred methods, the labeled HDACIs of the present invention
are excreted from tissues of the body quickly to prevent prolonged
exposure to the radiation of the radiolabeled compound administered
to the individual. Typically, labeled HDACIs of the present
invention are eliminated from the body in less than about 24 hours.
More preferably, labeled HDACIs are eliminated from the body in
less than about 16 hours, 12 hours, 8 hours, 6 hours, 4 hours, 2
hours, 90 minutes, or 60 minutes. Typically, preferred labeled
HDACIs are eliminated in about 60 to about 120 minutes.
In addition to isotopically labeled and fluorescently labeled
derivatives, the present invention also embodies the use of
derivatives containing tags (such as biotin) for the identification
of biomolecules associated with the HDAC isoforms of interest for
diagnostic, therapeutic or research purposes.
The present HDACIs also are useful in the treatment of autoimmune
diseases and inflammations. Compounds of the present invention are
particularly useful in overcoming graft and transplant rejections
and in treating forms of arthritis.
Despite successes of modern transplant programs, the
nephrotoxicity, cardiovascular disease, diabetes, and
hyperlipidemia associated with current therapeutic regimens, plus
the incidence of post-transplant malignancies and graft loss from
chronic rejection, drive efforts to achieve long-term allograft
function in association with minimal immunosuppression Likewise,
the incidence of inflammatory bowel disease (IBD), including
Crohn's disease and ulcerative colitis, is increasing. Animal
studies have shown that T regulatory cells (Tregs) expressing the
forkhead transcription family member, Foxp3, are key to limiting
autoreactive and alloreactive immunity. Moreover, after their
induction by costimulation blockade, immunosuppression, or other
strategies, Tregs may be adoptively transferred to naive hosts to
achieve beneficial therapeutic effects. However, attempts to
develop sufficient Tregs that maintain their suppressive functions
post-transfer in clinical trials have failed. Murine studies show
that HDACIs limit immune responses, at least in significant part,
by increasing Treg suppressive functions, (R. Tao et al., Nat Med,
13, 1299-1307, (2007)), and that selective targeting of HDAC6 is
especially efficacious in this regard.
With organ transplantation, rejection begins to develop in the days
immediately post-transplant, such that prevention rather than
treatment of rejection is a paramount consideration. The reverse
applies in autoimmunity, wherein a patient presents with the
disease already causing problems. Accordingly, HDAC6-/- mice
treated for 14 days with low-dose RPM (rapamycin) are assessed for
displaying signs of tolerance induction and resistance to the
development of chronic rejection, a continuing major loss of graft
function long-term in the clinical transplant population. Tolerance
is assessed by testing whether mice with long-surviving allografts
reject a subsequent third-party cardiac graft and accept additional
donor allografts without any immunosuppression, as can occur using
a non-selective HDACI plus RPM. These in vivo sutides are
accompanied by assessment of ELISPOT and MLR activities using
recipient lymphocytes challenged with donor cells. Protection
against chronic rejection is assessed by analysis of host
anti-donor humoral responses and analysis of graft transplant
arteriosclerosis and interstitial fibrosis in long-surviving
allograft recipients.
The importance of HDAC6 targeting is assessed in additional
transplant models seeking readouts of biochemical significance, as
is monitored clinically. Thus, the effects of HDAC6 in targeting in
renal transplant recipients (monitoring BUN, proteinuria) and islet
allografts (monitoring blood glucose levels) are assessed. Renal
transplants are the most common organ transplants performed, and
the kidney performs multiple functions, e.g., regulating acid/base
metabolism, blood pressure, red cell production, such that efficacy
in this model indicates the utility of HDAC6 targeting. Likewise,
islet transplantation is a major unmet need given that clinical
islet allografts are typically lost after the first one or two
years post-transplant. Having a safe and non-toxic means to extend
islet survival without maintenance CNI therapy would be an
important advance. Transplant studies also are strengthened by use
of mice with floxed HDAC6. Using existing Foxp3-Cre mice, for
example, the effects of deletion of HDAC6 just in Tregs is tested.
This approach can be extended to targeting of HDAC6 in T cells
(CD4-Cre) and dendritic cells (CD11c-Cre), for example. Using
tamoxifen-regulated Cre, the importance of HDAC6 in induction vs.
maintenance of transplants (with implications for short-term vs.
maintenance HDAC6I therapy) is assessed by administering tamoxifen
and inducing HDAC6 deletion at varying periods post-transplant.
