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WO2007067993A1 - Inhibitors of histone deacetylase for the treatment of disease - Google Patents

Inhibitors of histone deacetylase for the treatment of disease Download PDF

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Publication number
WO2007067993A1
WO2007067993A1 PCT/US2006/061820 US2006061820W WO2007067993A1 WO 2007067993 A1 WO2007067993 A1 WO 2007067993A1 US 2006061820 W US2006061820 W US 2006061820W WO 2007067993 A1 WO2007067993 A1 WO 2007067993A1
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optionally substituted
group
compound
inhibitors
recited
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PCT/US2006/061820
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French (fr)
Inventor
Celine Bonnefous
Joseph E. Payne
Nicholas D. Smith
Timothy Z. Hoffman
Michael Sertic
Paul L. Wash
James W. Malecha
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Kalypsys, Inc.
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Publication of WO2007067993A1 publication Critical patent/WO2007067993A1/en

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    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D209/00—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D209/02—Heterocyclic compounds containing five-membered rings, condensed with other rings, with one nitrogen atom as the only ring hetero atom condensed with one carbocyclic ring
    • C07D209/04—Indoles; Hydrogenated indoles
    • C07D209/08—Indoles; Hydrogenated indoles with only hydrogen atoms or radicals containing only hydrogen and carbon atoms, directly attached to carbon atoms of the hetero ring
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P35/00—Antineoplastic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07C—ACYCLIC OR CARBOCYCLIC COMPOUNDS
    • C07C327/00—Thiocarboxylic acids
    • C07C327/20—Esters of monothiocarboxylic acids
    • C07C327/30—Esters of monothiocarboxylic acids having sulfur atoms of esterified thiocarboxyl groups bound to carbon atoms of hydrocarbon radicals substituted by nitrogen atoms, not being part of nitro or nitroso groups
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D213/00—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members
    • C07D213/02—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members
    • C07D213/04—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D213/60—Heterocyclic compounds containing six-membered rings, not condensed with other rings, with one nitrogen atom as the only ring hetero atom and three or more double bonds between ring members or between ring members and non-ring members having three double bonds between ring members or between ring members and non-ring members having no bond between the ring nitrogen atom and a non-ring member or having only hydrogen or carbon atoms directly attached to the ring nitrogen atom with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D213/72—Nitrogen atoms
    • C07D213/76—Nitrogen atoms to which a second hetero atom is attached
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D231/00—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings
    • C07D231/02—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings not condensed with other rings
    • C07D231/10—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members
    • C07D231/12—Heterocyclic compounds containing 1,2-diazole or hydrogenated 1,2-diazole rings not condensed with other rings having two or three double bonds between ring members or between ring members and non-ring members with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to ring carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D235/00—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, condensed with other rings
    • C07D235/02—Heterocyclic compounds containing 1,3-diazole or hydrogenated 1,3-diazole rings, condensed with other rings condensed with carbocyclic rings or ring systems
    • C07D235/04—Benzimidazoles; Hydrogenated benzimidazoles
    • C07D235/06—Benzimidazoles; Hydrogenated benzimidazoles with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached in position 2
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D239/00—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings
    • C07D239/02—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings
    • C07D239/24—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members
    • C07D239/26—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to ring carbon atoms
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D239/00—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings
    • C07D239/02—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings
    • C07D239/24—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members
    • C07D239/28—Heterocyclic compounds containing 1,3-diazine or hydrogenated 1,3-diazine rings not condensed with other rings having three or more double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, directly attached to ring carbon atoms
    • C07D239/32—One oxygen, sulfur or nitrogen atom
    • C07D239/42—One nitrogen atom
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D271/00—Heterocyclic compounds containing five-membered rings having two nitrogen atoms and one oxygen atom as the only ring hetero atoms
    • C07D271/12—Heterocyclic compounds containing five-membered rings having two nitrogen atoms and one oxygen atom as the only ring hetero atoms condensed with carbocyclic rings or ring systems
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D277/00—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings
    • C07D277/60—Heterocyclic compounds containing 1,3-thiazole or hydrogenated 1,3-thiazole rings condensed with carbocyclic rings or ring systems
    • C07D277/62—Benzothiazoles
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
    • C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
    • C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
    • C07D311/06—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 2
    • C07D311/08—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 2 not hydrogenated in the hetero ring
    • C07D311/14—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 2 not hydrogenated in the hetero ring substituted in position 6 and unsubstituted in position 7
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
    • C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
    • C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
    • C07D311/58—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring other than with oxygen or sulphur atoms in position 2 or 4
    • C07D311/70—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring other than with oxygen or sulphur atoms in position 2 or 4 with two hydrocarbon radicals attached in position 2 and elements other than carbon and hydrogen in position 6
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D401/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom
    • C07D401/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings
    • C07D401/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, at least one ring being a six-membered ring with only one nitrogen atom containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

  • Histone proteins organize DNA into nucleosomes, which are regular repeating structures of chromatin. The acetylation status of histones alters chromatin structure, which, in turn, is involved in gene expression.
  • Two classes of enzymes can affect the acetylation of histones - histone acetyltransferases (HATs) and histone deacetylases (HDACs).
  • HATs histone acetyltransferases
  • HDACs histone deacetylases
  • a number of HDAC inhibitors have been characterized.
  • One of the potent inhibitors of HDAC is (SAHA), ahydroxamic acid-based compound. It is also known as vorinostat or ZOLINZA(TM), which is currently in clinical trials.
  • Histone proteins organize DNA into nucleosomes, which are regular repeating structures of chromatin. The acetylation status of histones alters chromatin structure, which, in turn, is involved in gene expression.
  • Two classes of enzymes can affect the acetylation of histones - histone acetyltransferases (HATs) and histone deacetylases (HDACs).
  • HATs histone acetyltransferases
  • HDACs histone deacetylases
  • Gi is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
  • G 2 is selected from the group consisting of an N-sulfonamide moiety having structure (II), an S-sulfonamide moiety having structure (III), an amide of the form -NR 3 C(O)- , and an amide of the form -C(O)NR 3 -:
  • G 3 is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
  • Ri and R2 are each independently selected from the group consisting of hydrogen, lower alkyl, halogen andperhaloalkyl, or R 1 and R 2 taken together may form an optionally substituted cycloalkyl or optionally substituted heterocycloalkyl;
  • R 3 and R 4 are each independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, and optionally substituted aryl;
  • T is selected from the group consisting of -(XO n1 NR 7 C(O)(X 2 )D 2 Rs and - (X 1 )mSO 2 (X 2 ) ⁇ 2 Rs;
  • nl 0-5;
  • n2 is 0-5;
  • R7 is selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted heteroalkyl, and optionally substituted lower alkoxy;
  • R 8 is selected from the group consisting of optionally substituted lower alkyl, optionally substituted heteroalkyl, optionally substituted heterocyclyl, optionally substituted heteroaryl and - NR 9 Ri 0 ;
  • R 9 and Rio are each independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted lower alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted cycloaklenyl, optionally substituted fused aryl, optionally substituted fused heteroaryl, optionally substituted fused heterocycloalkyl, and optionally substituted fused cycloalkyl; or R 9 and R 10 , taken together with the nitrogen to which they are attached, form an optionally substituted five-membered or six-membered heteroaromatic or heteroaliphatic ring; and
  • Ge is selected from the group consisting of hydrogen, optionally substituted acyl, optionally substituted aryl, optionally substituted alkyl, optionally substituted heteroaryl, optionally substituted alkylthio, optionally substituted arylthio and optionally substituted heteroarylthio; or G 6 may have the structural formula (IV) thereby forming a homodisulf ⁇ de or heterodisulf ⁇ de dimer of a compound of the present invention
  • G 7 is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
  • G 8 is selected from the group consisting of an N-sulfonamide moiety having structure (V), an S-sulfonamide moiety having structure (VI), an amide of the form -NR 13 C(O)- , and an amide of the form -C(O)NR 14 -:
  • G 9 is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
  • R 11 and R ⁇ are each independently selected from the group consisting of hydrogen, lower alkyl, halogen and perhaloalkyl, or Ru and R ⁇ taken together may form an optionally substituted cycloalkyl or optionally substituted heterocycloalkyl;
  • R 13 and R 14 are each independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, and optionally substituted aryl.
  • the invention provides methods and compositions for treating diseases in mammals using compounds of the invention, including but not limited to, treating cancers, autoimmune diseases, tissue damage, central nervous system disorders, neurodegenerative disorders, fibrosis, bone disorders, polyglutamine-repeat disorders, anemias, thalassemias, inflammatory conditions, cardiovascular conditions, and disorders in which angiogenesis plays a role in pathogenesis.
  • the invention further provides methods of inhibiting the catalytic activity and cellular function of histone deacetylase (HDAC).
  • HDAC histone deacetylase
  • G 6 is selected from the group consisting of optionally substituted acyl and hydrogen.
  • the compound of the invention wherein G 1 is optionally substituted 6 membered aryl or optionally substituted 6 membered heteroaryl.
  • the compound of the invention wherein G 2 is N-sulfonamide. In certain embodiments, the compound of the invention wherein G 3 is phenyl.
  • Gi is pyridinyl
  • T is -CXi) nI NR 7 C(O)R 8 -;
  • nl 0.
  • Gi is phenyl
  • T is -CXOmNR 7 C(O)Rs-;
  • nl 0.
  • the present invention provides compounds capable of inhibiting the catalytic activity of histone deacetylase (HDAC).
  • HDAC histone deacetylase
  • the present invention provides pharmaceutical compositions comprising compounds capable of inhibiting the catalytic activity of histone deacetylase CHDAC).
  • the present invention provides methods and compositions for treating certain diseases or disease states.
  • HDAC histone deacetylase
  • the disease is a hyperproliferative condition of the human or animal body.
  • said hyperproliferative condition is selected from the group consisting of hematologic and nonhematologic cancers.
  • said hematologic cancer is selected from the group consisting of multiple myeloma, leukemias, and lymphomas.
  • said leukemia is selected from the group consisting of acute and chronic leukemias.
  • said acute leukemia is selected from the group consisting of acute lymphocytic leukemia CALL) and acute nonlymphocytic leukemia (ANLL).
  • said chronic leukemia is selected from the group consisting of chronic lymphocytic leukemia (CLL) and chronic myelogenous leukemia (CML).
  • said lymphoma is selected from the group consisting of Hodgkin's lymphoma and non-Hodgkin's lymphoma.
  • said lymophoma is selected from the group consisting of cutaneous t-cell lymphoma (CTCL) and mantle cell lymphoma (MCL).
  • CTCL cutaneous t-cell lymphoma
  • MCL mantle cell lymphoma
  • said hematologic cancer is multiple myeloma (MM).
  • said hematologic cancer is of low, intermediate, or high grade.
  • said MM myeloma
  • nonhematologic cancer is selected from the group consisting of: brain cancer, cancers of the head and neck, lung cancer, breast cancer, cancers of the reproductive system, cancers of the digestive system, pancreatic cancer, and cancers of the urinary system.
  • said cancer of the digestive system is a cancer of the upper digestive tract or colorectal cancer.
  • said cancer of the urinary system is bladder cancer or renal cell carcinoma.
  • said cancer of the reproductive system is prostate cancer.
  • cancers of oral cavity and pharynx include: cancers of oral cavity and pharynx, cancers of the respiratory system, cancers of bones and joints, cancers of soft tissue, skin cancers, cancers of the genital system, cancers of the eye and orbit, cancers of the nervous system, cancers of the lymphatic system, and cancers of the endocrine system.
  • these cancer s may beselected from the group consisting of: cancer of the tongue, mouth, pharynx, or other oral cavity; esophageal cancer, stomach cancer, or cancer of the small intestine; colon cancer or rectal, anal, or anorectal cancer; cancer of the liver, intrahepatic bile duct, gallbladder, pancreas, or other biliary or digestive organs; laryngeal, bronchial, and other cancers of the respiratory organs; heart cancer, melanoma, basal cell carcinoma, squamous cell carcinoma, other non-epithelial skin cancer; uterine or cervical cancer; uterine corpus cancer; ovarian, vulvar, vaginal, or other female genital cancer; prostate, testicular, penile or other male genital cancer; urinary bladder cancer; cancer of the kidney; renal, pelvic, or urethral cancer or other cancer of the genito-urinary organs; thyroid cancer or other
  • cancers which may be treated using the compounds and methods described herein include: adenocarcinoma, angiosarcoma, astrocytoma, acoustic neuroma, anaplastic astrocytoma, basal cell carcinoma, blastoglioma, chondrosarcoma, choriocarcinoma, chordoma, craniopharyngioma, cutaneous melanoma, cystadenocarcinoma, endotheliosarcoma, embryonal carcinoma, ependymoma, Ewing's tumor, epithelial carcinoma, fibrosarcoma, gastric cancer, genitourinary tract cancers, glioblastoma multiforme, hemangioblastoma, hepatocellular carcinoma, hepatoma, Kaposi's sarcoma, large cell carcinoma, leiomyosarcoma, liposarcoma,
  • lymphangiosarcoma lymphangioendotheliosarcoma, medullary thyroid carcinoma
  • medulloblastoma meningioma mesothelioma, myelomas, myxosarcoma neuroblastoma, neurofibrosarcoma, oligodendroglioma, osteogenic sarcoma, epithelial ovarian cancer, papillary carcinoma, papillary adenocarcinomas, parathyroid tumors, pheochromocytoma, pinealoma, plasmacytomas, retinoblastoma, rhabdomyosarcoma, sebaceous gland carcinoma, seminoma, skin cancers, melanoma, small cell lung carcinoma, squamous cell carcinoma, sweat gland carcinoma, synovioma, thyroid cancer, uveal melanoma, and Wilm's tumor.
  • the disease to be treated by the methods of the present invention may be a hematologic disorder.
  • said hematologic disorder is selected from the group consisting of sickle cell anemia, myelodysplastic disorders (MDS), and myeloproliferative disorders.
  • said myeloproliferative disorder is selected from the group consisting of polycythemia vera, myelofibrosis and essential thrombocythemia.
  • the disease to be treated by the methods of the present invention may be a neurological disorder.
  • said neurological disorder is selected from the group consisting of epilepsy, neuropathic pain, depression and bipolar disorders.
  • the disease to be treated by the methods of the present invention may be a cardiovascular condition.
  • said cardiovascular condition is selected from the group consisting of cardiac hypertrophy, idiopathic cardiomyopathies, and heart failure.
  • the disease to be treated by the methods of the present invention may be an autoimmune disease.
  • said autoimmune disease is selected from the group consisting of systemic lupus erythromatosus (SLE), multiple sclerosis (MS), and systemic lupus nephritis.
  • the disease to be treated by the methods of the present invention may be a dermatologic disorder.
  • said dermatologic disorder is selected from the group consisting of psoriasis, melanoma, basal cell carcinoma, squamous cell carcinoma, and other non-epithelial skin cancers.
  • the disease to be treated by the methods of the present invention may be an ophthalmologic disorder.
  • said ophthalmologic disorder is selected from the group consisting of dry eye, closed angle glaucoma and wide angle glaucoma.
  • the disease to be treated by the methods of the present invention may be a polyglutamine-repeat disorders.
  • the polyglutamine- repeat disorder is selected from the group consisting of Huntington's disease, Spinocerebellar ataxia 1 (SCA 1), Machado-Joseph disease (MID)/Spinocerebella ataxia 3 (SCA 3), Kennedy disease/Spinal and bulbar muscular atrophy (SBMA) and Dentatorubral pallidolusyian atrophy (DRPLA).
  • the disease to be treated by the methods of the present invention may be an inflammatory condition.
  • the inflammatory condition is selected from the group consisting of Rheumatoid Arthritis (RA), Inflammatory Bowel Disease (IBD), ulcerative colitis and psoriasis.
  • HDAC histone deacetylase
  • acyl refers to a carbonyl attached to an alkenyl, alkyl, aryl, cycloalkyl, heteroaryl, heterocycle, or any other moiety were the atom attached to the carbonyl is carbon.
  • An “acetyl” group refers to a -C(O)CH 3 group.
  • An “alkylcarbonyl” or “alkanoyl” group refers to an alkyl group attached to the parent molecular moiety through a carbonyl group. Examples of such groups include methylcarbonyl and ethylcarbonyl. Examples of acyl groups include formyl, alkanoyl and aroyl.
  • alkenyl refers to a straight-chain or branched-chain hydrocarbon radical having one or more double bonds and containing from 2 to 20, preferably 2 to 6, carbon atoms.
  • suitable alkenyl radicals include ethenyl, propenyl, 2-methylpropenyl, 1,4-butadienyl and the like.
  • alkoxy refers to an alkyl ether radical, wherein the term alkyl is as defined below.
  • suitable alkyl ether radicals include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, iso-butoxy, sec-butoxy, tert-butoxy, and the like.
  • alkyl refers to a straight-chain or branched-chain alkyl radical containing from 1 to and including 20, preferably 1 to 10, and more preferably 1 to 6, carbon atoms. Alkyl groups may be optionally substituted as defined herein. Examples of alkyl radicals include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, iso-amyl, hexyl, octyl, noyl and the like.
  • alkylene as used herein, alone or in combination, refers to a saturated aliphatic group derived from a straight or branched chain saturated hydrocarbon attached at two or more positions, such as methylene (-CHr-).
  • alkylamino refers to an alkyl group attached to the parent molecular moiety through an amino group. Suitable alkylamino groups may be mono- or dialkylated, forming groups such as, for example, N-methylamino, N-ethylamino, N,N- dimethylamino, N,N-ethylmethylamino and the like.
  • alkylidene refers to an alkenyl group in which one carbon atom of the carbon-carbon double bond belongs to the moiety to which the alkenyl group is attached.
  • alkylthio refers to an alkyl thioether (R-S-) radical wherein the term alkyl is as defined above and wherein the sulfur may be singly or doubly oxidized.
  • suitable alkyl thioether radicals include methylthio, ethylthio, n- propylthio, isopropylthio, n-butylthio, iso-butylthio, sec-butylthio, tert-butylthio, methanesulfonyl, ethanesulfinyl, and the like.
  • alkynyl refers to a straight-chain or branched chain hydrocarbon radical having one or more triple bonds and containing from 2 to 20, preferably from 2 to 6, more preferably from 2 to 4, carbon atoms.
  • Alkynylene refers to a carbon- carbon triple bond attached at two positions such as ethynylene (-C:::C-, -OC-).
  • alkynyl radicals include ethynyl, propynyl, hydroxypropynyl, butyn-1-yl, butyn-2-yl, pentyn-1-yl, 3- methylbutyn-1-yl, hexyn-2-yl, and the like.
  • acylamino as used herein, alone or in combination, embraces an acyl group attached to the parent moiety through an amino group.
  • An example of an “acylamino” group is acetylamino (CH 3 C(O)NH-).
  • amino refers to— NRR ' , wherein R and R are independently selected from the group consisting of hydrogen, alkyl, acyl, heteroalkyl, aryl, cycloalkyl, heteroaryl, and heterocycloalkyl, any of which may themselves be optionally substituted.
  • aryl as used herein, alone or in combination, means a carbocyclic aromatic system containing one, two or three rings wherein such rings may be attached together in a pendent manner or may be fused.
  • aryl embraces aromatic radicals such as benzyl, phenyl, naphthyl, anthracenyl, phenanthryl, indanyl, indenyl, annulenyl, azulenyl, tetrahydronaphthyl, and biphenyl.
  • arylalkenyl or “aralkenyl,” as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkenyl group.
  • arylalkoxy or “aralkoxy,” as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkoxy group.
  • arylalkyl or “aralkyl,” as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkyl group.
  • arylalkynyl or “aralkynyl,” as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkynyl group.
  • arylalkanoyl or “aralkanoyl” or “aroyl,”as used herein, alone or in combination, refers to an acyl radical derived from an aryl-substituted alkanecarboxylic acid such as benzoyl, napthoyl, phenylacetyl, 3-phenylpropionyl (hydrocinnamoyl), 4-phenylbutyryl, (2-naphthyl)acetyl, 4-chlorohydrocinnamoyl, and the like.
  • aryloxy refers to an aryl group attached to the parent molecular moiety through an oxy.
  • carbamate refers to an ester of carbamic acid (-NHC00-) which may be attached to the parent molecular moiety from either the nitrogen or acid end, and which may be optionally substituted as defined herein.
  • O-carbamyl refers to a -OC(O)NRR', group-with R and R' as defined herein.
  • N-carbamyl as used herein, alone or in combination, refers to a ROC(O)NR'- group, with R and R' as defined herein.
  • carbonyl when alone includes formyl [-C(O)H] and in combination is a -C(O)- group.
  • carboxy refers to -C(O)OH or the corresponding "carboxylate” anion, such as is in a carboxylic acid salt.
  • An "O-carboxy” group refers to a RC(O)O- group, where R is as defined herein.
  • a "C-carboxy” group refers to a -C(O)OR groups where R is as defined herein.
  • cyano as used herein, alone or in combination, refers to -CN.
  • cycloalkyl refers to a saturated or partially saturated monocyclic, bicyclic or tricyclic alkyl radical wherein each cyclic moiety contains from 3 to 12, preferably five to seven, carbon atom ring members and which may optionally be a benzo fused ring system which is optionally substituted as defined herein.
  • cycloalkyl radicals include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, octahydronaphthyl, 2,3-dihydro-lH-indenyl, adamantyl and the like.
  • Bicyclic and tricyclic as used herein are intended to include both fused ring systems, such as decahydonapthalene, octahydronapthalene as well as the multicyclic (multicentered) saturated or partially unsaturated type.
  • the latter type of isomer is exemplified in general by, bicyclo[l,l,l]pentane, camphor, adamantane, and bicyclo[3,2,l]octane.
  • esters refers to a carboxy group bridging two moieties linked at carbon atoms.
  • ether refers to an oxy group bridging two moieties linked at carbon atoms.
  • halo or halogen, as used herein, alone or in combination, refers to fluorine, chlorine, bromine, or iodine.
  • haloalkoxy refers to a haloalkyl group attached to the parent molecular moiety through an oxygen atom.
  • haloalkyl refers to an alkyl radical having the meaning as defined above wherein one or more hydrogens are replaced with a halogen. Specifically embraced are monohaloalkyl, dihaloalkyl and polyhaloalkyl radicals.
  • a monohaloalkyl radical for one example, may have an iodo, bromo, chloro or fluoro atom within the radical.
  • Dihalo and polyhaloalkyl radicals may have two or more of the same halo atoms or a combination of different halo radicals.
  • haloalkyl radicals include fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, heptafluoropropyl, difluorochloromethyl, dichlorofluoromethyl, difluoroethyl, difluoropropyl, dichloroethyl and dichloropropyl.
  • Haloalkylene refers to a haloalkyl group attached at two or more positions. Examples include fluoromethylene
  • heteroalkyl refers to a stable straight or branched chain, or cyclic hydrocarbon radical, or combinations thereof, fully saturated or containing from 1 to 3 degrees of unsaturation, consisting of the stated number of carbon atoms and from one to three heteroatoms selected from the group consisting of O, N, and S, and wherein the nitrogen and sulfur atoms may optionally be oxidized and the nitrogen heteroatom may optionally be quaternized.
  • the heteroatom(s) O, N and S may be placed at any interior position of the heteroalkyl group. Up to two heteroatoms may be consecutive, such as, for example, -CH 2 -NH-OCH 3 .
  • heteroaryl refers to 3 to 7 membered, preferably 5 to 7 membered, unsaturated heteromonocyclic rings, or fused polycyclic rings in which at least one of the fused rings is unsaturated, wherein at least one atom is selected from the group consisting of O, S, and N.
  • the term also embraces fused polycyclic groups wherein heterocyclic radicals are fused with aryl radicals, wherein heteroaryl radicals are fused with other heteroaryl radicals, or wherein heteroaryl radicals are fused with cycloalkyl radicals.
  • heteroaryl groups include pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazolyl, pyranyl, furyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, isothiazolyl, indolyl, isoindolyl, indolizinyl, benzimidazolyl, qninolyl, isoquinolyl, quinoxalinyl, quinazolinyl, indazolyl, benzotriazolyl, benzodioxolyl, benzopyranyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzofuryl, benzothienyl, chro
  • Exemplary tricyclic heterocyclic groups include carbazolyl, benzidolyl, phenanthrolinyl, dibenzofuranyl, acridinyl, phenanthridinyl, xanthenyl and the like.
  • heterocycloalkyl and, interchangeably, “heterocycle,” as used herein, alone or in combination, each refer to a saturated, partially unsaturated, or fully unsaturated monocyclic, bicyclic, or tricyclic heterocyclic radical containing at least one, preferably 1 to 4, and more preferably 1 to 2 heteroatoms as ring members, wherein each said heteroatom may be independently selected from the group consisting of nitrogen, oxygen, and sulfur, and wherein there are preferably 3 to 8 ring members in each ring, more preferably 3 to 7 ring members in each ring, and most preferably 5 to 6 ring members in each ring.
  • Heterocycloalkyl and “heterocycle” are intended to include sulfones, sulfoxides, N-oxides of tertiary nitrogen ring members, and carbocyclic fused and benzo fused ring systems; additionally, both terms also include systems where a heterocycle ring is fused to an aryl group, as defined herein, or an additional heterocycle group.
  • Heterocycle groups of the invention are exemplified by aziridinyl, azetidinyl, 1,3 -benzodioxolyl, dihydroisoindolyl, dihydroisoquinolinyl, dihydrocinnolinyl, dihydrobenzodioxinyl, dihydro[l,3]oxazolo[4,5- b]pyridinyl, benzothiazolyl, dihydroindolyl, dihy-dropyridinyl, 1,3-dioxanyl, 1,4-dioxanyl, 1,3- dioxolanyl, isoindolinyl, morpholinyl, piperazinyl, pyrrolidinyl, tetrahydropyridinyl, piperidinyl, thiomorpholinyl, and the like.
  • the heterocycle groups may be optionally substituted unless specifically prohibited.
  • hydrazinyl as used herein, alone or in combination, refers to two amino groups joined by a single bond, i.e., -N-N-.
  • hydroxyalkyl refers to a hydroxy group attached to the parent molecular moiety through an alkyl group.
  • the phrase "in the main chain” refers to the longest contiguous or adjacent chain of carbon atoms starting at the point of attachment of a group to the compounds of this invention.
  • isocyanato refers to a -NCO group.
  • isothiocyanato refers to a -NCS group.
  • linear chain of atoms refers to the longest straight chain of atoms independently selected from carbon, nitrogen, oxygen and sulfur.
  • lower means containing from 1 to and including 6 carbon atoms.
  • mercaptyl as used herein, alone or in combination, refers to an RS- group, where R is as defined herein.
  • nitro refers to -NO 2 .
  • perhaloalkoxy refers to an alkoxy group where all of the hydrogen atoms are replaced by halogen atoms.
  • perhaloalkyl refers to an alkyl group where all of the hydrogen atoms are replaced by halogen atoms.
  • sulfonate refers the -SO 3 H group and its anion as the sulfonic acid is used in salt formation.
  • thia and thio refer to a -S- group or an ether wherein the oxygen is replaced with sulfur.
  • the oxidized derivatives of the thio group namely sulfinyl and sulfonyl, are included in the definition of thia and thio.
  • thiol as used herein, alone or in combination, refers to an -SH group.
  • thiocarbonyl when alone includes thioformyl -C(S)H and in combination is a -C(S)- group.
  • N-thiocarbamyl refers to an ROC(S)NR'- group, with R and R'as defined herein.
  • O-thiocarbamyl refers to a -OC(S)NRR', group with R and R'as defined herein.
  • thiocyanato refers to a -CNS group.
  • trihalomethanesulfonamido refers to a X 3 CS(O) 2 NR- group with X is a halogen and R as defined herein.
  • trihalomethanesulfonyl refers to a X 3 CS(O) 2 - group where X is a halogen.
  • trimihalomethoxy refers to a X 3 CO- group where X is a halogen.
  • trimethysilyl tert-butyldimethylsilyl, triphenylsilyl and the like.
  • Any definition herein may be used in combination with any other definition to describe a composite structural group.
  • the trailing element of any such definition is that which attaches to the parent moiety.
  • the composite group alkylamido would represent an alkyl group attached to the parent molecule through an amido group
  • the term alkoxyalkyl would represent an alkoxy group attached to the parent molecule through an alkyl group.
  • the term "optionally substituted” means the anteceding group may be substituted or unsubstituted.
  • the substituents of an "optionally substituted” group may include, without limitation, one or more substituents independently selected from the following groups or a particular designated set of groups, alone or in combination: lower alkyl, lower alkenyl, lower alkynyl, lower alkanoyl, lower heteroalkyl, lower heterocycloalkyl, lower haloalkyl, lower haloalkenyl, lower haloalkynyl, lower perhaloalkyl, lower perhaloalkoxy, lower cycloalkyl, phenyl, aryl, aryloxy, lower alkoxy, lower haloalkoxy, oxo, lower acyloxy, carbonyl, carboxyl, lower alkylcarbonyl, lower carboxyester, lower carboxamido, cyano, hydrogen, halogen, hydroxy, amino, lower alkylamino
  • Two substituents may be joined together to form a fused five-, six-, or seven-menbered carbocyclic or heterocyclic ring consisting of zero to three heteroatoms, for example forming methylenedioxy or ethylenedioxy.
  • An optionally substituted group may be unsubstituted (e.g., -CH 2 CH 3 ), fully substituted (e.g., -CF 2 CF 3 ), monosubstituted (e.g., -CH 2 CH 2 F) or substituted at a level anywhere in- between fully substituted and monosubstituted (e.g., -CH 2 CF 3 ).
  • aryl, heterocycle, R, etc. occur more than one time in a formula or generic structure, its definition at each occurrence is independent of the definition at every other occurrence.
  • certain groups may be attached to a parent molecule or may occupy a position in a chain of elements from either end as written.
  • an unsymmetrical group such as -C(O)N(R)- may be attached to the parent moiety at either the carbon or the nitrogen.
  • Asymmetric centers exist in the compounds of the present invention. These centers are designated by the symbols "R” or "S,” depending on the configuration of substituents around the chiral carbon atom.
  • the invention encompasses all stereochemical isomeric forms, including diastereomeric, enantiomeric, and epimeric forms, as well as d-isomers and 1-isomers, and mixtures thereof.
  • Individual stereoisomers of compounds can be prepared synthetically from commercially available starting materials which contain chiral centers or by preparation of mixtures of enantiomeric products followed by separation such as conversion to a mixture of diastereomers followed by separation or recrystallization, chromatographic techniques, direct separation of enantiomers on chiral chromatographic columns, or any other appropriate method known in the art.
  • Starting compounds of particular stereochemistry are either commercially available or can be made and resolved by techniques known in the art. Additionally, the compounds of the present invention may exist as geometric isomers.
  • the present invention includes all cis, trans, syn, anti,
  • E
  • Z
  • compounds may exist as tautomers; all tautomeric isomers are provided by this invention.
  • the compounds of the present invention can exist in unsolvated as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like. In general, the solvated forms are considered equivalent to the unsolvated forms for the purposes of the present invention.
  • bonds refers to a covalent linkage between two atoms, or two moieties when the atoms joined by the bond are considered to be part of larger substructure.
  • a bond may be single, double, or triple unless otherwise specified.
  • a dashed line between two atoms in a drawing of a molecule indicates that an additional bond may be present or absent at that position.
  • combination therapy means the administration of two or more therapeutic agents to treat a therapeutic condition or disorder described in the present disclosure. Such administration encompasses co-administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule having a fixed ratio of active ingredients or in multiple, separate capsules for each active ingredient. In addition, such administration also encompasses use of each type of therapeutic agent in a sequential manner. In either case, the treatment regimen will provide beneficial effects of the drug combination in treating the conditions or disorders described herein.
  • the terms “therapy” or “treating” as used herein refer to (1) reducing the rate of progress of a disease, or, in case of cancer reducing the size of the tumor; (2) inhibiting to some extent further progress of the disease, which in case of cancer may mean slowing to some extent, or preferably stopping, tumor metastasis or tumor growth; and/or, (3) relieving to some extent (or, preferably, eliminating) one or more symptoms associated with the disease.
  • therapeutically effective amount refers to that amount of the compound being administered which will provide therapy or affect treatment.
  • the term "patient” means all mammals including humans. Examples of patients include humans, cows, dogs, cats, goats, sheep, pigs, and rabbits. Preferably, the patient is a human.
  • the compounds of the present invention are also antitumor compounds and/or inhibit the growth of a tumor, i.e., they are tumor-growth-inhibiting compounds.
  • anti-tumor and “tumor-growth-inhibiting,” when modifying the term "compound,” and the terms “inhibiting” and “reducing”, when modifying the terms "compound” and/or “tumor,” mean that the presence of the subject compound is correlated with at least the slowing of the rate of growth of the tumor.
  • anti-tumor refers to a correlation between the presence of the subject compound and at least the temporary cessation of tumor growth.
  • anti-tumor refers to, a correlation between the presence of the subject compound and at least the temporary reduction in the mass of the tumor.
  • the term "function” refers to the cellular role of HDAC.
  • catalytic activity in the context of the invention, defines the rate at which HDAC deacetylates a substrate. Catalytic activity can be measured, for example, by determining the amount of a substrate converted to a product as a function of time. Deacetylation of a substrate occurs at the active-site of HDAC.
  • the active-site is normally a cavity in which the substrate binds to HDAC and is deacetylated.
  • substrate refers to a molecule deacetylated by HDAC.
  • the substrate is preferably a peptide and more preferably a protein.
  • the protein is a histone, whereas in other embodiments, the protein is not a histone.
  • inhibitor refers to decreasing the cellular function of HDAC. It is understood that compounds of the present invention may inhibit the cellular function of HDAC by various direct or indirect mechanisms, in particular by direct or indirect inhibition of the catalytic activityof HDAC.
  • activates refers to increasing the cellular function of HDAC.
  • activate refers to increasing the cellular function of HDAC.
  • HDAC function is preferably the interaction with a natural binding partner and most preferably catalytic activity.
  • modulate refers to altering the function of HDAC by increasing or decreasing the probability that a complex forms between HDAC and a natural binding partner.
  • a modulator may increase the probability that such a complex forms between HDAC and the natural binding partner, or may increase or decrease the probability that a complex forms between HDAC and the natural binding partner depending on the concentration of the compound exposed to HDAC, or may decrease the probability that a complex forms between HDAC and the natural binding partner.
  • a modulator may activate the catalytic activity of HDAC, or may activate or inhibit the catalytic activity of HDAC depending on the concentration of the compound exposed to HDAC, or may inhibit the catalytic activity of HDAC.
  • complex refers to an assembly of at least two molecules bound to one another.
  • natural binding partner refers to polypeptides that bind to HDAC in cells.
  • a change in the interaction between HDAC and a natural binding partner can manifest itself as an increased or decreased probability that the interaction forms, or an increased or decreased concentration of HDAC/natural binding partner complex.
  • the term "contacting" as used herein refers to mixing a solution comprising a compound of the invention with a liquid medium bathing the cells of the methods.
  • the solution comprising the compound may also comprise another component, such as dimethylsulfoxide (DMSO), which facilitates the uptake of the compound or compounds into the cells of the methods.
  • DMSO dimethylsulfoxide
  • the solution comprising the compound of the invention may be added to the medium bathing the cells by utilizing a delivery apparatus, such as a pipet-based device or syringe-based device.
  • monitoring refers to observing the effect of adding the compound to the cells of the method.
  • the effect can be manifested in a change in cell phenotype, cell proliferation, HDAC catalytic activity, substrate protein acetylation levels, gene expression changes, or in the interaction between HDAC and a natural binding partner.
  • effect describes a change or an absence of a change in cell phenotype or cell proliferation.
  • Effect can also describe a change or an absence of a change in the catalytic activity of HDAC.
  • Effect can also describe a change or an absence of a change in an interaction between HDAC and a natural binding partner.
  • cell phenotype refers to the outward appearance of a cell or tissue or the function of the cell or tissue.
  • Examples of cell phenotype are cell size (reduction or enlargement), cell proliferation (increased or decreased numbers of cells), cell differentiation (a change or absence of a change in cell shape), cell survival, apoptosis (cell death), or the utilization of a metabolic nutrient (e.g., glucose uptake). Changes or the absence of changes in cell phenotype are readily measured by techniques known in the art.
  • HDAC inhibitor is used herein to refer to a compound that exhibits an IC 50 with respect to HDAC activity of no more than about lOO ⁇ .M and more typically not more than about 50 ⁇ M, as measured in the biochemical in vitro HDAC-inhibition assay, cellular histone hyperacetylation assay, and differential cytotoxicity assay described generally herein below.
  • IC50 is that concentration of inhibitor which reduces the activity of an enzyme (e.g., HDAC) to half-maximal level. Representative compounds of the present invention have been discovered to exhibit inhibitory activity against HDAC.
  • Compounds of the present invention preferably exhibit an IC 5 0 with respect to HDAC of no more than about 10 ⁇ M, more preferably, no more than about 5 ⁇ M, even more preferably not more than about 1 ⁇ M, and most preferably, not more than about 200 nM, as measured in the HDAC assays described herein.
  • prodrug refers to a compound that is made more active in vivo.
  • Certain compounds of the present invention may also exist as prodrugs, as described in Hydrolysis in Drug and Prodrug Metabolism Chemistry, Biochemistry, and Enzymology (Testa, Bernard and Mayer, Joachim M. Wiley-VHCA, Zurich, Switzerland 2003).
  • Prodrugs of the compounds described herein are structurally modified forms of the compound that readily undergo chemical changes under physiological conditions to provide the compound.
  • prodrugs can be converted to the compound by chemical or biochemical methods in an ex vivo environment. For example, prodrugs can be slowly converted to a compound when placed in a transdermal patch reservoir with a suitable enzyme or chemical reagent.
  • Prodrugs are often useful because, in some situations, they may be easier to administer than the compound, or parent drug. They may, for instance, be bioavailable by oral administration whereas the parent drug is not. The prodrug may also have improved solubility in pharmaceutical compositions over the parent drug.
  • a wide variety of prodrug derivatives are known in the art, such as those that rely on hydrolytic cleavage or oxidative activation of the prodrug.
  • An example, without limitation, of a prodrug would be a compound which is administered as an ester (the "prodrug"), but then is metabolically hydrolyzed to the carboxylic acid, the active entity.
  • peptidyl derivatives of a compound include peptidyl derivatives of a compound.
  • a prodrug is protected thiol compounds. Thiols bearing hydrolyzable protecting groups can unmask protected SH groups prior to or simultaneous to use. As shown below, the moiety -C(O)-R E of a thioester may be hydrolyzed to yield a thiol and a pharmaceutically acceptable acid HO-C(O)-R E .
  • therapeutically acceptable prodrug refers to those prodrugs or zwitterions which are suitable for use in contact with the tissues of patients without undue toxicity, irritation, and allergic response, are commensurate with a reasonable benefit/risk ratio, and are effective for their intended use.
  • a "pharmaceutically active metabolite” is intended to mean a pharmacologically active product produced through metabolism in the body of a specified compound or salt thereof.
  • Metabolites of a compound may be identified using routine techniques known in the art and their activities determined using tests such as those decribed herein.
  • thiol protecting group refers to thiols bearing hydrolyzable protecting groups that can unmask protected SH groups prior to or simultaneous to use.
  • Preferred thiol protecting groups include but are not limited to thiol esters which release pharmaceutically acceptable acids along with an active thiol moiety. Such pharmaceutically acceptable acids are generally nontoxic and do not abrogate the biological activity of the active thiol moiety.
  • Examples of pharmaceutically acceptable acids include, but are not limited to: N,N-diethylglycine; 4-ethylpiperazinoacetic acid; ethyl 2-methoxy-2-phenylacetic acid; N,N-dimethylglycine; (nitrophenoxysulfonyl)benzoic acid; acetic acid; maleic acid; fumaric acid; benzoic acid; tartaric acid; natural amino acids (like glutamate, aspartate, cyclic amino acids such proline); D-amino acids; butyric acid; fatty acids like palmitic acid, stearic acid, oleate; pipecolic acid; phosphonic acid; phosphoric acid; pivalate (trimethylacetic acid); succinic acid; cinnamic acid; anthranilic acid; salicylic acid; lactic acid; and pyruvic acids.