Studies of autoimmunity also are undertaken. In this case,
interruption of existing disease is especially important and HDAC6
targeting can be efficacious without any requirement for additional
therapy (in contrast to a need for brief low-dose RPM in the very
aggressive, fully MHC-mismatched transplant models). Studies in
mice with colitis indicated that HDAC6-/- Tregs were more effective
than WT Tregs in regulating disease, and tubacin was able to rescue
mice if treatment was begun once colitis had developed. These
studies are extended by assessing whether deletion of HDAC6 in
Tregs (Foxp3/Cre) vs. T cells (CD4=Cre) vs. DC (CD11c-Cre)
differentially affect the development and severity of colitis.
Similarly, control of colitis is assessed by inducing HDAC6
deletion at varying intervals after the onset of colitis with
tamoxifen-regulated Cre.
The present compounds are envisioned to demonstrate anti-arthritic
efficacy in a collagen-induced arthritis model in DBA1/J mice. In
this test, DBA1/J mice (male, 7-8 weeks) are used, with 8 animals
per group. Systemic arthritis is induced with bovine collagen type
II and CFA, plus an IFA booster injection on day 21. A present
HDACI is dosed at 50 mg/kg and 100 mg/kg on day 28 for 2
consecutive weeks, and the effects determined from the Average
Arthritic Score vs. Days of Treatment data.
Despite efforts to avoid graft rejection through host-donor tissue
type matching, in the majority of transplantation procedures,
immunosuppressive therapy is critical to the viability of the donor
organ in the host. A variety of immunosuppressive agents have been
employed in transplantation procedures, including azathioprine,
methotrexate, cyclophosphamide, FK-506, rapamycin, and
corticosteroids.
The present HDACIs are potent immunosuppressive agents that
suppress humoral immunity and cell-mediated immune reactions, such
as allograft rejection, delayed hypersensitivity, experimental
allergic encephalomyelitis, Freund's adjuvant arthritis and graft
versus host disease. HDACIs of the present invention are useful for
the prophylaxis of organ rejection subsequent to organ
transplantation, for treatment of rheumatoid arthritis, for the
treatment of psoriasis, and for the treatment of other autoimmune
diseases, such as type I diabetes, Crohn's disease, and lupus.
A therapeutically effective amount of a present HDACI can be used
for immunosuppression including, for example, to prevent organ
rejection or graft vs. host disease, and to treat diseases and
conditions, in particular, autoimmune and inflammatory diseases and
conditions. Examples of autoimmune and inflammatory diseases
include, but are not limited to, Hashimoto's thyroiditis,
pernicious anemia, Addison's disease, psoriasis, diabetes,
rheumatoid arthritis, systemic lupus erythematosus,
dermatomyositis, Sjogren's syndrome, dermatomyositis, lupus
erythematosus, multiple sclerosis, myasthenia gravis, Reiter's
syndrome, arthritis (rheumatoid arthritis, arthritis chronic
progrediente, and arthritis deformans) and rheumatic diseases,
autoimmune hematological disorder (hemolytic anaemia, aplastic
anaemia, pure red cell anaemia and idiopathic thrombocytopaenia),
systemic lupus erythematosus, polychondritis, sclerodoma, Wegener
granulamatosis, dermatomyositis, chronic active hepatitis,
psoriasis, Steven-Johnson syndrome, idiopathic sprue, autoimmune
inflammatory bowel disease (ulcerative colitis and Crohn's disease)
endocrine opthalmopathy, Graves disease, sarcoidosis, primary
biliary cirrhosis, juvenile diabetes (diabetes mellitus type I),
uveitis (anterior and posterior), keratoconjunctivitis sicca and
vernal keratoconjunctivitis, interstitial lung fibrosis, psoriatic
arthritis, and glomerulonephritis.
A present HDACI can be used alone, or in conjunction with a second
therapeutic agent known to be useful in the treatment of autoimmune
diseases, inflammations, transplants, and grafts, such as
cyclosporin, rapamycin, methotrexate, cyclophosphamide,
azathioprine, corticosteroids, and similar agents known to persons
skilled in the art.
Additional diseases and conditions mediated by HDACs, and
particularly HDAC6, include, but are not limited to asthma, cardiac
hypertrophy, giant axonal neuropathy, mononeuropathy, mononeuritis,
polyneuropathy, autonomic neuropathy, neuritis in general, and
neuropathy in general. These disease and conditions also can be
treated by a method of the present invention.