  • Another aspect of the present invention are compounds containing at least one thiol in a protected form, which can be released to provide a SH group prior to or simultaneous to use.
  • Thiol moieties are known to be unstable in the presence of air and are oxidized to the corresponding disulfide.
  • Protected thiol groups are those that can be converted under mild conditions into free thiol groups without other undesired side reactions taking place.
  • Suitable thiol protecting groups include but are not limited to trityl (Trt), allyloxycarbonyl (Alloc), l-(4,4-dimethyl-2,6-dioxocyclohex-l- ylidene)ethyl (Dde), acetamidomethyl (Acm), t-buryl (tBu), or the like.
  • Preferred thiol protecting groups include lower alkanoyl, e.g. acetyl. Free thiol, disulfides, and protected thiols are understood to be within the scope of this invention.
  • terapéuticaally acceptable salt represents salts or zwitterionic forms of the compounds of the present invention which are water or oil-soluble or dispersible; which are suitable for treatment of diseases without undue toxicity, irritation, and allergic-response; which are commensurate with a reasonable benefit/risk ratio; and which are effective for their intended use.
  • the salts can be prepared during the final isolation and purification of the compounds or separately by reacting the appropriate compound in the form of the free base with a suitable acid.
  • Representative acid addition salts include acetate, adipate, alginate, L-ascorbate, aspartate, benzoate, benzenesulfonate (besylate), bisulfate, butyrate, camphorate, camphorsulfonate, citrate, digluconate, formate, fumarate, gentisate, glutarate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2- hydroxyethansulfonate (isethionate), lactate, maleate, malate, malonate, DL-mandelate, mesitylenesulfonate, methanesulfonate, naphthylenesulfonate, nicotinate, 2-na ⁇ hthalenesulfonate, oxalate, pamoate, pectinate, persulf
  • basic groups in the compounds of the present invention can be quaternized with methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides; dimethyl, diethyl, dibulyl, and diamyl sulfates; decyl, lauryl, myristyl, and steryl chlorides, bromides, and iodides; and benzyl and phenethyl bromides.
  • acids which can be employed to form therapeutically acceptable addition salts include inorganic acids such as hydrochloric, hydrobromic, sulfuric, and phosphoric, and organic acids such as oxalic, maleic, succinic, and citric. Salts can also be formed by coordination of the compounds with an alkali metal or alkaline earth ion.
  • the present invention contemplates sodium, potassium, magnesium, and calcium salts of the compounds of the compounds of the present invention and the like.
  • Basic addition salts can be prepared during the final isolation and purification of the compounds by reacting a carboxy group with a suitable base such as the hydroxide, carbonate, or bicarbonate of a metal cation or with ammonia or an organic primary, secondary, or tertiary amine.
  • a suitable base such as the hydroxide, carbonate, or bicarbonate of a metal cation or with ammonia or an organic primary, secondary, or tertiary amine.
  • the cations of therapeutically acceptable salts include lithium, sodium, potassium, calcium, magnesium, and aluminum, as well as nontoxic quaternary amine cations such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, iV,iV-dimethylaniline, N- methylpiperidine, iV-methylmorpholine, dicyclohexylamine, procaine, dibenzylamine, N,N- dibenzylphenethylamine, 1-ephenamine, and ⁇ iV-dibenzylethylenediamine.
  • Other representative organic amines useful for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, and piperazine.
  • the compounds of the present invention can exist as therapeutically acceptable salts.
  • the present invention includes compounds listed above in the form of salts, in particular acid addition salts. Suitable salts include those formed with both organic and inorganic acids. Such acid addition salts will normally be pharmaceutically acceptable. However, salts of non-pharmaceutically acceptable salts may be of utility in the preparation and purification of the compound in question. While it may be possible for the compounds of the subject invention to be administered as the raw chemical, it is also possible to present them as a pharmaceutical formulation. Accordingly, the subject invention provides a pharmaceutical formulation comprising a compound or a pharmaceutically acceptable salt, ester, prodrug or solvate thereof, together with one or more pharmaceutically acceptable carriers thereof and optionally one or more other therapeutic ingredients.
  • the carrier(s) must be "acceptable” in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. Proper formulation is dependent upon the route of administration chosen. Any of the well-known techniques, carriers, and excipients may be used as suitable and as understood in the art; e.g., in Remington's Pharmaceutical Sciences.
  • the pharmaceutical compositions of the present invention may be manufactured in a manner that is itself known, e.g., by means of conventional mixing, dissolving, granulating, dragee- making, levigating, emulsifying, encapsulating, entrapping or compression processes.
  • the formulations include those suitable for oral, parenteral (including subcutaneous, intradermal, intramuscular, intravenous, intraarticular, and intramedullary), intraperitoneal, transmucosal, transdermal, rectal and topical (including dermal, buccal, sublingual and intraocular) administration although the most suitable route may depend upon for example the condition and disorder of the recipient.
  • the formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy. AU methods include the step of bringing into association a compound of the subject invention or a pharmaceutically acceptable salt, ester, prodrug or solvate thereof ("active ingredient”) with the carrier which constitutes one or more accessory ingredients.
  • active ingredient a pharmaceutically acceptable salt, ester, prodrug or solvate thereof
  • the formulations are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both and then, if necessary, shaping the product into the desired formulation.
  • Formulations of the present invention suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous liquid or a non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion.
  • the active ingredient may also be presented as a bolus, electuary or paste.
  • compositions which can be used orally include tablets, push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. Tablets may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with binders, inert diluents, or lubricating, surface active or dispersing agents. Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent.
  • the tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein. All formulations for oral administration should be in dosages suitable for such administration.
  • the push-fit capsules can contain the active ingredients in admixture with filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers.
  • the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols.
  • stabilizers may be added.
  • Dragee cores are provided with suitable coatings.
  • concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures.
  • Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses.
  • the compounds may be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion.
  • Formulations for injection may be presented in unit dosage form, e.g., in ampoules or in multi-dose containers, with an added preservative.
  • the compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents.
  • the formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in powder form or in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example, saline or sterile pyrogen-free water, immediately prior to use.
  • sterile liquid carrier for example, saline or sterile pyrogen-free water
  • Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.
  • Formulations for parenteral administration include aqueous and non-aqueous (oily) sterile injection solutions of the active compounds which may contain antioxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents.
  • Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes.
  • Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran.
  • the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds to allow for the preparation of highly concentrated solutions.
  • the compounds may also be formulated as a depot preparation. Such long acting formulations may be administered by implantation (for example subcutaneously or intramuscularly) or by intramuscular injection.
  • the compounds may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.
  • compositions may take the form of tablets, lozenges, pastilles, or gels formulated in conventional manner.
  • Such compositions may comprise the active ingredient in a flavored basis such as sucrose and acacia or tragacanth.
  • the compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter, polyethylene glycol, or other glycerides.
  • Compounds of the present invention may be administered topically, that is by non- systemic administration. This includes the application of a compound of the present invention externally to the epidermis or the buccal cavity and the instillation of such a compound into the ear, eye and nose, such that the compound does not significantly enter the blood stream.
  • systemic administration refers to oral, intravenous, intraperitoneal and intramuscular administration.
  • Formulations suitable for topical administration include liquid or semi-liquid preparations suitable for penetration through the skin to the site of inflammation such as gels, liniments, lotions, creams, ointments or pastes, and drops suitable for administration to the eye, ear or nose.
  • the active ingredient may comprise, for topical administration, from 0.001% to 10% w/w, for instance from 1% to 2% by weight of the formulation. It may however comprise as much as 10% w/w but preferably will comprise less than 5% w/w, more preferably from 0.1% to 1% w/w of the formulation.
  • the compounds according to the invention are conveniently delivered from an insufflator, nebulizer pressurized packs or other convenient means of delivering an aerosol spray.
  • Pressurized packs may comprise a suitable propellant such as
  • the dosage unit may be determined by providing a valve to deliver a metered amount.
  • the compounds according to the invention may take the form of a dry powder composition, for example a powder mix of the compound and a suitable powder base such as lactose or starch.
  • the powder composition may be presented in unit dosage form, in for example, capsules, cartridges, gelatin or blister packs from which the powder may be administered with the aid of an inhalator or insufflator.
  • pharmaceutical preparations of compound(s) or active ingredient(s) of the present invention may be formulated by Latitude Pharmaceuticals Inc. located in 9865 Mesa Rim Road, STE 201, San Diego, CA 92121 using their trade secret and proprietary formulation named "FlOl".
  • the composition of said formulation FlOl is known to contain triglyceride, soy lecithin, vitamin E and PEG400.
  • Preferred unit dosage formulations are those containing an effective dose, as herein below recited, or an appropriate fraction thereof, of the active ingredient.
  • formulations of this invention may include other agents conventional in the art having regard to the type of formulation in question, for example those suitable for oral administration may include flavoring agents.
  • the compounds of the invention may be administered orally or via injection at a dose of from 0.1 to 500 mg/kg per day.
  • the dose range for adult humans is generally from 5 mg to 2 g/day.
  • Tablets or other forms of presentation provided in discrete units may conveniently contain an amount of compound of the invention which is effective at such dosage or as a multiple of the same, for instance, units containing 5 mg to 500 mg, usually around 10 mg to 200 mg.
  • the amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration.
  • the compounds of the subject invention can be administered in various modes, e.g. orally, topically, or by injection.
  • the precise amount of compound administered to a patient will be the responsibility of the attendant physician.
  • the specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diets, time of administration, route of administration, rate of excretion, drug combination, the precise disorder being treated, and the severity of the indication or condition being treated.
  • the route of administration may vary depending on the condition and its severity.
  • the compounds described herein may be administered in combination with another therapeutic agent.
  • another therapeutic agent such as a pharmaceutically acceptable salt, ester, or prodrug thereof.
  • the therapeutic effectiveness of one of the compounds described herein may be enhanced by administration of an adjuvant (i.e., by itself the adjuvant may only have minimal therapeutic benefit, but in combination with another therapeutic agent, the overall therapeutic benefit to the patient is enhanced).
  • the benefit of experienced by a patient may be increased by administering one of the compounds described herein with another therapeutic agent (which also includes a therapeutic regimen) that also has therapeutic benefit.
  • another therapeutic agent which also includes a therapeutic regimen
  • increased therapeutic benefit may result by also providing the patient with another therapeutic agent for cancer.
  • the overall benefit experienced by the patient may simply be additive of the two therapeutic agents or the patient may experience a synergistic benefit.
  • combination therapies include use of the compounds of the invention with another chemotherapeutic agent such as aromatase inhibitors, antiestrogen, anti-androgen, or a gonadorelin agonists, topoisomerase 1 and 2 inhibitors, microtubule active agents, alkylating agents, antimeoplastic antimetabolite, or platin containing compound, lipid or protein kinase targeting agents, protein or lipid phosphatase targeting agents, anti-angiogentic agents, agents that induce cell differentiation, bradykinin 1 receptor and angiotensin II antagonists, cyclooxygenase inhibitors, heparanase inhibitors, lymphokines or cytokine inhibitors, bisphosphanates, rapamycin derivatives, anti-apoptotic pathway inhibitors, apoptotic pathway agonists, PPAR agonists, inhibitors of Ras isoforms, telomerase inhibitors, protease inhibitors, metalloproteinas
  • the chemotherapeutic agents that are useful for the treatment of multiple myeloma include, but are not limited to, alkylating agents (eg, melphalan), anthracyclines (eg. doxorubicin), corticosteroids (eg. dexamethasome), IMiDs (eg. thalidomide, lenalidomide), protease inhibitors (eg. bortezomib, NPI0052), IGF-I inhibitors, CD40 antibodies, Smac mimetics (eg. telomestatin), FGF3 modulator (eg. CHIR258), mTOR inhibitor (Rad 001), HDAC inhibitors (eg. SAHA, Tubacin), IKX inhibitors, P38MAPK inhibitors, HSP90 inhibitors (eg 17-AAG), and akt inhibitors (eg. Perifosine).
  • alkylating agents eg, melphalan
  • anthracyclines eg
  • the preferred chemotherapeiitic agents used in combination with the compounds of the present invention include without limitation melphalan, doxorubicin (including lyophilized), dexamethasone, prednisone, thalidomide, lenalidomide, bortezomib, andNPI0052.
  • the multiple chemotherapeutic agents may be administered in any order or even simultaneously. If simultaneously, the multiple chemotherapeutic agents may be provided in a single, unified form, or in multiple forms (by way of example only, either as a single pill or as two separate pills).
  • the multiple chemotherapeutic agents may be provided in a single, unified form, or in multiple forms (by way of example only, either as a single pill or as two separate pills).
  • chemotherapeutic agents may be given in multiple doses, or both may be given as multiple doses. If not simultaneous, the timing between the multiple doses may be any duration of time ranging from a few minutes to four weeks.
  • the present invention provides methods for treating HDAC- mediated disorders in a human or animal subject in need of such treatment comprising administering to said subject an amount of a compound of the present invention effective to reduce or prevent said disorder in the subject in combination with at least one additional agent for the treatment of said disorder that is known in the art.
  • the present invention provides therapeutic compositions comprising at least one compound of the present invention in combination with one or more additional agents for the treatment of HDAC -mediated disorders.
  • the present invention includes compounds listed above in the form of salts, in particular acid addition salts.
  • Suitable salts include those formed with both organic and inorganic acids.
  • Such acid addition salts will normally be pharmaceutically acceptable.
  • salts of non- pharmaceutically acceptable salts may be of utility in the preparation and purification of the compound in question.
  • preferred salts include those formed from hydrochloric, hydrobromic, sulphuric, citric, tartaric, phosphoric, lactic, pyruvic, acetic, succinic, oxalic, fumaric, maleic, oxaloacetic, methanesulphonic, ethanesulphonic, p-toluenesulphonic, benzenesulphonic and isethionic acids.
  • a salt of a compound can be made by reacting the appropriate compound in the form of the free base with the appropriate acid.
  • Reagents (a) H 2 , Raney Nickel, toluene, (b) 1] NaNO 2 , HCl 2] EtOCS 2 K. (c) LiOH, THF,
  • Reagents (a) Pyridine, 4O 0 C, 6h (b) Copper iodide, 1,10-phenanthroline, cesium carbonate,
  • Reagents (a) Acetic anhydride, pyridine, 6O 0 C, 18h. (b) PCl 5 , 60°C, 6h. (c) Pyridine, THF,
  • Reagents (a) Pyridine, THF, 40°C, 6h. (b) Trimethylphenylammonium tribromide, THF,
  • Reagents (a) Pyridine, THF, 40°C, 6h. (b) Trimethylphenylammonium tribromide, THF,
  • Step 1 N-(4 ⁇ Acetyl-phenyl)-4-butoxy-benzenesuIfbnamide
  • Step 3 Thioacetic acid S- ⁇ 2-[4-(4-butoxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl ⁇ ester
  • Step 1 N-[4-(4-Acetyl-phenylsuIfamoyl)-phenyl]-acetamide
  • N-[4-(4-Acetyl- ⁇ henylsulfamoyl)-phenyl]-acetamide was synthesized as described in EXAMPLE 1, Step 1 using 4-acetylamino-benzenesulfonyl chloride and l-(4-amino-phenyl)-ethanone as starting materials.
  • N- ⁇ 4-[4-(2-Bromo-acefyl)-phenylsulfamoyl]-phenyl ⁇ -acetamide was synthesized according to EXAMPLE 1, Step 2 using N-[4-(4-acetyl-phenylsulfamoyl)-phenyl]-acetamide as starting material.
  • N- ⁇ 4-[4-(2-Bromo-acetyl)-phenylsulfamoyl]-phenyl ⁇ -acetamide (530 mg, 1.29 mmol) was dissolved in 1,4-dioxane (8 mL) and aqueous HCl (IM, 4 mL) and the resulting mixture was heated to 110 0 C for 2h. The reaction mixture was then cooled to room temperature and slowly poured into water containing Na 2 CO 3 . The aqueous layer was extracted with CH 2 Cl 2 (3x).
  • Step 1 N-(5-Acetyl-pyridin-2-yl)-4-iodo-benzenesulfonamide
  • N-(5-Acetyl-pyridin-2-yl)-4-iodo-benzenesulfonamide was synthesized as described in EXAMPLE 1, Step 1 using 4-iodo-benzenesulfonyl chloride and l-(6-ammo-pyridin-3-yl)-ethanone as starting materials and pyridine as solvent.
  • 1 HNMR 400 MHz, DMSO-dg) ⁇ 8.59 (s, IH), 8.13 (dd, IH), 7.92 (d, 2H), 7.64 (d, 2H), 7.20 (d, IH), 2.45 (s, 3H).
  • Step 2 N-(5-Acetyl-pyridin-2-yl)-4-(2-methoxy-ethoxy)-benzenesulfonamide
  • Step 3 N-[5-(2-Bromo-acetyl)-pyridin-2-yl]-4-(2-methoxy-ethoxy)-benzenesulfonainide
  • N-[5-(2-Bromo-acetyl)-pyridin-2-yl]-4-(2-methoxy-ethoxy)-benzenesulfonamide was synthesized as described in EXAMPLE 1 , Step 2 using N-(5-acetyl-pyridin-2-yl)-4-(2-methoxy-ethoxy)- benzenesulfonamide as starting material.
  • Step 4 Thioacetic acid S-(2- ⁇ 6-[4-(2-methoxy-ethoxy)-benzenesulfonylamino]-pyridin-3-yl ⁇ -2- oxo-ethyl) ester
  • This example is intentionally left blank.
  • Raney nickel was washed with iPrOH followed by toluene. Under N 2 atmosphere, l-methyl-5- nitroimidazole (0.5Og, 2,8 mmol) was added to toluene (20 mL) containing the pre-washed Raney nickel. The reaction flask was purged with H 2 and the reaction mixture was warmed to 40 0 C for two hours. The resulting mixture was filtered through celite and concentrated to give 0.342 g (82%) of a white solid.
  • Step 2 Dithiocarbonic acid O-ethyl ester S-(l ⁇ methyl-lH-benzoimidazol-5-yl) ester:
  • l-Methyl-lH-benzoimidazol-5-ylamine (2.3 Ig, 15.7 mmol) obtained from step 1 was suspended in 30 mL of water, the solution was then cooled to 0 0 C and aqueous HCl (3.14 mL, 37.9 mmol) was added. The reaction mixture was stirred in an ice/ water bath. NaNO 2 in 5 mL OfH 2 O was added in small portion to the reaction mixture over a period of 30 minutes. The reaction mixture was then added portionwise to a 45 0 C solution OfEtOCS 2 K in 10 mL of water. The reaction mixture was stirred at 45 0 C for an additional 30 minutes, and then it was extracted with Et 2 O (2 x 25 mL).
  • Step 4 l-Methyl-lH-benzoimidazole-5-sulfonyl chloride
  • Step 5 l-Methyl-lH-be ⁇ zoimidazole-5-sulfonic acid (4-acetyl-phenyl)-amide
  • Step 6 l-Methyl-lH-benzoimidazole-5-sulfonic acid [4-(2-bromo-acetyl)-phenyl]-amide
  • Step 7 Thioacetic acid S- ⁇ 2-[4-(l-methyl-lH-benzoimidazole-5-sulfonylamino)-phenyl]-2-oxo- ethyl ⁇ ester
  • Step 1 l-(2,3-Dihydro-indol-l-yl)-ethanone
  • Step 2 l-Acetyl-2,3-dihydro-lH-indole-5-sulfonyl chloride
  • Step 3 l-Acetyl-2,3 ⁇ dihydro-lH-indole-5-sulfonic acid [4-(2-bromo-acetyl)-phenyl]-amide
  • Step 1 l-(5-Bromo-2,3-dihydro-indol-l-yl)-ethanone
  • Step 3 l-Acetyl-5-bromo-2,3-dihydro-lH-indole-6-sulfonic acid (4-acetyl-phenyl)-amide
  • Step 4 l-Acetyl-2,3-dihydro-lH-indole-6-sulfonic acid (4-acetyl-phenyl)-amide
  • Step 1 lH-Indole-6-sulfonic acid [4-(2-chloro-acetyl)-phenyl]-amide
  • Step 2 Thioacetic acid S- ⁇ 2-[4-(lH-indole-6-sulfonylamlno)-phenyl]-2-oxo-ethyl ⁇ ester
  • Step 1 Sodium 4-acetoxy-benzenesulfonate
  • Acetic anhydride 250 mL.and pyridine (16g, 202.28 mmol) were added to sodium 4- hydroxybenzenesulfonate (36g, 183.52 mmol) and the resulting solution was allowed to react, with stirring, overnight while the temperature was maintained at 60-70°C until done by TLC. The mixture was concentrated by evaporation under vacuum using a rotary evaporator. This resulted in 35g (80%) of sodium 4-acetoxybenzenesulfonate as a dark red solid.
  • Acetic acid 4-(4-acetyl-phenylsulfamoyl)-phenyl ester was synthesized as described in EXAMPLE 1, Step 1 using acetic acid 4-chlorosulfonyl-phenyl ester and l-(4-amino-phenyl)-ethanone as starting materials.
  • Step 4 N-(4-acetyl-phenyl)-4-hydroxy-be ⁇ zenesulfonamide
  • Adjustment of the pH to 8 was accomplished by the addition of Na 2 COs (2-3%). The resulting solution was extracted three times with 200 ml of CH 2 Cl 2 and one time with 200 ml of EtOAc, the aqueous layers combined. Adjustment of the pH to 6 was accomplished by the addition of HCl (2-3%). The resulting solution was extracted three times with 200 ml of CH 2 Cl 2 and the organic layers combined, dried over MgSO 4 and concentrated by evaporation under vacuum using a rotary evaporator. This resulted in 3 g (43%) of N- (4- acetylphenyl)-4-hydroxybenzenesulfonamide as a light gray solid.
  • Step 5 Thioacetic acid S- ⁇ 2-[4-(4-hydroxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl ⁇ ester
  • Step 2 Thioacetic acid S- ⁇ 2-[2-(4-methoxy-be ⁇ zenesulfonylamino)-pyrimidin-5-yl]-2-oxo- ethyl ⁇ ester
  • This example is intentionally left blank.
  • Step 1 N-(5-Acetyl-pyridin-2-yl)-4-amino-benzenesulfonamide
  • N-[4-(5-acetyl-pyridine-2-ylsulfamoyl)-phenyl]-acetamide (1.33 g, 3.98 mmol) was dissolved in a dioxane HCl solution (10 ml, 4 M). Water (20 ml) was added, and the resulting tan solution was heated to reflux for 1 hour. The volatiles were removed, and the resulting tan solid was purified by HPLC to leave the desired amine as a tan solid (0.78 g, 67 %).
  • Step 2 N-[4-(5-Acetyl-pyridin-2-ylsulfamoyl)-phenyl]-succinamic acid methyl ester
  • N-(5-acetyl-pyridin-2-yl)-4-amino-benzenesulfonamide (149 mg, 0.51 mmol) was suspended in THFVCH 2 Cl 2 (1:1, 4 ml) before triethylamine (0.21 ml, 1.53 mmol) and DMF (0.5 ml) were added to afford a tan solution.
  • Methyl 4-chloro-4-oxo-butyrate (0.07 ml, 0.56 mmol) was then added as a liquid. The resulting yellow mixture was then stirred at room temperature for 15 minutes, before being filtered, and the mother liquor evaporated to leave a yellow oil which was purified by HPLC.
  • Step 3 N- ⁇ 4-[5-(2-Bromo-acetyl)-pyridin-2-ylsulfamoyl]-phenyl ⁇ -succinamic acid methyl ester
  • N- ⁇ 4-[5-(2-Bromo-acetyl)-pyridin-2-ylsulfamoyl]-phenyl ⁇ -succinamic acid methyl ester was synthesized as described in EXAMPLE 1, Step 2, using N-[4-(5-Acetyl- ⁇ yridin-2-ylsulfamoyl)- phenyl]-succinamic acid methyl ester as starting material
  • Step 4 N- ⁇ 4-[5-(2-Acetylsulfanyl-acetyl)-pyridin-2-ylsulfamoyl]-phenyl ⁇ -succinamic acid methyl ester
  • N- ⁇ 4-[5-(2-Acetylsulfanyl-acetyl)-pyridin-2-ylsulfamoyl]- ⁇ henyl ⁇ -succinamic acid methyl ester was synthesized as described in EXAMPLE 1, Step 3, using N- ⁇ 4-[5-(2-Bromo-acetyl)- ⁇ yridin-2- ylsulfamoylj-phenylj-succinamic acid methyl ester as starting material.
  • This assay measures a compound's ability to inhibit acetyl-lysine deacetylation in vitro and was used as both a primary screening method as well as for IC50 determinations of confirmed inhibitors.
  • the assay is performed in vitro using an HDAC enzyme source ⁇ e.g. partially purified nuclear extract or immunopurified HDAC complexes) and a proprietary fluorescent source ⁇ e.g. partially purified nuclear extract or immunopurified HDAC complexes.
  • HDAC Quantizyme Fluor de Lys Fluorescent Activity Assay BIOMOL
  • BIOMOL HDAC Quantizyme Fluor de Lys Fluorescent Activity Assay
  • Step 1 Enzyme (2.5 ⁇ L) source added to plate (from refrigerated container)
  • Step 2 Compounds (50 nL) added with pin transfer device
  • Step 3 Fluor de Lys (2.5 ⁇ L) substrate added, incubate at RT, 30 minutes
  • Step 4 Developer (5 ⁇ L) solution is added (containing TSA), to stop reaction
  • Step 5 Plate Reader - data collection
  • the deacetylated fluorophore is excited with 360 nm light and the emitted light
  • Transformed cell lines e.g. HeLa, A549, MCF-7 are cultured under standard media and culture conditions prior to plating.
  • Cells (approx.2,500/well) are allowed to adhere 10-24 hours to wells of a 384-well Greiner PS assay plate in media containing 1-5% serum. Cells are treated with appropriate compound and specific concentrations for 0 to 24 hours. Cells are washed once with PBS (60 ⁇ L) and then fixed (95% ethanol, 5% acetic acid or 2% PFA) for 1 minute at RT (30 ⁇ L). Cells are blocked with 1% BSA for 1 hour and washed and stained with antibody (e.g. anti-Acetylated Histone H3, Upstate Biotechnology), followed by washing and incubation with an appropriate secondary antibody conjugated to HRP or fluorophore. For luminescence assays, signal is generated using Luminol substrate (Santa Cruz Biotechnology) and detected using an Aquest plate reader (Molecular Devices).
  • Cells (4 x 10 5 /well) are plated into Corning 6-well dish and allowed to adhere overnight. Cells are treated with compound at appropriate concentration for 12-18 hours at 37 degrees. Cells are washed with PBS on ice. Cells are dislodged with rubber policeman and lysed in buffer containing 25 mM Tris, pH7.6; 150 mM NaCl, 25 mM MgCl 2 , 1% Tween-20, and nuclei collected by centrifugation (750Og). Nuclei are washed once in 25 mM Tris, pH7.6; 10 mM EDTA, collected by centrifugation (750Og). Supernatant is removed and histones are extracted using 0.4 M HCl. Samples are centrifuged at 1400Og and supernatants are precipitated in 1 ml cold acetone. The histone pellet is dissolved in water and histones are separated and analyzed by SDS-PAGE
  • HDAC inhibitors display differential cytotoxicity toward certain transformed cell lines.
  • Cells are cultured according to standard ATCC recommended conditions that are appropriate to each cell type. Compounds were tested for their ability to kill different cell types (normal and transformed) using the ATPlite luminescence ATP detection assay system (Perkin Elmer). Assays are run in either 384-well or 1536-well Greiner PS plates. Cells (30 ⁇ L or 5 ⁇ L, respectively) are dispensed using either multichannel pipette for 384-well plates, or proprietary Kalypsys bulk liquid dispenser for 1536-well plates. Compounds added using proprietary pin-transfer device (500 nL or 5 nL) and incubated 5 to 30 hours prior to analysis. Luminescence is measured using Aquest plate reader (Molecular Devices).

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Abstract

Disclosed herein are compounds of formula I and methods used for treating disease states including, but not limited to cancers, autoimmune diseases, tissue damage, central nervous system disorders, neurodegenerative disorders, fibrosis, bone disorders, polyglutamine- repeat disorders, anemias, thalassemias, inflammatory conditions, cardiovascular conditions, and disorders in which angiogenesis play a role in pathogenesis. In addition, methods of modulating the activity of histone deacetylase (HDAC) are also disclosed.

Description

INHIBITORS OF HISTONE DEACETYLASE FOR THE TREATMENT OF DISEASE
This application claims the benefit of priority of United States provisional application No.s 60/748,822 and 60/748,823, both filed December 9, 2005, United States provisional application No. 60/784,644, filed March 20, 2006, and United States provisional application No. 60/802,823, filed May 22, 2006, the disclosures of which are hereby incorporated by reference as if written herein in their entireties.
FIELD OF THE INVENTION
Histone proteins organize DNA into nucleosomes, which are regular repeating structures of chromatin. The acetylation status of histones alters chromatin structure, which, in turn, is involved in gene expression. Two classes of enzymes can affect the acetylation of histones - histone acetyltransferases (HATs) and histone deacetylases (HDACs). A number of HDAC inhibitors have been characterized. One of the potent inhibitors of HDAC is (SAHA), ahydroxamic acid-based compound. It is also known as vorinostat or ZOLINZA(TM), which is currently in clinical trials. ("Merck Announces Pivotal Phase lib Study Results of the Company's Investigational HDAC Inhibitor ZOLINZA(TM) and Glaxo's Cancer Vaccine Shows Response," M2 Presswire, 5 June 2006.) The Food and Drug Administration (FDA) has also accepted the New Drug Application (NDA) for ZOLINZA(TM) for the treatment of advanced cutaneous T-cell-lymphoma (CTCL) in June 2006. (WHITEHOUSE STATION, NJ., "ZOLINZA(TM), Merck's Investigational Medicine for Advanced Cutaneous T-CeIl Lymphoma (CTCL), to Receive Priority Review from U.S. Food and Drug Administration," Business Wire, 7 June 2006.)
Certain non-hydroxamate carbonyl compounds as HDAC inhibitors have previously been published. In PCT Patent Application Publication No. WO 04/110418, published December 23, 2004, Wash et al. first discloses a novel, non-hydroxamate carbonyl class of HDAC inhibitor. In
PCT Patent Application Publication No. WO 05/123089, published December 29, 2005, Malecha et al. describes multicyclic sulfonamide carbonyl compounds as HDAC inhibitors. In PCT Patent Application Publication No. WO 05/120515, published December 29, 2005, Malecha et al. discloses somewhat different sulfonamide carbonyl compounds as HDAC inhibitors.
BACKGROUND OF THE INVENTION
Histone proteins organize DNA into nucleosomes, which are regular repeating structures of chromatin. The acetylation status of histones alters chromatin structure, which, in turn, is involved in gene expression. Two classes of enzymes can affect the acetylation of histones - histone acetyltransferases (HATs) and histone deacetylases (HDACs). A number of HDAC inhibitors have been characterized. However, to date no effective candidate for cancer therapy has been identified. Therefore, there is a need in the art to discover HDAC inhibitors that have effective anti-tumor activity. SUMMARY OF THE INVENTION
Disclosed herein are compounds useful for treating diseases by inhibiting HDAC.
In certain embodiments of carbonyl compounds of Formula (I), as described herein, including their pharmaceutical acceptable salts, esters, and prodrugs. Compounds of Formula (I) have the following structure
Figure imgf000003_0001
wherein:
Gi is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
G2 is selected from the group consisting of an N-sulfonamide moiety having structure (II), an S-sulfonamide moiety having structure (III), an amide of the form -NR3C(O)- , and an amide of the form -C(O)NR3-:
Figure imgf000003_0002
G3 is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
Ri and R2 are each independently selected from the group consisting of hydrogen, lower alkyl, halogen andperhaloalkyl, or R1 and R2 taken together may form an optionally substituted cycloalkyl or optionally substituted heterocycloalkyl;
R3 and R4 are each independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, and optionally substituted aryl;
T is selected from the group consisting of -(XOn1NR7C(O)(X2)D2Rs and - (X1)mSO2(X2)π2Rs;
nl is 0-5;
n2 is 0-5;
R7 is selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted heteroalkyl, and optionally substituted lower alkoxy;
R8 is selected from the group consisting of optionally substituted lower alkyl, optionally substituted heteroalkyl, optionally substituted heterocyclyl, optionally substituted heteroaryl and - NR9Ri0;
R9 and Rio are each independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted lower alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted cycloaklenyl, optionally substituted fused aryl, optionally substituted fused heteroaryl, optionally substituted fused heterocycloalkyl, and optionally substituted fused cycloalkyl; or R9 and R10, taken together with the nitrogen to which they are attached, form an optionally substituted five-membered or six-membered heteroaromatic or heteroaliphatic ring; and
Ge is selected from the group consisting of hydrogen, optionally substituted acyl, optionally substituted aryl, optionally substituted alkyl, optionally substituted heteroaryl, optionally substituted alkylthio, optionally substituted arylthio and optionally substituted heteroarylthio; or G6 may have the structural formula (IV) thereby forming a homodisulfϊde or heterodisulfϊde dimer of a compound of the present invention
Figure imgf000004_0001
wherein:
G7 is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
G8 is selected from the group consisting of an N-sulfonamide moiety having structure (V), an S-sulfonamide moiety having structure (VI), an amide of the form -NR13C(O)- , and an amide of the form -C(O)NR14-:
Figure imgf000004_0002
G9 is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
R11 and R^ are each independently selected from the group consisting of hydrogen, lower alkyl, halogen and perhaloalkyl, or Ru and R^ taken together may form an optionally substituted cycloalkyl or optionally substituted heterocycloalkyl; and
R13 and R14 are each independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, and optionally substituted aryl.
The invention provides methods and compositions for treating diseases in mammals using compounds of the invention, including but not limited to, treating cancers, autoimmune diseases, tissue damage, central nervous system disorders, neurodegenerative disorders, fibrosis, bone disorders, polyglutamine-repeat disorders, anemias, thalassemias, inflammatory conditions, cardiovascular conditions, and disorders in which angiogenesis plays a role in pathogenesis.
The invention further provides methods of inhibiting the catalytic activity and cellular function of histone deacetylase (HDAC). DETAILRD DESCRIPTION OF THE INVENTION
In further embodiments of compounds of Formula (I) wherein G6 is selected from the group consisting of optionally substituted acyl and hydrogen.
In yet further embodiments, the compound of the invention wherein G1 is optionally substituted 6 membered aryl or optionally substituted 6 membered heteroaryl.
In yet further embodiments, the compound of the invention wherein G2 is N-sulfonamide. In certain embodiments, the compound of the invention wherein G3 is phenyl.
In further embodiments, the compound of the invention wherein:
Gi is pyridinyl; and
T is -CXi)nINR7C(O)R8-; and
nl is 0.
In further embodiments, the compound of the invention wherein:
Gi is phenyl;
T is -CXOmNR7C(O)Rs-; and
nl is 0.
In accordance with one aspect, the present invention provides compounds capable of inhibiting the catalytic activity of histone deacetylase (HDAC). In another aspect, the present invention provides pharmaceutical compositions comprising compounds capable of inhibiting the catalytic activity of histone deacetylase CHDAC).
In accordance with yet another aspect of the invention, the present invention provides methods and compositions for treating certain diseases or disease states.
In another aspect are compounds or compositions comprising compounds capable of inhibiting the catalytic activity of histone deacetylase (HDAC).
In some aspects of the invention, the disease is a hyperproliferative condition of the human or animal body.
In another aspects of the invention, said hyperproliferative condition is selected from the group consisting of hematologic and nonhematologic cancers. In yet further embodiments, said hematologic cancer is selected from the group consisting of multiple myeloma, leukemias, and lymphomas. In yet another aspects of the invention, said leukemia is selected from the group consisting of acute and chronic leukemias. In yet further embodiments, said acute leukemia is selected from the group consisting of acute lymphocytic leukemia CALL) and acute nonlymphocytic leukemia (ANLL). In yet further embodiments, said chronic leukemia is selected from the group consisting of chronic lymphocytic leukemia (CLL) and chronic myelogenous leukemia (CML). In further embodiments, said lymphoma is selected from the group consisting of Hodgkin's lymphoma and non-Hodgkin's lymphoma. In further embodiments, said lymophoma is selected from the group consisting of cutaneous t-cell lymphoma (CTCL) and mantle cell lymphoma (MCL). In further embodiments, said hematologic cancer is multiple myeloma (MM). In other embodiments, said hematologic cancer is of low, intermediate, or high grade. In other embodiments, said
nonhematologic cancer is selected from the group consisting of: brain cancer, cancers of the head and neck, lung cancer, breast cancer, cancers of the reproductive system, cancers of the digestive system, pancreatic cancer, and cancers of the urinary system. In further embodiments, said cancer of the digestive system is a cancer of the upper digestive tract or colorectal cancer. In further embodiments, said cancer of the urinary system is bladder cancer or renal cell carcinoma. In further embodiments, said cancer of the reproductive system is prostate cancer.
Additional types of cancers which may be treated using the compounds and methods described herein include: cancers of oral cavity and pharynx, cancers of the respiratory system, cancers of bones and joints, cancers of soft tissue, skin cancers, cancers of the genital system, cancers of the eye and orbit, cancers of the nervous system, cancers of the lymphatic system, and cancers of the endocrine system. In certain embodiments, these cancer s may beselected from the group consisting of: cancer of the tongue, mouth, pharynx, or other oral cavity; esophageal cancer, stomach cancer, or cancer of the small intestine; colon cancer or rectal, anal, or anorectal cancer; cancer of the liver, intrahepatic bile duct, gallbladder, pancreas, or other biliary or digestive organs; laryngeal, bronchial, and other cancers of the respiratory organs; heart cancer, melanoma, basal cell carcinoma, squamous cell carcinoma, other non-epithelial skin cancer; uterine or cervical cancer; uterine corpus cancer; ovarian, vulvar, vaginal, or other female genital cancer; prostate, testicular, penile or other male genital cancer; urinary bladder cancer; cancer of the kidney; renal, pelvic, or urethral cancer or other cancer of the genito-urinary organs; thyroid cancer or other endocrine cancer; chronic lymphocytic leukemia; and cutaneous T-cell lymphoma, both granulocytic and monocytic.
Yet other types of cancers which may be treated using the compounds and methods described herein include: adenocarcinoma, angiosarcoma, astrocytoma, acoustic neuroma, anaplastic astrocytoma, basal cell carcinoma, blastoglioma, chondrosarcoma, choriocarcinoma, chordoma, craniopharyngioma, cutaneous melanoma, cystadenocarcinoma, endotheliosarcoma, embryonal carcinoma, ependymoma, Ewing's tumor, epithelial carcinoma, fibrosarcoma, gastric cancer, genitourinary tract cancers, glioblastoma multiforme, hemangioblastoma, hepatocellular carcinoma, hepatoma, Kaposi's sarcoma, large cell carcinoma, leiomyosarcoma, liposarcoma,
lymphangiosarcoma, lymphangioendotheliosarcoma, medullary thyroid carcinoma,
medulloblastoma, meningioma mesothelioma, myelomas, myxosarcoma neuroblastoma, neurofibrosarcoma, oligodendroglioma, osteogenic sarcoma, epithelial ovarian cancer, papillary carcinoma, papillary adenocarcinomas, parathyroid tumors, pheochromocytoma, pinealoma, plasmacytomas, retinoblastoma, rhabdomyosarcoma, sebaceous gland carcinoma, seminoma, skin cancers, melanoma, small cell lung carcinoma, squamous cell carcinoma, sweat gland carcinoma, synovioma, thyroid cancer, uveal melanoma, and Wilm's tumor.