In the present method, a therapeutically effective amount of one or
more HDACI of the present invention, typically formulated in
accordance with pharmaceutical practice, is administered to a human
being in need thereof. Whether such a treatment is indicated
depends on the individual case and is subject to medical assessment
(diagnosis) that takes into consideration signs, symptoms, and/or
malfunctions that are present, the risks of developing particular
signs, symptoms and/or malfunctions, and other factors.
A present HDACI can be administered by any suitable route, for
example by oral, buccal, inhalation, topical, sublingual, rectal,
vaginal, intracisternal or intrathecal through lumbar puncture,
transurethral, nasal, percutaneous, i.e., transdermal, or
parenteral (including intravenous, intramuscular, subcutaneous,
intracoronary, intradermal, intramammary, intraperitoneal,
intraarticular, intrathecal, retrobulbar, intrapulmonary injection
and/or surgical implantation at a particular site) administration.
Parenteral administration can be accomplished using a needle and
syringe or using a high pressure technique.
Pharmaceutical compositions include those wherein a present HDACI
is present in a sufficient amount to be administered in an
effective amount to achieve its intended purpose. The exact
formulation, route of administration, and dosage is determined by
an individual physician in view of the diagnosed condition or
disease. Dosage amount and interval can be adjusted individually to
provide levels of a present HDACI that is sufficient to maintain
therapeutic effects.
Toxicity and therapeutic efficacy of the present HDACI compounds
can be determined by standard pharmaceutical procedures in cell
cultures or experimental animals, e.g., for determining the
LD.sub.50 (the dose lethal to 50% of the population) and the
ED.sub.50 (the dose therapeutically effective in 50% of the
population). The dose ratio between toxic and therapeutic effects
is the therapeutic index, which is expressed as the ratio between
LD.sub.50 and ED.sub.50. Compounds that exhibit high therapeutic
indices are preferred. The data obtained from such procedures can
be used in formulating a dosage range for use in humans. The dosage
preferably lies within a range of circulating compound
concentrations that include the ED.sub.50 with little or no
toxicity. The dosage can vary within this range depending upon the
dosage form employed, and the route of administration utilized.
Determination of a therapeutically effective amount is well within
the capability of those skilled in the art, especially in light of
the detailed disclosure provided herein.
A therapeutically effective amount of a present HDACI required for
use in therapy varies with the nature of the condition being
treated, the length of time that activity is desired, and the age
and the condition of the patient, and ultimately is determined by
the attendant physician. Dosage amounts and intervals can be
adjusted individually to provide plasma levels of the HDACI that
are sufficient to maintain the desired therapeutic effects. The
desired dose conveniently can be administered in a single dose, or
as multiple doses administered at appropriate intervals, for
example as one, two, three, four or more subdoses per day. Multiple
doses often are desired, or required. For example, a present HDACI
can be administered at a frequency of: four doses delivered as one
dose per day at four-day intervals (q4d.times.4); four doses
delivered as one dose per day at three-day intervals (q3d.times.4);
one dose delivered per day at five-day intervals (qd.times.5); one
dose per week for three weeks (qwk3); five daily doses, with two
days rest, and another five daily doses (5/2/5); or, any dose
regimen determined to be appropriate for the circumstance.
The dosage of a composition containing a present HDACI, or a
composition containing the same, can be from about 1 ng/kg to about
200 mg/kg, about 1 .mu.g/kg to about 100 mg/kg, or about 1 mg/kg to
about 50 mg/kg of body weight. The dosage of a composition may be
at any dosage including, but not limited to, about 1 .mu.g/kg, 10
.mu.g/kg, 25 .mu.g/kg, 50 .mu.g/kg, 75 .mu.g/kg, 100 .mu.g/kg, 125
.mu.g/kg, 150 .mu.g/kg, 175 .mu.g/kg, 200 .mu.g/kg, 225 .mu.g/kg,
250 .mu.g/kg, 275 .mu.g/kg, 300 .mu.g/kg, 325 .mu.g/kg, 350
.mu.g/kg, 375 .mu.g/kg, 400 .mu.g/kg, 425 .mu.g/kg, 450 .mu.g/kg,
475 .mu.g/kg, 500 .mu.g/kg, 525 .mu.g/kg, 550 .mu.g/kg, 575
.mu.g/kg, 600 .mu.g/kg, 625 .mu.g/kg, 650 .mu.g/kg, 675 .mu.g/kg,
700 .mu.g/kg, 725 .mu.g/kg, 750 .mu.g/kg, 775 .mu.g/kg, 800
.mu.g/kg, 825 .mu.g/kg, 850 .mu.g/kg, 875 .mu.g/kg, 900 .mu.g/kg,
925 .mu.g/kg, 950 .mu.g/kg, 975 .mu.g/kg, 1 mg/kg, 5 mg/kg, 10
mg/kg, 15 mg/kg, 20 mg/kg, 25 mg/kg, 30 mg/kg, 35 mg/kg, 40 mg/kg,
45 mg/kg, 50 mg/kg, 60 mg/kg, 70 mg/kg, 80 mg/kg, 90 mg/kg, 100
mg/kg, 125 mg/kg, 150 mg/kg, 175 mg/kg, or 200 mg/kg. The above
dosages are exemplary of the average case, but there can be
individual instances in which higher or lower dosages are merited,
and such are within the scope of this invention. In practice, the
physician determines the actual dosing regimen that is most
suitable for an individual patient, which can vary with the age,
weight, and response of the particular patient.