In some aspects of the invention, the disease to be treated by the methods of the present invention may be a hematologic disorder. In certain embodiments, said hematologic disorder is selected from the group consisting of sickle cell anemia, myelodysplastic disorders (MDS), and myeloproliferative disorders. In further embodiments, said myeloproliferative disorder is selected from the group consisting of polycythemia vera, myelofibrosis and essential thrombocythemia. In some aspects of the invention, the disease to be treated by the methods of the present invention may be a neurological disorder. In further embodiments, said neurological disorder is selected from the group consisting of epilepsy, neuropathic pain, depression and bipolar disorders.
In some aspects of the invention, the disease to be treated by the methods of the present invention may be a cardiovascular condition. In certain embodiments, said cardiovascular condition is selected from the group consisting of cardiac hypertrophy, idiopathic cardiomyopathies, and heart failure.
In some aspects of the invention, the disease to be treated by the methods of the present invention may be an autoimmune disease. In certain embodiments, said autoimmune disease is selected from the group consisting of systemic lupus erythromatosus (SLE), multiple sclerosis (MS), and systemic lupus nephritis.
In some aspects of the invention, the disease to be treated by the methods of the present invention may be a dermatologic disorder. In certain embodiments, said dermatologic disorder is selected from the group consisting of psoriasis, melanoma, basal cell carcinoma, squamous cell carcinoma, and other non-epithelial skin cancers.
In some aspects of the invention, the disease to be treated by the methods of the present invention may be an ophthalmologic disorder. Jh certain embodiments, said ophthalmologic disorder is selected from the group consisting of dry eye, closed angle glaucoma and wide angle glaucoma.
In some aspects of the invention, the disease to be treated by the methods of the present invention may be a polyglutamine-repeat disorders. In some embodiments, the polyglutamine- repeat disorder is selected from the group consisting of Huntington's disease, Spinocerebellar ataxia 1 (SCA 1), Machado-Joseph disease (MID)/Spinocerebella ataxia 3 (SCA 3), Kennedy disease/Spinal and bulbar muscular atrophy (SBMA) and Dentatorubral pallidolusyian atrophy (DRPLA).
In some aspects of the invention, the disease to be treated by the methods of the present invention may be an inflammatory condition. In some embodiments, the inflammatory condition is selected from the group consisting of Rheumatoid Arthritis (RA), Inflammatory Bowel Disease (IBD), ulcerative colitis and psoriasis.
In another aspect are compounds or compositions comprising compounds capable of inhibiting the catalytic or cellular activity of histone deacetylase (HDAC).
As used in the present specification, the following terms have the meanings indicated.
The term "acyl," as used herein, alone or in combination, refers to a carbonyl attached to an alkenyl, alkyl, aryl, cycloalkyl, heteroaryl, heterocycle, or any other moiety were the atom attached to the carbonyl is carbon. An "acetyl" group refers to a -C(O)CH3 group. An "alkylcarbonyl" or "alkanoyl" group refers to an alkyl group attached to the parent molecular moiety through a carbonyl group. Examples of such groups include methylcarbonyl and ethylcarbonyl. Examples of acyl groups include formyl, alkanoyl and aroyl. The term "alkenyl," as used herein, alone or in combination, refers to a straight-chain or branched-chain hydrocarbon radical having one or more double bonds and containing from 2 to 20, preferably 2 to 6, carbon atoms. Alkenylene refers to a carbon-carbon double bond system attached at two or more positions such as ethenylene [(-CH=CH-),(-C::C-)]. Examples of suitable alkenyl radicals include ethenyl, propenyl, 2-methylpropenyl, 1,4-butadienyl and the like.
The term "alkoxy," as used herein, alone or in combination, refers to an alkyl ether radical, wherein the term alkyl is as defined below. Examples of suitable alkyl ether radicals include methoxy, ethoxy, n-propoxy, isopropoxy, n-butoxy, iso-butoxy, sec-butoxy, tert-butoxy, and the like.
The term "alkyl," as used herein, alone or in combination, refers to a straight-chain or branched-chain alkyl radical containing from 1 to and including 20, preferably 1 to 10, and more preferably 1 to 6, carbon atoms. Alkyl groups may be optionally substituted as defined herein. Examples of alkyl radicals include methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, tert-butyl, pentyl, iso-amyl, hexyl, octyl, noyl and the like. The term "alkylene," as used herein, alone or in combination, refers to a saturated aliphatic group derived from a straight or branched chain saturated hydrocarbon attached at two or more positions, such as methylene (-CHr-).
The term "alkylamino," as used herein, alone or in combination, refers to an alkyl group attached to the parent molecular moiety through an amino group. Suitable alkylamino groups may be mono- or dialkylated, forming groups such as, for example, N-methylamino, N-ethylamino, N,N- dimethylamino, N,N-ethylmethylamino and the like.
The term "alkylidene," as used herein, alone or in combination, refers to an alkenyl group in which one carbon atom of the carbon-carbon double bond belongs to the moiety to which the alkenyl group is attached.
The term "alkylthio," as used herein, alone or in combination, refers to an alkyl thioether (R-S-) radical wherein the term alkyl is as defined above and wherein the sulfur may be singly or doubly oxidized. Examples of suitable alkyl thioether radicals include methylthio, ethylthio, n- propylthio, isopropylthio, n-butylthio, iso-butylthio, sec-butylthio, tert-butylthio, methanesulfonyl, ethanesulfinyl, and the like.
The term "alkynyl," as used herein, alone or in combination, refers to a straight-chain or branched chain hydrocarbon radical having one or more triple bonds and containing from 2 to 20, preferably from 2 to 6, more preferably from 2 to 4, carbon atoms. "Alkynylene" refers to a carbon- carbon triple bond attached at two positions such as ethynylene (-C:::C-, -OC-). Examples of alkynyl radicals include ethynyl, propynyl, hydroxypropynyl, butyn-1-yl, butyn-2-yl, pentyn-1-yl, 3- methylbutyn-1-yl, hexyn-2-yl, and the like.
The terms "amido" and "carbamoyl,"as used herein, alone or in combination, refer to an amino group as described below attached to the parent molecular moiety through a carbonyl group, or vice versa. The term "C-amido" as used herein, alone or in combination, refers to a -C(=O)-NR2 group with R as defined herein. The term "N-amido" as used herein, alone or in combination, refers to a RC(=0)NH- group, with R as defined herein. The term "acylamino" as used herein, alone or in combination, embraces an acyl group attached to the parent moiety through an amino group. An example of an "acylamino" group is acetylamino (CH3C(O)NH-).
The term "amino," as used herein, alone or in combination, refers to— NRR', wherein R and R are independently selected from the group consisting of hydrogen, alkyl, acyl, heteroalkyl, aryl, cycloalkyl, heteroaryl, and heterocycloalkyl, any of which may themselves be optionally substituted.
The term "aryl," as used herein, alone or in combination, means a carbocyclic aromatic system containing one, two or three rings wherein such rings may be attached together in a pendent manner or may be fused. The term "aryl" embraces aromatic radicals such as benzyl, phenyl, naphthyl, anthracenyl, phenanthryl, indanyl, indenyl, annulenyl, azulenyl, tetrahydronaphthyl, and biphenyl.
The term "arylalkenyl" or "aralkenyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkenyl group.
The term "arylalkoxy" or "aralkoxy," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkoxy group.
The term "arylalkyl" or "aralkyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkyl group.
The term "arylalkynyl" or "aralkynyl," as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an alkynyl group.
The term "arylalkanoyl" or "aralkanoyl" or "aroyl,"as used herein, alone or in combination, refers to an acyl radical derived from an aryl-substituted alkanecarboxylic acid such as benzoyl, napthoyl, phenylacetyl, 3-phenylpropionyl (hydrocinnamoyl), 4-phenylbutyryl, (2-naphthyl)acetyl, 4-chlorohydrocinnamoyl, and the like.
The term aryloxy as used herein, alone or in combination, refers to an aryl group attached to the parent molecular moiety through an oxy.
The terms "benzo" and "benz," as used herein, alone or in combination, refer to the divalent radical CeKt= derived from benzene. Examples include benzothiophene and benzimidazole.
The term "carbamate," as used herein, alone or in combination, refers to an ester of carbamic acid (-NHC00-) which may be attached to the parent molecular moiety from either the nitrogen or acid end, and which may be optionally substituted as defined herein.
The term "O-carbamyl" as used herein, alone or in combination, refers to a -OC(O)NRR', group-with R and R' as defined herein.
The term "N-carbamyl" as used herein, alone or in combination, refers to a ROC(O)NR'- group, with R and R' as defined herein.
The term "carbonyl," as used herein, when alone includes formyl [-C(O)H] and in combination is a -C(O)- group.
The term "carboxy," as used herein, refers to -C(O)OH or the corresponding "carboxylate" anion, such as is in a carboxylic acid salt. An "O-carboxy" group refers to a RC(O)O- group, where R is as defined herein. A "C-carboxy" group refers to a -C(O)OR groups where R is as defined herein. The term "cyano," as used herein, alone or in combination, refers to -CN.
The term "cycloalkyl," as used herein, alone or in combination, refers to a saturated or partially saturated monocyclic, bicyclic or tricyclic alkyl radical wherein each cyclic moiety contains from 3 to 12, preferably five to seven, carbon atom ring members and which may optionally be a benzo fused ring system which is optionally substituted as defined herein. Examples of such cycloalkyl radicals include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, octahydronaphthyl, 2,3-dihydro-lH-indenyl, adamantyl and the like. "Bicyclic" and "tricyclic" as used herein are intended to include both fused ring systems, such as decahydonapthalene, octahydronapthalene as well as the multicyclic (multicentered) saturated or partially unsaturated type. The latter type of isomer is exemplified in general by, bicyclo[l,l,l]pentane, camphor, adamantane, and bicyclo[3,2,l]octane.
The term "ester," as used herein, alone or in combination, refers to a carboxy group bridging two moieties linked at carbon atoms.
The term "ether," as used herein, alone or in combination, refers to an oxy group bridging two moieties linked at carbon atoms.
The term "halo," or "halogen," as used herein, alone or in combination, refers to fluorine, chlorine, bromine, or iodine.
The term "haloalkoxy," as used herein, alone or in combination, refers to a haloalkyl group attached to the parent molecular moiety through an oxygen atom.
The term "haloalkyl," as used herein, alone or in combination, refers to an alkyl radical having the meaning as defined above wherein one or more hydrogens are replaced with a halogen. Specifically embraced are monohaloalkyl, dihaloalkyl and polyhaloalkyl radicals. A monohaloalkyl radical, for one example, may have an iodo, bromo, chloro or fluoro atom within the radical. Dihalo and polyhaloalkyl radicals may have two or more of the same halo atoms or a combination of different halo radicals. Examples of haloalkyl radicals include fluoromethyl, difluoromethyl, trifluoromethyl, chloromethyl, dichloromethyl, trichloromethyl, pentafluoroethyl, heptafluoropropyl, difluorochloromethyl, dichlorofluoromethyl, difluoroethyl, difluoropropyl, dichloroethyl and dichloropropyl. "Haloalkylene" refers to a haloalkyl group attached at two or more positions. Examples include fluoromethylene
(-CFH-), difluoromethylene (-CF2 -), chloromethylene (-CHC1-) and the like.
The term "heteroalkyl," as used herein, alone or in combination, refers to a stable straight or branched chain, or cyclic hydrocarbon radical, or combinations thereof, fully saturated or containing from 1 to 3 degrees of unsaturation, consisting of the stated number of carbon atoms and from one to three heteroatoms selected from the group consisting of O, N, and S, and wherein the nitrogen and sulfur atoms may optionally be oxidized and the nitrogen heteroatom may optionally be quaternized. The heteroatom(s) O, N and S may be placed at any interior position of the heteroalkyl group. Up to two heteroatoms may be consecutive, such as, for example, -CH2-NH-OCH3.
The term "heteroaryl," as used herein, alone or in combination, refers to 3 to 7 membered, preferably 5 to 7 membered, unsaturated heteromonocyclic rings, or fused polycyclic rings in which at least one of the fused rings is unsaturated, wherein at least one atom is selected from the group consisting of O, S, and N. The term also embraces fused polycyclic groups wherein heterocyclic radicals are fused with aryl radicals, wherein heteroaryl radicals are fused with other heteroaryl radicals, or wherein heteroaryl radicals are fused with cycloalkyl radicals. Examples of heteroaryl groups include pyrrolyl, pyrrolinyl, imidazolyl, pyrazolyl, pyridyl, pyrimidinyl, pyrazinyl, pyridazinyl, triazolyl, pyranyl, furyl, thienyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, thiadiazolyl, isothiazolyl, indolyl, isoindolyl, indolizinyl, benzimidazolyl, qninolyl, isoquinolyl, quinoxalinyl, quinazolinyl, indazolyl, benzotriazolyl, benzodioxolyl, benzopyranyl, benzoxazolyl, benzoxadiazolyl, benzothiazolyl, benzothiadiazolyl, benzofuryl, benzothienyl, chromonyl, coumarinyl, benzopyranyl, tetrahydroquinolinyl, tetrazolopyridazinyl, tetrahydroisoquinolinyl, thienopyridinyl, furopyridinyl, pyrrolopyridinyl and the like. Exemplary tricyclic heterocyclic groupsinclude carbazolyl, benzidolyl, phenanthrolinyl, dibenzofuranyl, acridinyl, phenanthridinyl, xanthenyl and the like.
The terms "heterocycloalkyl" and, interchangeably, "heterocycle," as used herein, alone or in combination, each refer to a saturated, partially unsaturated, or fully unsaturated monocyclic, bicyclic, or tricyclic heterocyclic radical containing at least one, preferably 1 to 4, and more preferably 1 to 2 heteroatoms as ring members, wherein each said heteroatom may be independently selected from the group consisting of nitrogen, oxygen, and sulfur, and wherein there are preferably 3 to 8 ring members in each ring, more preferably 3 to 7 ring members in each ring, and most preferably 5 to 6 ring members in each ring. "Heterocycloalkyl" and "heterocycle" are intended to include sulfones, sulfoxides, N-oxides of tertiary nitrogen ring members, and carbocyclic fused and benzo fused ring systems; additionally, both terms also include systems where a heterocycle ring is fused to an aryl group, as defined herein, or an additional heterocycle group. Heterocycle groups of the invention are exemplified by aziridinyl, azetidinyl, 1,3 -benzodioxolyl, dihydroisoindolyl, dihydroisoquinolinyl, dihydrocinnolinyl, dihydrobenzodioxinyl, dihydro[l,3]oxazolo[4,5- b]pyridinyl, benzothiazolyl, dihydroindolyl, dihy-dropyridinyl, 1,3-dioxanyl, 1,4-dioxanyl, 1,3- dioxolanyl, isoindolinyl, morpholinyl, piperazinyl, pyrrolidinyl, tetrahydropyridinyl, piperidinyl, thiomorpholinyl, and the like. The heterocycle groups may be optionally substituted unless specifically prohibited.
The term "hydrazinyl" as used herein, alone or in combination, refers to two amino groups joined by a single bond, i.e., -N-N-.
The term "hydroxy," as used herein, alone or in combination, refers to -OH.
The term "hydroxyalkyl," as used herein, alone or in combination, refers to a hydroxy group attached to the parent molecular moiety through an alkyl group.
The term "imino," as used herein, alone or in combination, refers to =N-.
The term "iminohydroxy," as used herein, alone or in combination, refers to =N(0H) and
=N-O-.
The phrase "in the main chain" refers to the longest contiguous or adjacent chain of carbon atoms starting at the point of attachment of a group to the compounds of this invention.
The term "isocyanato" refers to a -NCO group.
The term "isothiocyanato" refers to a -NCS group. The phrase "linear chain of atoms" refers to the longest straight chain of atoms independently selected from carbon, nitrogen, oxygen and sulfur.
The term "lower," as used herein, alone or in combination, means containing from 1 to and including 6 carbon atoms.
The term "mercaptyl" as used herein, alone or in combination, refers to an RS- group, where R is as defined herein.
The term "nitro," as used herein, alone or in combination, refers to -NO2.
The terms "oxy" or "oxa," as used herein, alone or in combination, refer to -Q-.
The term "oxo," as used herein, alone or in combination, refers to =0.
The term "perhaloalkoxy" refers to an alkoxy group where all of the hydrogen atoms are replaced by halogen atoms.
The term "perhaloalkyl" as used herein, alone or in combination, refers to an alkyl group where all of the hydrogen atoms are replaced by halogen atoms.
The terms "sulfonate," "sulfonic acid," and "sulfonic," as used herein, alone or in combination, refer the -SO3H group and its anion as the sulfonic acid is used in salt formation.
The term "sulfanyl," as used herein, alone or in combination, refers to -S-.
The term "sulfinyl," as used herein, alone or in combination, refers to -S(O)-.
The term "sulfonyl," as used herein, alone or in combination, refers to -SO2-.
The term "N-sulfonamide" refers to a -S(=O)2NR- group with R as defined herein.
The term "S-sulfonamide" refers to a -NRS(=O)2-, group, with R as defined herein.
The terms "thia" and "thio," as used herein, alone or in combination, refer to a -S- group or an ether wherein the oxygen is replaced with sulfur. The oxidized derivatives of the thio group, namely sulfinyl and sulfonyl, are included in the definition of thia and thio.
The term "thiol," as used herein, alone or in combination, refers to an -SH group.
The term "thiocarbonyl," as used herein, when alone includes thioformyl -C(S)H and in combination is a -C(S)- group.
The term "N-thiocarbamyl" refers to an ROC(S)NR'- group, with R and R'as defined herein.
The term "O-thiocarbamyl" refers to a -OC(S)NRR', group with R and R'as defined herein.
The term "thiocyanato" refers to a -CNS group.
The term "trihalomethanesulfonamido" refers to a X3CS(O)2NR- group with X is a halogen and R as defined herein.
The term "trihalomethanesulfonyl" refers to a X3CS(O)2- group where X is a halogen. The term "trihalomethoxy" refers to a X3CO- group where X is a halogen.
The term "trisubstituted silyl," as used herein, alone or in combination, refers to a silicone group substituted at its three free valences with groups as listed herein under the definition of substituted amino. Examples include trimethysilyl, tert-butyldimethylsilyl, triphenylsilyl and the like. Any definition herein may be used in combination with any other definition to describe a composite structural group. By convention, the trailing element of any such definition is that which attaches to the parent moiety. For example, the composite group alkylamido would represent an alkyl group attached to the parent molecule through an amido group, and the term alkoxyalkyl would represent an alkoxy group attached to the parent molecule through an alkyl group.
When a group is defined to be "null," what is meant is that said group is absent.
The term "optionally substituted" means the anteceding group may be substituted or unsubstituted. When substituted, the substituents of an "optionally substituted" group may include, without limitation, one or more substituents independently selected from the following groups or a particular designated set of groups, alone or in combination: lower alkyl, lower alkenyl, lower alkynyl, lower alkanoyl, lower heteroalkyl, lower heterocycloalkyl, lower haloalkyl, lower haloalkenyl, lower haloalkynyl, lower perhaloalkyl, lower perhaloalkoxy, lower cycloalkyl, phenyl, aryl, aryloxy, lower alkoxy, lower haloalkoxy, oxo, lower acyloxy, carbonyl, carboxyl, lower alkylcarbonyl, lower carboxyester, lower carboxamido, cyano, hydrogen, halogen, hydroxy, amino, lower alkylamino, arylamino, amido, nitro, thiol, lower alkylthio, arylthio, lower alkylsulfinyl, lower alkylsulfonyl, arylsulfinyl, arylsulfonyl, arylthio, sulfonate, sulfonic acid, trisubstituted silyl, N3, SH, SCH3, C(O)CH3, COaCH3, CO2H, pyridinyl, thiophene, furanyl, lower carbamate, and lower urea. Two substituents may be joined together to form a fused five-, six-, or seven-menbered carbocyclic or heterocyclic ring consisting of zero to three heteroatoms, for example forming methylenedioxy or ethylenedioxy. An optionally substituted group may be unsubstituted (e.g., -CH2CH3), fully substituted (e.g., -CF2CF3), monosubstituted (e.g., -CH2CH2F) or substituted at a level anywhere in- between fully substituted and monosubstituted (e.g., -CH2CF3). Where substituents are recited without qualification as to substitution, both substituted and unsubstituted forms are encompassed. Where a substituent is qualified as "substituted," the substituted form is specifically intended. Additionally, different sets of optional substituents to a particuar moiety may be defined as needed; in these cases, the optional substitution will be as defined, often immediately following the phrase, "optionally substituted with."
The term R or the term R', appearing by itself and without a number designation, unless otherwise defined, refers to a moiety selected from the group consisting of null, hydrogen, alkyl, cycloalkyl, heteroalkyl, aryl, heteroaryl and heterocycloalkyl, any of which may be optionally substituted. Such R and R' groups should be understood to be optionally substituted as defined herein. Whether an R group has a number designation or not, every R group, including R, R' and Rn where n=(l, 2, 3, ...n), every substituent, and every term should be understood to be independent of every other in terms of selection from a group. Should any variable, substituent, or term (e.g. aryl, heterocycle, R, etc.) occur more than one time in a formula or generic structure, its definition at each occurrence is independent of the definition at every other occurrence. Those of skill in the art will further recognize that certain groups may be attached to a parent molecule or may occupy a position in a chain of elements from either end as written. Thus, by way of example only, an unsymmetrical group such as -C(O)N(R)- may be attached to the parent moiety at either the carbon or the nitrogen. Asymmetric centers exist in the compounds of the present invention. These centers are designated by the symbols "R" or "S," depending on the configuration of substituents around the chiral carbon atom. It should be understood that the invention encompasses all stereochemical isomeric forms, including diastereomeric, enantiomeric, and epimeric forms, as well as d-isomers and 1-isomers, and mixtures thereof. Individual stereoisomers of compounds can be prepared synthetically from commercially available starting materials which contain chiral centers or by preparation of mixtures of enantiomeric products followed by separation such as conversion to a mixture of diastereomers followed by separation or recrystallization, chromatographic techniques, direct separation of enantiomers on chiral chromatographic columns, or any other appropriate method known in the art. Starting compounds of particular stereochemistry are either commercially available or can be made and resolved by techniques known in the art. Additionally, the compounds of the present invention may exist as geometric isomers. The present invention includes all cis, trans, syn, anti, entgegen (E), and zusammen (Z) isomers as well as the appropriate mixtures thereof. Additionally, compounds may exist as tautomers; all tautomeric isomers are provided by this invention. Additionally, the compounds of the present invention can exist in unsolvated as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like. In general, the solvated forms are considered equivalent to the unsolvated forms for the purposes of the present invention.
The term "bond" refers to a covalent linkage between two atoms, or two moieties when the atoms joined by the bond are considered to be part of larger substructure. A bond may be single, double, or triple unless otherwise specified. A dashed line between two atoms in a drawing of a molecule indicates that an additional bond may be present or absent at that position.
The term "combination therapy" means the administration of two or more therapeutic agents to treat a therapeutic condition or disorder described in the present disclosure. Such administration encompasses co-administration of these therapeutic agents in a substantially simultaneous manner, such as in a single capsule having a fixed ratio of active ingredients or in multiple, separate capsules for each active ingredient. In addition, such administration also encompasses use of each type of therapeutic agent in a sequential manner. In either case, the treatment regimen will provide beneficial effects of the drug combination in treating the conditions or disorders described herein.
The terms "therapy" or "treating" as used herein refer to (1) reducing the rate of progress of a disease, or, in case of cancer reducing the size of the tumor; (2) inhibiting to some extent further progress of the disease, which in case of cancer may mean slowing to some extent, or preferably stopping, tumor metastasis or tumor growth; and/or, (3) relieving to some extent (or, preferably, eliminating) one or more symptoms associated with the disease. Thus, the term "therapeutically effective amount" as used herein refers to that amount of the compound being administered which will provide therapy or affect treatment.
The term "patient" means all mammals including humans. Examples of patients include humans, cows, dogs, cats, goats, sheep, pigs, and rabbits. Preferably, the patient is a human. In some aspects of the invention, the compounds of the present invention are also antitumor compounds and/or inhibit the growth of a tumor, i.e., they are tumor-growth-inhibiting compounds. The terms "anti-tumor" and "tumor-growth-inhibiting," when modifying the term "compound," and the terms "inhibiting" and "reducing", when modifying the terms "compound" and/or "tumor," mean that the presence of the subject compound is correlated with at least the slowing of the rate of growth of the tumor. More preferably, the terms "anti-tumor," "tπmor-growth- inhibiting," "inhibiting," and "reducing" refer to a correlation between the presence of the subject compound and at least the temporary cessation of tumor growth. The terms "anti-tumor," "tumor- growth-inhibiting," "inhibiting," and "reducing" also refer to, a correlation between the presence of the subject compound and at least the temporary reduction in the mass of the tumor.
The term "function" refers to the cellular role of HDAC. The term "catalytic activity", in the context of the invention, defines the rate at which HDAC deacetylates a substrate. Catalytic activity can be measured, for example, by determining the amount of a substrate converted to a product as a function of time. Deacetylation of a substrate occurs at the active-site of HDAC. The active-site is normally a cavity in which the substrate binds to HDAC and is deacetylated.
The term "substrate" as used herein refers to a molecule deacetylated by HDAC. The substrate is preferably a peptide and more preferably a protein. In some embodiments, the protein is a histone, whereas in other embodiments, the protein is not a histone.
The term "inhibit" refers to decreasing the cellular function of HDAC. It is understood that compounds of the present invention may inhibit the cellular function of HDAC by various direct or indirect mechanisms, in particular by direct or indirect inhibition of the catalytic activityof HDAC. The term "activates" refers to increasing the cellular function of HDAC.
The term "activate" refers to increasing the cellular function of HDAC.
HDAC function is preferably the interaction with a natural binding partner and most preferably catalytic activity.
The term "modulate" refers to altering the function of HDAC by increasing or decreasing the probability that a complex forms between HDAC and a natural binding partner. A modulator may increase the probability that such a complex forms between HDAC and the natural binding partner, or may increase or decrease the probability that a complex forms between HDAC and the natural binding partner depending on the concentration of the compound exposed to HDAC, or may decrease the probability that a complex forms between HDAC and the natural binding partner. A modulator may activate the catalytic activity of HDAC, or may activate or inhibit the catalytic activity of HDAC depending on the concentration of the compound exposed to HDAC, or may inhibit the catalytic activity of HDAC.
The term "complex" refers to an assembly of at least two molecules bound to one another.
The term "natural binding partner" refers to polypeptides that bind to HDAC in cells. A change in the interaction between HDAC and a natural binding partner can manifest itself as an increased or decreased probability that the interaction forms, or an increased or decreased concentration of HDAC/natural binding partner complex. The term "contacting" as used herein refers to mixing a solution comprising a compound of the invention with a liquid medium bathing the cells of the methods. The solution comprising the compound may also comprise another component, such as dimethylsulfoxide (DMSO), which facilitates the uptake of the compound or compounds into the cells of the methods. The solution comprising the compound of the invention may be added to the medium bathing the cells by utilizing a delivery apparatus, such as a pipet-based device or syringe-based device.
The term "monitoring" refers to observing the effect of adding the compound to the cells of the method. The effect can be manifested in a change in cell phenotype, cell proliferation, HDAC catalytic activity, substrate protein acetylation levels, gene expression changes, or in the interaction between HDAC and a natural binding partner.
The term "effect" describes a change or an absence of a change in cell phenotype or cell proliferation. "Effect" can also describe a change or an absence of a change in the catalytic activity of HDAC. "Effect" can also describe a change or an absence of a change in an interaction between HDAC and a natural binding partner.
The term "cell phenotype" refers to the outward appearance of a cell or tissue or the function of the cell or tissue. Examples of cell phenotype are cell size (reduction or enlargement), cell proliferation (increased or decreased numbers of cells), cell differentiation (a change or absence of a change in cell shape), cell survival, apoptosis (cell death), or the utilization of a metabolic nutrient (e.g., glucose uptake). Changes or the absence of changes in cell phenotype are readily measured by techniques known in the art.
"HDAC inhibitor" is used herein to refer to a compound that exhibits an IC50 with respect to HDAC activity of no more than about lOOμ.M and more typically not more than about 50 μM, as measured in the biochemical in vitro HDAC-inhibition assay, cellular histone hyperacetylation assay, and differential cytotoxicity assay described generally herein below. "IC50" is that concentration of inhibitor which reduces the activity of an enzyme (e.g., HDAC) to half-maximal level. Representative compounds of the present invention have been discovered to exhibit inhibitory activity against HDAC. Compounds of the present invention preferably exhibit an IC50 with respect to HDAC of no more than about 10 μM, more preferably, no more than about 5 μM, even more preferably not more than about 1 μM, and most preferably, not more than about 200 nM, as measured in the HDAC assays described herein.
The term "prodrug" refers to a compound that is made more active in vivo. Certain compounds of the present invention may also exist as prodrugs, as described in Hydrolysis in Drug and Prodrug Metabolism Chemistry, Biochemistry, and Enzymology (Testa, Bernard and Mayer, Joachim M. Wiley-VHCA, Zurich, Switzerland 2003). Prodrugs of the compounds described herein are structurally modified forms of the compound that readily undergo chemical changes under physiological conditions to provide the compound. Additionally, prodrugs can be converted to the compound by chemical or biochemical methods in an ex vivo environment. For example, prodrugs can be slowly converted to a compound when placed in a transdermal patch reservoir with a suitable enzyme or chemical reagent. Prodrugs are often useful because, in some situations, they may be easier to administer than the compound, or parent drug. They may, for instance, be bioavailable by oral administration whereas the parent drug is not. The prodrug may also have improved solubility in pharmaceutical compositions over the parent drug. A wide variety of prodrug derivatives are known in the art, such as those that rely on hydrolytic cleavage or oxidative activation of the prodrug. An example, without limitation, of a prodrug would be a compound which is administered as an ester (the "prodrug"), but then is metabolically hydrolyzed to the carboxylic acid, the active entity.
Additional examples include peptidyl derivatives of a compound. Yet another example of a prodrug is protected thiol compounds. Thiols bearing hydrolyzable protecting groups can unmask protected SH groups prior to or simultaneous to use. As shown below, the moiety -C(O)-RE of a thioester may be hydrolyzed to yield a thiol and a pharmaceutically acceptable acid HO-C(O)-RE.
* r s & RE + H2° *" \ I"SH HO SL RE
The term "therapeutically acceptable prodrug," refers to those prodrugs or zwitterions which are suitable for use in contact with the tissues of patients without undue toxicity, irritation, and allergic response, are commensurate with a reasonable benefit/risk ratio, and are effective for their intended use.
A "pharmaceutically active metabolite" is intended to mean a pharmacologically active product produced through metabolism in the body of a specified compound or salt thereof.
Metabolites of a compound may be identified using routine techniques known in the art and their activities determined using tests such as those decribed herein.
The term "thiol protecting group" refers to thiols bearing hydrolyzable protecting groups that can unmask protected SH groups prior to or simultaneous to use. Preferred thiol protecting groups include but are not limited to thiol esters which release pharmaceutically acceptable acids along with an active thiol moiety. Such pharmaceutically acceptable acids are generally nontoxic and do not abrogate the biological activity of the active thiol moiety. Examples of pharmaceutically acceptable acids include, but are not limited to: N,N-diethylglycine; 4-ethylpiperazinoacetic acid; ethyl 2-methoxy-2-phenylacetic acid; N,N-dimethylglycine; (nitrophenoxysulfonyl)benzoic acid; acetic acid; maleic acid; fumaric acid; benzoic acid; tartaric acid; natural amino acids (like glutamate, aspartate, cyclic amino acids such proline); D-amino acids; butyric acid; fatty acids like palmitic acid, stearic acid, oleate; pipecolic acid; phosphonic acid; phosphoric acid; pivalate (trimethylacetic acid); succinic acid; cinnamic acid; anthranilic acid; salicylic acid; lactic acid; and pyruvic acids.
Another aspect of the present invention are compounds containing at least one thiol in a protected form, which can be released to provide a SH group prior to or simultaneous to use. Thiol moieties are known to be unstable in the presence of air and are oxidized to the corresponding disulfide. Protected thiol groups are those that can be converted under mild conditions into free thiol groups without other undesired side reactions taking place. Suitable thiol protecting groups include but are not limited to trityl (Trt), allyloxycarbonyl (Alloc), l-(4,4-dimethyl-2,6-dioxocyclohex-l- ylidene)ethyl (Dde), acetamidomethyl (Acm), t-buryl (tBu), or the like. Preferred thiol protecting groups include lower alkanoyl, e.g. acetyl. Free thiol, disulfides, and protected thiols are understood to be within the scope of this invention. The term "therapeutically acceptable salt," as used herein, represents salts or zwitterionic forms of the compounds of the present invention which are water or oil-soluble or dispersible; which are suitable for treatment of diseases without undue toxicity, irritation, and allergic-response; which are commensurate with a reasonable benefit/risk ratio; and which are effective for their intended use. The salts can be prepared during the final isolation and purification of the compounds or separately by reacting the appropriate compound in the form of the free base with a suitable acid.
Representative acid addition salts include acetate, adipate, alginate, L-ascorbate, aspartate, benzoate, benzenesulfonate (besylate), bisulfate, butyrate, camphorate, camphorsulfonate, citrate, digluconate, formate, fumarate, gentisate, glutarate, glycerophosphate, glycolate, hemisulfate, heptanoate, hexanoate, hippurate, hydrochloride, hydrobromide, hydroiodide, 2- hydroxyethansulfonate (isethionate), lactate, maleate, malate, malonate, DL-mandelate, mesitylenesulfonate, methanesulfonate, naphthylenesulfonate, nicotinate, 2-naρhthalenesulfonate, oxalate, pamoate, pectinate, persulfate, 3-phenylproprionate, phosphonate, picrate, pivalate, propionate, pyroglutamate, succinate, sulfonate, tartrate, L-tartrate, trichloroacetate, trifluoroacetate, phosphate, glutamate, bicarbonate, para-toluenesulfonate (p-tosylate), and undecanoate. Also, basic groups in the compounds of the present invention can be quaternized with methyl, ethyl, propyl, and butyl chlorides, bromides, and iodides; dimethyl, diethyl, dibulyl, and diamyl sulfates; decyl, lauryl, myristyl, and steryl chlorides, bromides, and iodides; and benzyl and phenethyl bromides. Examples of acids which can be employed to form therapeutically acceptable addition salts include inorganic acids such as hydrochloric, hydrobromic, sulfuric, and phosphoric, and organic acids such as oxalic, maleic, succinic, and citric. Salts can also be formed by coordination of the compounds with an alkali metal or alkaline earth ion. Hence, the present invention contemplates sodium, potassium, magnesium, and calcium salts of the compounds of the compounds of the present invention and the like.
Basic addition salts can be prepared during the final isolation and purification of the compounds by reacting a carboxy group with a suitable base such as the hydroxide, carbonate, or bicarbonate of a metal cation or with ammonia or an organic primary, secondary, or tertiary amine. The cations of therapeutically acceptable salts include lithium, sodium, potassium, calcium, magnesium, and aluminum, as well as nontoxic quaternary amine cations such as ammonium, tetramethylammonium, tetraethylammonium, methylamine, dimethylamine, trimethylamine, triethylamine, diethylamine, ethylamine, tributylamine, pyridine, iV,iV-dimethylaniline, N- methylpiperidine, iV-methylmorpholine, dicyclohexylamine, procaine, dibenzylamine, N,N- dibenzylphenethylamine, 1-ephenamine, andΛζiV-dibenzylethylenediamine. Other representative organic amines useful for the formation of base addition salts include ethylenediamine, ethanolamine, diethanolamine, piperidine, and piperazine.
The compounds of the present invention can exist as therapeutically acceptable salts. The present invention includes compounds listed above in the form of salts, in particular acid addition salts. Suitable salts include those formed with both organic and inorganic acids. Such acid addition salts will normally be pharmaceutically acceptable. However, salts of non-pharmaceutically acceptable salts may be of utility in the preparation and purification of the compound in question. While it may be possible for the compounds of the subject invention to be administered as the raw chemical, it is also possible to present them as a pharmaceutical formulation. Accordingly, the subject invention provides a pharmaceutical formulation comprising a compound or a pharmaceutically acceptable salt, ester, prodrug or solvate thereof, together with one or more pharmaceutically acceptable carriers thereof and optionally one or more other therapeutic ingredients. The carrier(s) must be "acceptable" in the sense of being compatible with the other ingredients of the formulation and not deleterious to the recipient thereof. Proper formulation is dependent upon the route of administration chosen. Any of the well-known techniques, carriers, and excipients may be used as suitable and as understood in the art; e.g., in Remington's Pharmaceutical Sciences. The pharmaceutical compositions of the present invention may be manufactured in a manner that is itself known, e.g., by means of conventional mixing, dissolving, granulating, dragee- making, levigating, emulsifying, encapsulating, entrapping or compression processes.
The formulations include those suitable for oral, parenteral (including subcutaneous, intradermal, intramuscular, intravenous, intraarticular, and intramedullary), intraperitoneal, transmucosal, transdermal, rectal and topical (including dermal, buccal, sublingual and intraocular) administration although the most suitable route may depend upon for example the condition and disorder of the recipient. The formulations may conveniently be presented in unit dosage form and may be prepared by any of the methods well known in the art of pharmacy. AU methods include the step of bringing into association a compound of the subject invention or a pharmaceutically acceptable salt, ester, prodrug or solvate thereof ("active ingredient") with the carrier which constitutes one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately bringing into association the active ingredient with liquid carriers or finely divided solid carriers or both and then, if necessary, shaping the product into the desired formulation.
Formulations of the present invention suitable for oral administration may be presented as discrete units such as capsules, cachets or tablets each containing a predetermined amount of the active ingredient; as a powder or granules; as a solution or a suspension in an aqueous liquid or a non-aqueous liquid; or as an oil-in-water liquid emulsion or a water-in-oil liquid emulsion. The active ingredient may also be presented as a bolus, electuary or paste.
Pharmaceutical preparations which can be used orally include tablets, push-fit capsules made of gelatin, as well as soft, sealed capsules made of gelatin and a plasticizer, such as glycerol or sorbitol. Tablets may be made by compression or molding, optionally with one or more accessory ingredients. Compressed tablets may be prepared by compressing in a suitable machine the active ingredient in a free-flowing form such as a powder or granules, optionally mixed with binders, inert diluents, or lubricating, surface active or dispersing agents. Molded tablets may be made by molding in a suitable machine a mixture of the powdered compound moistened with an inert liquid diluent. The tablets may optionally be coated or scored and may be formulated so as to provide slow or controlled release of the active ingredient therein. All formulations for oral administration should be in dosages suitable for such administration. The push-fit capsules can contain the active ingredients in admixture with filler such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers. In soft capsules, the active compounds may be dissolved or suspended in suitable liquids, such as fatty oils, liquid paraffin, or liquid polyethylene glycols. In addition, stabilizers may be added. Dragee cores are provided with suitable coatings. For this purpose, concentrated sugar solutions may be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, carbopol gel, polyethylene glycol, and/or titanium dioxide, lacquer solutions, and suitable organic solvents or solvent mixtures. Dyestuffs or pigments may be added to the tablets or dragee coatings for identification or to characterize different combinations of active compound doses.
The compounds may be formulated for parenteral administration by injection, e.g., by bolus injection or continuous infusion. Formulations for injection may be presented in unit dosage form, e.g., in ampoules or in multi-dose containers, with an added preservative. The compositions may take such forms as suspensions, solutions or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. The formulations may be presented in unit-dose or multi-dose containers, for example sealed ampoules and vials, and may be stored in powder form or in a freeze-dried (lyophilized) condition requiring only the addition of the sterile liquid carrier, for example, saline or sterile pyrogen-free water, immediately prior to use. Extemporaneous injection solutions and suspensions may be prepared from sterile powders, granules and tablets of the kind previously described.