A present HDACI used in a method of the present invention typically
is administered in an amount of about 0.005 to about 500 milligrams
per dose, about 0.05 to about 250 milligrams per dose, or about 0.5
to about 100 milligrams per dose. For example, a present HDACI can
be administered, per dose, in an amount of about 0.005, 0.05, 0.5,
5, 10, 20, 30, 40, 50, 100, 150, 200, 250, 300, 350, 400, 450, or
500 milligrams, including all doses between 0.005 and 500
milligrams.
The HDACIs of the present invention typically are administered in
admixture with a pharmaceutical carrier selected with regard to the
intended route of administration and standard pharmaceutical
practice. Pharmaceutical compositions for use in accordance with
the present invention are formulated in a conventional manner using
one or more physiologically acceptable carriers comprising
excipients and auxiliaries that facilitate processing of the
present HDACIs.
The term "carrier" refers to a diluent, adjuvant, or excipient,
with which a present HDACI is administered. Such pharmaceutical
carriers can be liquids, such as water and oils, including those of
petroleum, animal, vegetable or synthetic origin, such as peanut
oil, soybean oil, mineral oil, sesame oil, and the like. The
carriers can be saline, gum acacia, gelatin, starch paste, talc,
keratin, colloidal silica, urea, and the like. In addition,
auxiliary, stabilizing, thickening, lubricating and coloring agents
can be used. The pharmaceutically acceptable carriers are sterile.
Water is a preferred carrier when a present HDACI is administered
intravenously. Saline solutions and aqueous dextrose and glycerol
solutions can also be employed as liquid carriers, particularly for
injectable solutions. Suitable pharmaceutical carriers also include
excipients such as starch, glucose, lactose, sucrose, gelatin,
malt, rice, flour, chalk, silica gel, sodium stearate, glycerol
monostearate, talc, sodium chloride, dried skim milk, glycerol,
propylene, glycol, water, ethanol, and the like. The present
compositions, if desired, can also contain minor amounts of wetting
or emulsifying agents, or pH buffering agents.
These pharmaceutical compositions can be manufactured, for example,
by conventional mixing, dissolving, granulating, dragee-making,
emulsifying, encapsulating, entrapping, or lyophilizing processes.
Proper formulation is dependent upon the route of administration
chosen. When a therapeutically effective amount of a present HDACI
is administered orally, the composition typically is in the form of
a tablet, capsule, powder, solution, or elixir. When administered
in tablet form, the composition additionally can contain a solid
carrier, such as a gelatin or an adjuvant. The tablet, capsule, and
powder contain about 0.01% to about 95%, and preferably from about
1% to about 50%, of a present HDACI. When administered in liquid
form, a liquid carrier, such as water, petroleum, or oils of animal
or plant origin, can be added. The liquid form of the composition
can further contain physiological saline solution, dextrose or
other saccharide solutions, or glycols. When administered in liquid
form, the composition contains about 0.1% to about 90%, and
preferably about 1% to about 50%, by weight, of a present
compound.
When a therapeutically effective amount of a present HDACI is
administered by intravenous, cutaneous, or subcutaneous injection,
the composition is in the form of a pyrogen-free, parenterally
acceptable aqueous solution. The preparation of such parenterally
acceptable solutions, having due regard to pH, isotonicity,
stability, and the like, is within the skill in the art. A
preferred composition for intravenous, cutaneous, or subcutaneous
injection typically contains an isotonic vehicle. A present HDACI
can be infused with other fluids over a 10-30 minute span or over
several hours.