Formulations for parenteral administration include aqueous and non-aqueous (oily) sterile injection solutions of the active compounds which may contain antioxidants, buffers, bacteriostats and solutes which render the formulation isotonic with the blood of the intended recipient; and aqueous and non-aqueous sterile suspensions which may include suspending agents and thickening agents. Suitable lipophilic solvents or vehicles include fatty oils such as sesame oil, or synthetic fatty acid esters, such as ethyl oleate or triglycerides, or liposomes. Aqueous injection suspensions may contain substances which increase the viscosity of the suspension, such as sodium carboxymethyl cellulose, sorbitol, or dextran. Optionally, the suspension may also contain suitable stabilizers or agents which increase the solubility of the compounds to allow for the preparation of highly concentrated solutions.
In addition to the formulations described previously, the compounds may also be formulated as a depot preparation. Such long acting formulations may be administered by implantation (for example subcutaneously or intramuscularly) or by intramuscular injection. Thus, for example, the compounds may be formulated with suitable polymeric or hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, or as sparingly soluble derivatives, for example, as a sparingly soluble salt.
For buccal or sublingual administration, the compositions may take the form of tablets, lozenges, pastilles, or gels formulated in conventional manner. Such compositions may comprise the active ingredient in a flavored basis such as sucrose and acacia or tragacanth.
The compounds may also be formulated in rectal compositions such as suppositories or retention enemas, e.g., containing conventional suppository bases such as cocoa butter, polyethylene glycol, or other glycerides. Compounds of the present invention may be administered topically, that is by non- systemic administration. This includes the application of a compound of the present invention externally to the epidermis or the buccal cavity and the instillation of such a compound into the ear, eye and nose, such that the compound does not significantly enter the blood stream. In contrast, systemic administration refers to oral, intravenous, intraperitoneal and intramuscular administration.
Formulations suitable for topical administration include liquid or semi-liquid preparations suitable for penetration through the skin to the site of inflammation such as gels, liniments, lotions, creams, ointments or pastes, and drops suitable for administration to the eye, ear or nose. The active ingredient may comprise, for topical administration, from 0.001% to 10% w/w, for instance from 1% to 2% by weight of the formulation. It may however comprise as much as 10% w/w but preferably will comprise less than 5% w/w, more preferably from 0.1% to 1% w/w of the formulation.
For administration by inhalation the compounds according to the invention are conveniently delivered from an insufflator, nebulizer pressurized packs or other convenient means of delivering an aerosol spray. Pressurized packs may comprise a suitable propellant such as
dichlorodifluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gas. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve to deliver a metered amount. Alternatively, for administration by inhalation or insufflation, the compounds according to the invention may take the form of a dry powder composition, for example a powder mix of the compound and a suitable powder base such as lactose or starch. The powder composition may be presented in unit dosage form, in for example, capsules, cartridges, gelatin or blister packs from which the powder may be administered with the aid of an inhalator or insufflator.
In certain embodiments, pharmaceutical preparations of compound(s) or active ingredient(s) of the present invention may be formulated by Latitude Pharmaceuticals Inc. located in 9865 Mesa Rim Road, STE 201, San Diego, CA 92121 using their trade secret and proprietary formulation named "FlOl". The composition of said formulation FlOl is known to contain triglyceride, soy lecithin, vitamin E and PEG400.
Preferred unit dosage formulations are those containing an effective dose, as herein below recited, or an appropriate fraction thereof, of the active ingredient.
It should be understood that in addition to the ingredients particularly mentioned above, the formulations of this invention may include other agents conventional in the art having regard to the type of formulation in question, for example those suitable for oral administration may include flavoring agents.
The compounds of the invention may be administered orally or via injection at a dose of from 0.1 to 500 mg/kg per day. The dose range for adult humans is generally from 5 mg to 2 g/day. Tablets or other forms of presentation provided in discrete units may conveniently contain an amount of compound of the invention which is effective at such dosage or as a multiple of the same, for instance, units containing 5 mg to 500 mg, usually around 10 mg to 200 mg. The amount of active ingredient that may be combined with the carrier materials to produce a single dosage form will vary depending upon the host treated and the particular mode of administration.
The compounds of the subject invention can be administered in various modes, e.g. orally, topically, or by injection. The precise amount of compound administered to a patient will be the responsibility of the attendant physician. The specific dose level for any particular patient will depend upon a variety of factors including the activity of the specific compound employed, the age, body weight, general health, sex, diets, time of administration, route of administration, rate of excretion, drug combination, the precise disorder being treated, and the severity of the indication or condition being treated. Also, the route of administration may vary depending on the condition and its severity.
In certain instances, it may be appropriate to administer at least one of the compounds described herein (or a pharmaceutically acceptable salt, ester, or prodrug thereof) in combination with another therapeutic agent. By way of example only, if one of the side effects experienced by a patient upon receiving one of the compounds herein is hypertension, then it may be appropriate to administer an anti-hypertensive agent in combination with the initial therapeutic agent. Or, by way of example only, the therapeutic effectiveness of one of the compounds described herein may be enhanced by administration of an adjuvant (i.e., by itself the adjuvant may only have minimal therapeutic benefit, but in combination with another therapeutic agent, the overall therapeutic benefit to the patient is enhanced). Or, by way of example only, the benefit of experienced by a patient may be increased by administering one of the compounds described herein with another therapeutic agent (which also includes a therapeutic regimen) that also has therapeutic benefit. By way of example only, in a treatment for cancer involving administration of one of the compounds described herein, increased therapeutic benefit may result by also providing the patient with another therapeutic agent for cancer. In any case, regardless of the disease, disorder, or condition being treated, the overall benefit experienced by the patient may simply be additive of the two therapeutic agents or the patient may experience a synergistic benefit.
Specific, non-limiting examples of possible combination therapies include use of the compounds of the invention with another chemotherapeutic agent such as aromatase inhibitors, antiestrogen, anti-androgen, or a gonadorelin agonists, topoisomerase 1 and 2 inhibitors, microtubule active agents, alkylating agents, antimeoplastic antimetabolite, or platin containing compound, lipid or protein kinase targeting agents, protein or lipid phosphatase targeting agents, anti-angiogentic agents, agents that induce cell differentiation, bradykinin 1 receptor and angiotensin II antagonists, cyclooxygenase inhibitors, heparanase inhibitors, lymphokines or cytokine inhibitors, bisphosphanates, rapamycin derivatives, anti-apoptotic pathway inhibitors, apoptotic pathway agonists, PPAR agonists, inhibitors of Ras isoforms, telomerase inhibitors, protease inhibitors, metalloproteinase inhibitors, aminopeptidase inhibitors, and biologic drugs including but not limited to antibodies, cytokines and growth factors.
In some aspects of the invention, the chemotherapeutic agents that are useful for the treatment of multiple myeloma include, but are not limited to, alkylating agents (eg, melphalan), anthracyclines (eg. doxorubicin), corticosteroids (eg. dexamethasome), IMiDs (eg. thalidomide, lenalidomide), protease inhibitors (eg. bortezomib, NPI0052), IGF-I inhibitors, CD40 antibodies, Smac mimetics (eg. telomestatin), FGF3 modulator (eg. CHIR258), mTOR inhibitor (Rad 001), HDAC inhibitors (eg. SAHA, Tubacin), IKX inhibitors, P38MAPK inhibitors, HSP90 inhibitors (eg 17-AAG), and akt inhibitors (eg. Perifosine).
Further, the preferred chemotherapeiitic agents used in combination with the compounds of the present invention include without limitation melphalan, doxorubicin (including lyophilized), dexamethasone, prednisone, thalidomide, lenalidomide, bortezomib, andNPI0052.
In any case, the multiple chemotherapeutic agents (at least one of which is a compound of the present invention) may be administered in any order or even simultaneously. If simultaneously, the multiple chemotherapeutic agents may be provided in a single, unified form, or in multiple forms (by way of example only, either as a single pill or as two separate pills). One of the
chemotherapeutic agents may be given in multiple doses, or both may be given as multiple doses. If not simultaneous, the timing between the multiple doses may be any duration of time ranging from a few minutes to four weeks.
Thus, in another aspect, the present invention provides methods for treating HDAC- mediated disorders in a human or animal subject in need of such treatment comprising administering to said subject an amount of a compound of the present invention effective to reduce or prevent said disorder in the subject in combination with at least one additional agent for the treatment of said disorder that is known in the art. In a related aspect, the present invention provides therapeutic compositions comprising at least one compound of the present invention in combination with one or more additional agents for the treatment of HDAC -mediated disorders.
The present invention includes compounds listed above in the form of salts, in particular acid addition salts. Suitable salts include those formed with both organic and inorganic acids. Such acid addition salts will normally be pharmaceutically acceptable. However, salts of non- pharmaceutically acceptable salts may be of utility in the preparation and purification of the compound in question.
Thus, preferred salts include those formed from hydrochloric, hydrobromic, sulphuric, citric, tartaric, phosphoric, lactic, pyruvic, acetic, succinic, oxalic, fumaric, maleic, oxaloacetic, methanesulphonic, ethanesulphonic, p-toluenesulphonic, benzenesulphonic and isethionic acids. A salt of a compound can be made by reacting the appropriate compound in the form of the free base with the appropriate acid.
All references, patents or applications, U.S. or foreign, cited in the application are hereby incorporated by reference as if written herein. GENERAL SYNTHETIC METHODS FOR PREPARING COMPOT JNDS
Molecular embodiments of the present invention can be synthesized using standard synthetic techniques known to those of skill in the art. Compounds of the present invention can be synthesized using the general synthetic procedures set forth in Schemes I-VI.
Figure imgf000024_0001
Reagents: (a) Acetic anhydride, Et3N, DMAP, CH2Cl2, room temperature, 2h. (b) Bromine,
Acetic Acid, room temperature, 45 min. (c) Chlorosulfonic acid, 70 C, 18h. (d) 4- aminoacetophenone, pyridine, 50 C, 18h. (e) H2, Pd(C), Et3N, DMF, 18h. (f) trimethylphenylammonium tribromide, THF, 40 C, 3h. (g) HCl, 1,4-dioxane, 100 C, Ih. (h) DDQ, acetonitrile, room temperature, Ih. (i) Potassium thioacetate, MeOH, room temperature, 18h. Scheme II
Figure imgf000025_0001
Reagents: (a) H2, Raney Nickel, toluene, (b) 1] NaNO2, HCl 2] EtOCS2K. (c) LiOH, THF,
MeOH. (d) Cl2, CCl4, H2O. (e) 4-Aminoacetophenone, pyridine, (f) trimethylphenylammonium tribromide, THF. (g) Potassium thioacetate, MeOH, room temperature, 18h.
Scheme IH
Figure imgf000025_0002
Reagents: (a) Pyridine, 4O0C, 6h (b) Copper iodide, 1,10-phenanthroline, cesium carbonate,
120°C, 24h. (c) Trimethylphenylammonium tribromide, THF, 50°C, 5h or HBr/AcOH, trimethylphenylammonium tribromide, CH2Cl2, MeOH, THF, room temperature, 30 min. (d) Potassium thioacetate, MeOH, room temp., 18h. Scheme IV
Figure imgf000026_0001
Reagents: (a) Acetic anhydride, pyridine, 6O0C, 18h. (b) PCl5, 60°C, 6h. (c) Pyridine, THF,
50°C, 18h. (d) K2CO3, H2O, EtOH, room temperature, 5h. (e) Trimethylphenylammonium tribromide, THF, 50°C, 18h. (f) Potassium thioacetate, MeOH, room temp., 30min.
Scheme V
Figure imgf000026_0002
Reagents: (a) Pyridine, THF, 40°C, 6h. (b) Trimethylphenylammonium tribromide, THF,
50°C, 18h. (c) IM HCl, 1,4-dioxane, 110°C, 6h. (d) Potassium thioacetate, MeOH, room temp., 18h. Scheme VI
Figure imgf000027_0001
Reagents: (a) Pyridine, THF, 40°C, 6h. (b) Trimethylphenylammonium tribromide, THF,
50°C, 18h. (c) Potassium thioacetate, MeOH, room temp., 18h.
The invention is further illustrated by the following examples.
EXAMPLE 1
Thioacetic acid S-{2-[4-(4-butoxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000027_0002
Step 1 : N-(4~Acetyl-phenyl)-4-butoxy-benzenesuIfbnamide
Figure imgf000027_0003
A mixture of 4-butoxy-benzenesulfonyl chloride (2g, 8 mmol), l-(4-amino-phenyl)-ethanone (1.41g, 10.45 mmol), and pyridine (4.5 mL, 32 mmol) in THF (20 mL) was heated to 4O0C for 6h. The solvent was removed in vacuo and the residue was partitioned between EtOAc and IM aqueous HCl. The aqueous layer was extracted with EtOAc (3x). The organics were combined, dried over Na2SO4 and evaporated to dryness to afford 1.44g of N-(4-acetyl-phenyl)-4-butoxy-benzenesulfonamide as a white solid. 1H NMR (400 MHz, CDCl3) β 7.84 (d, 2H), 7.75 (d, 2H), 7.15 (d, 2H), 6.90 (d, 2H), 3.97 (t, 2H), 2.53 (s, 3H), 1.76 (m, 2H), 1.46 (m, 2H), 0.96 (t, 3H). Step 2: N-[4-(2-Bromo-acetyl)-phenyl]-4-butoxy-benzenesulfoπamide
Figure imgf000028_0001
A mixture of N-(4-acetyl-phenyl)-4-butoxy-benzenesulfonamide (1.44g, 4.15 mmol) and trimethylphenylammonium tribromide (1.7g, 4.6 mmol) in THF (20 mL) was heated to 40 0C for 2h. The solvent was removed in vacuo and the residue was partitioned between EtOAc and water. The aqueous layer was extracted with EtOAc (3x). The organics were combined, dried over Na2SO4 and evaporated to dryness to afford 2g of a mixture of N-[4-(2-bromo-acetyl)-ρhenyl]-4-butoxy- benzenesulfonamide, 4-butoxy-N-[4-(2,2-dibromo -acetyl)-phenyl]-benzenesulfonamide and unreacted starting material.
Step 3: Thioacetic acid S-{2-[4-(4-butoxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000028_0002
A mixture of N-[4-(2-bromo-acetyl)-phenyl]-4-butoxy-benzenesulfonamide (2g, 4.7 mmol) and potassium thioacetate (641mg, 5.6 mmol) in MeOH (20 mL) was stirred at room temperature for 18h. The solvent was removed in vacuo and the residue was partitioned between CH2Cl2 and water. The aqueous layer was extracted with CH2CI2 (3x). The organics were combined, dried over Na2Sθ4 and evaporated to dryness to afford a crude compound that was purified by flash chromatography (silica gel, 1:1 EtOAc±exane). Thioacetic acid S-{2-[4-(4-butoxy- benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester was obtained as a white solid (1.12g). 1H NMR (400 MHz, DMSO-d6) δ 10.81 (s, IH), 7.88 (d, 2H), 7.75 (d, 2H), 7.21 (d, 2H), 7.06 (d, 2H), 4.40 (s, 2H), 4.01 (m, 2H), 2.35 (s, 3H), 1.66 (m, 2H), 1.38 (m, 2H), 0.90 (t, 3H). LCMS: 422 (M+l)+. EXAMPLE 2
Thioacetic acid S-{2-[4-(benzotWazole-6-sulfonyIamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000029_0001
Thioacetic acid S-{2-[4-(benzothiazole-6-sulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using benzothiazole-6-sulfonyl chloride and l-(4-amino- phenyl)-ethanone as starting materials. 1HNMR (400 MHz, CDCl3) δ 9.20(s, IH), 8.57(s, IH), 8.21(d, IH), 7.97(d, IH), 7.86(d, 2H), 7.20(d, 2H), 4.24(s, 2H), 2.39(s, 3H). LCMS: 408 (M+l)+.
EXAMPLE 3
Thioacetic acid S-{2-oxo-2-[4-(2-oxo-2H-chromene-6-suIfonylamino)-pheπyl]-ethyl} ester
Figure imgf000029_0002
Thioacetic acid S-{2-oxo-2-[4-(2-oxo-2H-chromene-6-sulfonylamino)-phenyl]-ethyl} ester was synthesized as described in EXAMPLE 1 using 2-oxo-2H-chromene-6-sulfonyl chloride and l-(4- amino-phenyl)-ethanone as starting materials. 1H NMR (400 MHz, CDCl3) δ 8.01(s, IH), 7.98(d, IH), 7.89(d, 2H), 7.70(d, IH), 7.41(d, IH), 7.19(d, 2H), 6.52(d, IH), 4.28(s, 2H), 2.39(s, 3H). LCMS: 418 (M+l)+.
EXAMPLE 4
Thioacetic acid S-{2-[4-(2,2-dimethyl-chroman-6-sulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000029_0003
Thioacetic acid S- {2-[4-(2,2-dimethyl-chroman-6-sulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 2,2-dimethyl-chroman-6-sulfonyl chloride and l-(4- amino-phenyl)-ethanone as starting materials. 1H NMR (400 MHz, CDCl3) δ 7.89(d, 2H), 7.60(s, IH), 7.17(d, 2H), 6.88(d, IH), 6.80(4 IH), 4.29(s, 2H), 2.78(t, 2H), 2.40(s, 3H), 1.80ft 2H), 1.3 l(s, 6H). LCMS: 434 (M+l)+.
EXAMPLE 5
Thioacetic acid S-{2-oxo-2-[4-(4-pyrazol-l-yl-benzenesulfonyIamino)-phenyl]-ethyl} ester
Figure imgf000030_0001
Thioacetic acid S-{2-oxo-2-[4-(4-pyrazol-l-yl-benzenesulfonylamino)-phenyl]-ethyl} ester was synthesized as described in EXAMPLE 1 using 4-ρyrazol-l-yl-benzenesulfonyl chloride and l-(4- amino-phenyl)-ethanone as starting materials. 1H NMR (400 MHz, CDCl3) δ 8.01(d, IH), 7.92, (d, 2H), 7.88(d, 2H), 7.80(d, 2H), 7.78(d, IH), 7.19(d, 2H), 6.52(m, IH), 4.29(s, 2H), 2.40(s, 3H). LCMS: 416 (M+l)+.
EXAMPLE 6
Thioacetic acid S-{2-[4-(7-chloro-benzo[l,2,5]oxadiazole-4-sulfonylamino)-phenyI]-2-oxo- ethyl} ester
Figure imgf000030_0002
Thioacetic acid S-{2-[4-(7-chloro-benzotl,2,5]oxadiazole-4-sulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 7-chloro-benzo[l,2,5]oxadiazole-4- sulfonyl chloride and l-(4-amino-phenyl)-ethanone as starting materials. 1H NMR (400 MHz, CDCl3) δ 8.03(d, (IH), 7.82(d, 2H), 7.51(d, IH), 7.21(d, 2H), 4.24(s, 2H), 2.39(s, 3H). LCMS: 426 (M+l)+. EXAMPLE 7
Thioacetic acid S-(2-{4-[3-(2-methyl-pyrimidin-4-yl)-benzenesuIfonylamino]-phenyl}-2-oxo- ethyl) ester
Figure imgf000031_0001
Thioacetic acid S-(2-{4-[3-(2-methyl-ρyrimidin-4-yl)-benzenesulfonylamino]-phenyl}-2-oxo-ethyl) ester was synthesized as described in EXAMPLE 1 using 3-(2-methyl-pyrimidin-4-yl)- benzenesulfonyl chloride and l-(4-amino-phenyl)-ethanone as starting materials. 1HNMR (400 MHz, CDCl3) δ 8.80(d, IH), 8.02(s, IH), 7.98(d, IH), 7.90(d, IH), 7.82(4 2H), 7.78(d, IH), 7.64(m, IH), 7.61(d, 2H), 4.26(s, 2H), 2.90(s, 3H), 2.39(s, 3H). LCMS: 442 (M+l)+.