The present HDACIs can be readily combined with pharmaceutically
acceptable carriers well-known in the art. Such carriers enable the
active agents to be formulated as tablets, pills, dragees,
capsules, liquids, gels, syrups, slurries, suspensions and the
like, for oral ingestion by a patient to be treated. Pharmaceutical
preparations for oral use can be obtained by adding a present HDACI
to a solid excipient, optionally grinding the resulting mixture,
and processing the mixture of granules, after adding suitable
auxiliaries, if desired, to obtain tablets or dragee cores.
Suitable excipients include, for example, fillers and cellulose
preparations. If desired, disintegrating agents can be added.
A present HDACI can be formulated for parenteral administration by
injection, e.g., by bolus injection or continuous infusion.
Formulations for injection can be presented in unit dosage form,
e.g., in ampules or in multidose containers, with an added
preservative. The compositions can take such forms as suspensions,
solutions, or emulsions in oily or aqueous vehicles, and can
contain formulatory agents such as suspending, stabilizing, and/or
dispersing agents.
Pharmaceutical compositions for parenteral administration include
aqueous solutions of the active agent in water-soluble form.
Additionally, suspensions of a present HDACI can be prepared as
appropriate oily injection suspensions. Suitable lipophilic
solvents or vehicles include fatty oils or synthetic fatty acid
esters. Aqueous injection suspensions can contain substances which
increase the viscosity of the suspension. Optionally, the
suspension also can contain suitable stabilizers or agents that
increase the solubility of the compounds and allow for the
preparation of highly concentrated solutions. Alternatively, a
present composition can be in powder form for constitution with a
suitable vehicle, e.g., sterile pyrogen-free water, before use.
A present HDACI also can be formulated in rectal compositions, such
as suppositories or retention enemas, e.g., containing conventional
suppository bases. In addition to the formulations described
previously, a present HDACI also can be formulated as a depot
preparation. Such long-acting formulations can be administered by
implantation (for example, subcutaneously or intramuscularly) or by
intramuscular injection. Thus, for example, a present HDACI can be
formulated with suitable polymeric or hydrophobic materials (for
example, as an emulsion in an acceptable oil) or ion exchange
resins.
In particular, a present HDACI can be administered orally,
buccally, or sublingually in the form of tablets containing
excipients, such as starch or lactose, or in capsules or ovules,
either alone or in admixture with excipients, or in the form of
elixirs or suspensions containing flavoring or coloring agents.
Such liquid preparations can be prepared with pharmaceutically
acceptable additives, such as suspending agents. The present HDACIs
also can be injected parenterally, for example, intravenously,
intramuscularly, subcutaneously, or intracoronarily. For parenteral
administration, the present HDACIs are best used in the form of a
sterile aqueous solution which can contain other substances, for
example, salts or monosaccharides, such as mannitol or glucose, to
make the solution isotonic with blood.
As an additional embodiment, the present invention includes kits
which comprise one or more compounds or compositions packaged in a
manner that facilitates their use to practice methods of the
invention. In one simple embodiment, the kit includes a compound or
composition described herein as useful for practice of a method
(e.g., a composition comprising a present HDACI and an optional
second therapeutic agent), packaged in a container, such as a
sealed bottle or vessel, with a label affixed to the container or
included in the kit that describes use of the compound or
composition to practice the method of the invention. Preferably,
the compound or composition is packaged in a unit dosage form. The
kit further can include a device suitable for administering the
composition according to the intended route of administration, for
example, a syringe, drip bag, or patch. In another embodiment, the
present compounds is a lyophilate. In this instance, the kit can
further comprise an additional container which contains a solution
useful for the reconstruction of the lyophilate.
Prior HDACIs possessed properties that hindered their development
as therapeutic agents. In accordance with an important feature of
the present invention, the present HDACIs were synthesized and
evaluated as inhibitors for HDAC. The present compounds demonstrate
an increased HDAC6 potency and selectivity against HDAC1 and HDAC8
with improvements in BEI relative to prior compounds. The improved
properties of the present compounds, particularly the increase in
BEI and reduced potency at HDAC8, indicate that the present
compounds are useful for applications such as, but not limited to,
immunosuppressive and neuroprotective agents. For example,
compounds of the present invention typically have a bonding
affinity (IC.sub.50) to HDAC6 of less than 100 .mu.M, less than 25
.mu.M, less than 10 .mu.M, less than 1 .mu.M, less than 0.5 .mu.M,
and less than 0.2 .mu.M.
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