EXAMPLE 8
Thioacetic acid S-{2-[4-(6-morpholin-4-yl-pyridine-3-sulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000031_0002
Thioacetic acid S-{2-[4-(6-morpholin-4-yl-pyridine-3-sulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 6-moφholin-4-yl-pyridine-3-sulfonyl chloride and l-(4-amino-phenyl)-ethanone as starting materials. 1H NMR (400 MHz, CDCl3) δ 8.60(s, IH), 7.90(d, 2H), 7.79(d, IH), 7.19(d, 2H), 6.54(d, IH), 4.29(s, 2H), 3.70(t, 4H), 3.62(t, 4H), 2.39(s, 3H). LCMS: 436 (M+l)+. EXAMPLE 9
Thioacetic acid S-{2-[4-(6-methoxy-pyridine-3-sulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000032_0001
Thioacetic acid S-{2-[4-(6-methoxy-pyridine-3-sulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 6-methoxy-pyridine-3-sulfonyl chloride and l-(4- amino-phenyl)-ethanone as starting materials. 1HNMR (400 MHz, CDCl3) δ 8.66(s, IH), 7.9 l(d, 2H), 7.20(d, 2H), 6.88(d, IH), 6.79(d, IH), 4.29(s, 2H), 3.99(s, 3H), 2.41(s, 3H). LCMS: 381 (M+l)+.
EXAMPLE 10
Thioacetic acid S-{2-[6-(6-methoxy-pyridine-3-sulfonylamino)-pyridin-3-yl]-2-oxo-ethyl} ester
Figure imgf000032_0002
Thioacetic acid S-{2-[6-(6-methoxy-pyridine-3-sulfonylamino)-pyridin-3-yl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 6-methoxy-pyridine-3-sulfonyl chloride and l-(6- amino-pyridin-3-yl)-ethanone as starting materials. 1H NMR (400 MHz, CDCl3) δ 8.73(s, IH), 8.71(s, IH), 7.99(d, IH), 7.96(d, IH), 7.04(d, IH), 6.60(d, IH), 3.89(s, 2H), 3.34(s, 3H), 2.39(s, 3H). LCMS: 382 (M+l)+.
EXAMPLE I l
Thioacetic acid S-{2-[4-(4-nitro-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000032_0003
Thioacetic acid S-{2-[4-(4-nitro-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 4-nitro-benzenesulfonyl chloride and l-(4-amino-phenyl)- ethanone as starting materials. 1HNMR (400 MHz, DMSO-d6) δ 11.25 (s, IH)1 8.39 (d, 2H), 8.09 (d, 2H), 7.92 (d, 2H), 7.25 (d, 2H), 4.41 (s, 2H), 2.35 (s, 3H). LCMS: 395 (M+l)+. RXAMPLE 12
Thioacetic acid S-{2-[4-(4-amino-benzeπesulfonyIamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000033_0001
Step 1: N-[4-(4-Acetyl-phenylsuIfamoyl)-phenyl]-acetamide
Figure imgf000033_0002
N-[4-(4-Acetyl-ρhenylsulfamoyl)-phenyl]-acetamide was synthesized as described in EXAMPLE 1, Step 1 using 4-acetylamino-benzenesulfonyl chloride and l-(4-amino-phenyl)-ethanone as starting materials.
Step 2: N-{4-[4-(2-Bromo-acetyl)-phenylsulfamoyl]-phenyl}-acetamide
Figure imgf000033_0003
N-{4-[4-(2-Bromo-acefyl)-phenylsulfamoyl]-phenyl}-acetamide was synthesized according to EXAMPLE 1, Step 2 using N-[4-(4-acetyl-phenylsulfamoyl)-phenyl]-acetamide as starting material.
Step 3: 4-Amino-N-[4-(2-chloro-acetyl)-phenyl]-benzenesulfonamide
Figure imgf000033_0004
N-{4-[4-(2-Bromo-acetyl)-phenylsulfamoyl]-phenyl}-acetamide (530 mg, 1.29 mmol) was dissolved in 1,4-dioxane (8 mL) and aqueous HCl (IM, 4 mL) and the resulting mixture was heated to 110 0C for 2h. The reaction mixture was then cooled to room temperature and slowly poured into water containing Na2CO3. The aqueous layer was extracted with CH2Cl2 (3x). The organics were combined, dried over Na2SO4 and evaporated to dryness to afford 414 mg of 4-amino-N-[4-(2- chloro-acetyl)-phenyl]-benzenesulfonamide as a yellow solid. Step 4: Thioacetic acid S-{2-[4-(4-amino-beπzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester
Thioacetic acid S-{2-[4-(4-amino-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1, Step 3 using 4-amino-N-[4-(2-chloro-acetyl)-phenyl]- benzenesulfonamide as starting material. 1HNMR (400 MHz, DMSOd6) δ 10.55 (s, IH), 7.86 (d, 2H), 7.47 (d, 2H), 7.17 (d, 2H), 6.55 (d, 2H), 6.05 (s, 2H), 4.40 (s, 2H), 2.36 (s, 3H). LCMS: 365
(M+l)+.
EXAMPLE 13
Thioacetic acid S-{2-[4-(2,4-dimethoxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000034_0002
Thioacetic acid S-{2-[4-(2,4-dimethoxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 2,4-dimethoxy-benzenesulfonyl chloride and l-(4- amino-phenyl)-ethanone as starting materials. 1HNMR (400 MHz, DMSOd6) δ 10.59 (s, IH), 7.84 (d, 2H), 7.79 (d, IH), 7.17 (d, 2H), 6.63 (m, 2H), 4.39 (s, 2H), 3.83 (s, 3H), 3.79 (s, 3H), 2.34 (s, 3H).
EXAMPLE 14
Thioacetic acid S-(2-{6- [4-(2-methoxy-ethoxy)-benzenesulfonylamino]-pyridin-3-yl}-2-oxo- ethyl) ester
Figure imgf000034_0003
Step 1 : N-(5-Acetyl-pyridin-2-yl)-4-iodo-benzenesulfonamide
Figure imgf000034_0004
N-(5-Acetyl-pyridin-2-yl)-4-iodo-benzenesulfonamide was synthesized as described in EXAMPLE 1, Step 1 using 4-iodo-benzenesulfonyl chloride and l-(6-ammo-pyridin-3-yl)-ethanone as starting materials and pyridine as solvent. 1HNMR (400 MHz, DMSO-dg) β 8.59 (s, IH), 8.13 (dd, IH), 7.92 (d, 2H), 7.64 (d, 2H), 7.20 (d, IH), 2.45 (s, 3H).
Step 2 : N-(5-Acetyl-pyridin-2-yl)-4-(2-methoxy-ethoxy)-benzenesulfonamide
Figure imgf000035_0001
A mixture ofN-(5-Acetyl-pyridin-2-yl)-4-iodo-benzenesulfonamide (Ig, 2.5 mmol), 2-Methoxy- ethanol (2 mL, 25 mmol), copper iodide (71mg, 0.37 mmol), 1,10-ρhenanthroline (134mg, 0.75 mmol), and cesium carbonate (2.2g, 6.2 mmol) was heated to 12O0C for 24h. The black reaction mixture was cooled to room temperature and partitioned between CH2CI2 and a saturated aqueous ammonium chloride solution. The aqueous layer was extracted with CH2Cl2 (4x), the organics were combined, dried over Na2SO4 and evaporated to dryness to afford a crude that was purified by flash chromatography (silica gel, 50 to 100% Haxane:EtOAc). N-(5-Acetyl-pyridin-2-yl)-4-(2-methoxy- ethoxy)-benzenesulfonamide (535mg) was obtained as yellow solid. 1H NMR (400 MHz, CDCl3) δ 8.95 (s, IH), 8.19 (d, IH), 7.81 (d, 2H), 7.40 (d, IH), 6.94 (d, 2H), 4.13 (t, 2H), 3.74 (t, 2H), 3.42 (s, 3H), 2.53 (s, 3H).
Step 3 : N-[5-(2-Bromo-acetyl)-pyridin-2-yl]-4-(2-methoxy-ethoxy)-benzenesulfonainide
Figure imgf000035_0002
N-[5-(2-Bromo-acetyl)-pyridin-2-yl]-4-(2-methoxy-ethoxy)-benzenesulfonamide was synthesized as described in EXAMPLE 1 , Step 2 using N-(5-acetyl-pyridin-2-yl)-4-(2-methoxy-ethoxy)- benzenesulfonamide as starting material.
Step 4: Thioacetic acid S-(2-{6-[4-(2-methoxy-ethoxy)-benzenesulfonylamino]-pyridin-3-yl}-2- oxo-ethyl) ester
Figure imgf000035_0003
Thioacetic acid S-(2-{6-[4-(2-methoxy-ethoxy)-benzenesulfonylamino]-pyridin-3-yl}-2-oxo-ethyl) ester was synthesized as described in EXAMPLE 1, Step 3 using N-[5-(2-bromo-acetyl)-pyridin-2- yl]-4-(2-methoxy-ethoxy)-benzenesulfonamide as starting material. 1H NMR (400 MHz, CDCl3) δ 9.02 (s, IH), 8.20 (d, IH), 7.85 (d, 2H), 7.41 (d, IH), 6.97 (d, 2H), 4.23 (s, 2H), 4.14 (t, 2H), 3.74 (t, 2H), 3.42 (s, 3H), 2.37 (s, 3H). LCMS: 425 (M+l)+.
EXAMPLE 15
Thioacetic acid S-(2-{6-[4-(3-methoxy-propoxy)-benzenesulfonylamino]-pyridin-3-yl}-2-oxo- ethyl) ester
Figure imgf000036_0001
Thioacetic acid S-(2-{6-[4-(3-methoxy-propoxy)-benzenesulfonylamino]-pyridin-3-yl}-2-oxo-ethyl) ester was synthesized as described in EXAMPLE 14 using 3-methoxy-propan-l-ol and N-(5-acetyl- pyridin-2-ylM-iodo-benzenesulfonamide as starting materials. 1H NMR (400 MHz, CDCl3) δ 9.04 (s, IH), 8.19 (d, IH), 7.84 (d, 2H), 7.42 (d, IH), 6.94 (d, 2H), 4.23 (s, 2H), 4.08 (t, 2H), 3.52 (t, 2H), 3.33 (s, 3H), 2.37 (s, 3H), 2.02 (m, 2H). LCMS: 439 (M+l)+.
EXAMPLE 16
This example is intentionally left blank.
EXAMPLE 17
Thioacetic acid S-(2-oxo-2-[6-(4-pyrrolidin-l-yl-benzenesulfonylamino)-pyridin-3-yl]-ethyl} ester
Figure imgf000036_0002
Thioacetic acid S-(2-oxo-2-[6-(4-pyrrolidin-l-yl-benzenesulfonylamino)-pyridin-3-yl]-ethyl} ester was synthesized as described in EXAMPLE 14 using N-(5-acetyl-pyridin-2-yl)-4-iodo- benzenesulfonamide and 1.2 equivalents of pyrrolidine as starting materials and using DMSO (3mL) as solvent in Step 1. 1HNMR (400 MHz, CDCl3) δ 8.98 (s, IH), 8.18 (dd, IH), 7.71 (d, 2H), 7.40 (d, IH), 6.54 (d, 2H), 4.25 (s, 2H), 3.25-3.35 (m, 4H), 2.39 (s, 3H), 1.95-2.05 (m, 4H). LCMS: 420.4 (M+l)+. EXAMPLE 18
Thioacetic acid S-{2-[4-(l-methyl-lH-benzoimidazoIe-5-sulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000037_0001
Step 1: l-Methyl-lH-benzoimidazol-5-ylamine
Figure imgf000037_0002
Raney nickel was washed with iPrOH followed by toluene. Under N2 atmosphere, l-methyl-5- nitroimidazole (0.5Og, 2,8 mmol) was added to toluene (20 mL) containing the pre-washed Raney nickel. The reaction flask was purged with H2 and the reaction mixture was warmed to 400C for two hours. The resulting mixture was filtered through celite and concentrated to give 0.342 g (82%) of a white solid.
Step 2: Dithiocarbonic acid O-ethyl ester S-(l~methyl-lH-benzoimidazol-5-yl) ester:
Figure imgf000037_0003
l-Methyl-lH-benzoimidazol-5-ylamine (2.3 Ig, 15.7 mmol) obtained from step 1 was suspended in 30 mL of water, the solution was then cooled to 00C and aqueous HCl (3.14 mL, 37.9 mmol) was added. The reaction mixture was stirred in an ice/ water bath. NaNO2 in 5 mL OfH2O was added in small portion to the reaction mixture over a period of 30 minutes. The reaction mixture was then added portionwise to a 45 0C solution OfEtOCS2K in 10 mL of water. The reaction mixture was stirred at 450C for an additional 30 minutes, and then it was extracted with Et2O (2 x 25 mL). The organic layer was washed with IN NaOH (2 x 100 mL), brine (1 x 75 mL), dried over Mg2SO4, filtered and concentrated to give 0.61 g (42 %) of a yellow oil. Step 3: l-Methyl-lH-benzoimidazole-5-thiol
Figure imgf000037_0004
Dithiocarbonic acid O-ethyl ester S-(l-methyl-lH-benzoimidazol-5-yl) ester (1.38 g, 5.47 mmol) from step 2 and LiOH (0.52 g, 21.9 mmol) was added to 30 mL of THF/MeOH (3 : 1) followed by the addition of H2O (8 mL). The suspension was stirred at 600C under N2 for 3.5 hours. The resulting mixtures was diluted in 300 mL OfH2O and washed with CH2Cl2 (2 x 100 mL). The aqueous layer was acidified to pH < 1 with concentrated HCl and extracted with CH2Cl2 (3 x 100 mL). The organic extracts was separated and dried with Mg2SO,!, filtered and concentrated to give a while solid (0.76 g, 85 %).
Step 4: l-Methyl-lH-benzoimidazole-5-sulfonyl chloride
Figure imgf000038_0001
1 -Methyl- lH-benzoimidazole-5 -thiol from step 3 was dissolved in 35 mL OfCCVH2O (6:1). Cl2 gas was bubbled through the solution at 00C for 50 minutes. The reaction mixture was diluted in 100 mL of CH2Cl2 and 100 mL OfH2O. The organic layer was dried over Mg2SO4, filtered, and concentrated to give a white solid (0.91 g, 85 %).
Step 5: l-Methyl-lH-beπzoimidazole-5-sulfonic acid (4-acetyl-phenyl)-amide
Figure imgf000038_0002
l-Methyl-lH-benzoimidazole-5-sulfonyl chloride (0.310 g, 1.3 mmol) and l-(4-Amino-phenyl)- ethanone (0.175 g, 1.3 mmol) were dissolved in pyridine. The reaction mixture was heated to 600C under N2 for 1.5 hours. The resulting solution was concentrated and dissolved in CH2Cl2 (75 mL), washed with 0.5 N HCl (1 x 75 mL), and dried over Mg2SO4, filtered and concentrated to give of an off-white solid (0.345 g, 78 %).
Step 6: l-Methyl-lH-benzoimidazole-5-sulfonic acid [4-(2-bromo-acetyl)-phenyl]-amide
Figure imgf000038_0003
l-Methyl-lH-benzoimidazole-5-sulfonic acid (4-acetyl-phenyl)-amide (0.345 g, 1.05 mmol) and trimethyl-phenyl-ammonium bromide (0.433 g, 1.15 mmol) were dissolved in THF (10 mL). The reaction solution was heated to 450C under N2 for 1 hour. Additional 50 mL of CH2Cl2 was added to the reaction solution and the resulting solution was washed with 50 mL OfH2O. The organic layer was dried over Mg2SO4, filtered and concentrated to give of an off-white solid. The resulting residue was purified using a preparative HPLC to afford a white solid (0.110 g, 26 %). Step 7: Thioacetic acid S-{2-[4-(l-methyl-lH-benzoimidazole-5-sulfonylamino)-phenyl]-2-oxo- ethyl} ester
Figure imgf000039_0001
To 5 mL of l-Methyl-lH-benzoiinidazole-5-sulfoπic acid [4-(2-bromo-acetyl)-phenyl]-amide (0.174 g, 0.43 mmol) in MeOH was added potassium thioacetate (0.054 g, 0.47 mmol). The reaction mixture was stirred under N2 at room temperature for 45 minutes. The resulting reaction mixture was dissolved in 50 mL of CH2Cl2 and washed with 50 mL of H2O. The organic layer was dried over Mg2SO4, filtered and concentrated to give of an off-white solid. The resulting residue was purified using a preparative HPLC to afford a white solid (140 mg). 1H-NMR (DMSO): 10.91 (s, IH), 8.39 (s, IH) 8.26 (s, IH), 7.81 (m, 4H), 7.23 (d, 2H), 4.37 (s, 2H), 4.05 (s, 3H), 2.34 (s, 3H) ppm; LC-MS (ES+): 404 [MH]+ m/e.
EXAMPLE 19
Thioacetic acid S-{2-[4-(2,3-dihydro-lH-indole-S-sulfonyIamiπo)-phenyl]-2-oxo-ethyl} ester
Figure imgf000039_0002
Step 1: l-(2,3-Dihydro-indol-l-yl)-ethanone
Figure imgf000039_0003
Indoline (10 mL, 89.2 mmol)and Et3N (38 mL, 268 mmol) were dissolved in CH2Cl2, followed by an addition of a catalytic amount of DMAP. After the reaction mixture was cooled to 00C, acetic anhydride was added. The resulting reacting mixture was stirred for 10 minutes under N2 and then warmed up to room temperature. The reaction mixture was continued to be stirred for another two hours. The reaction mixture was dissolved in EtOAc (200 mL), washed with IN HCl (2 x 150 mL), saturated bicarbonate (2 x 150 mL), brine (1 x 100 mL). The organic layer was dried over Mg2SO4, filtered, and concentrated. The crude material was recrystallized from hot EtOAc / hexanes to give a pink solid (10.45 g, 73 %). Step 2: l-Acetyl-2,3-dihydro-lH-indole-5-sulfonyl chloride
Figure imgf000040_0001
Chlorosulfonic acid (25 mL) was cooled to 00C in an ice bath. l-(2,3-Dihydro-indol-l-yl)-ethanone (5.00 g, 31 mmol) obtained from step 1 was added slowly into the cold chlorosulfonic acid. The reaction mixture was stirred at 00C under N2 for 20 minutes, and then it was heated to 700C and continued stirring for 5 hours. The resulting mixture was quenched by pipetting into 500 mL of crushed ice. The precipitated sulfonyl chloride / H2O was extracted with CH2Cl2 (3 x 250 mL). The organic layer was dried over Mg2SO4, filtered and concentrated to give an off-white solid which was used directly in the next step.
Step 3: l-Acetyl-2,3~dihydro-lH-indole-5-sulfonic acid [4-(2-bromo-acetyl)-phenyl]-amide
Figure imgf000040_0002
The compound l-Acetyl-2,3-dihydro-lH-indole-5-sulfonic acid [4-(2-bromo-acetyl)-ρhenyl]-amide was synthesized according to the procedures described in EXAMPLE 18 Steps 5 and 6.
Step 4: 2,3-Dihydro-lH-indole-5-sulfonic acid [4-(2-chloro-acetyl)-phenyl]-amide
Figure imgf000040_0003
To l-Acetyl-2,3-&hydro-lH-indole-5-sulfonic acid [4-(2-bromo-acetyl)-phenyl]-amide (0.600 g, 1.37 mmol) in 1,4-dioxane (2.4 mL) was added 12N HCl (1.2 mL). The reaction mixture was heated to 1000C under N2 for 4 hours. The reaction was cooled to room temperature, dissolved in 75 mL of MeOHTH2O (3:1) and neutralized with saturated aqueous NaHCO3. The reaction solution was filtered, concentrated and diluted with H2O, extracted with CH2Cl2 (3 x 50 mL), dried over Mg2SO4, filtered and concentrated to give an off-white solid (0.268 g, 56 %). Step 5: Thioacetic acid S-{2-[4-(2,3-dihydro-lH-indole-5-sulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000041_0001
The compound thioacetic acid S-{2-[4-(2,3-dmydro-lH-indole-5-sulfonylamino)~phenyl]-2-oxo- ethyl} ester was synthesized according to the procedure described in EXAMPLE 18, Step 7 using 2,3-Dihydro-lH-indole-5-sulfonic acid [4-(2-chloro-acetyl)-phenyl]-amide obtained from the previous step. 1H-NMR (DMSO): 10.57 (s, IH), 7.87 (d, 2H) 7.40 (m, 2H), 7.18 (d, 2H), 6.43 (s, IH), 4.41 (s, 2H), 3.50 (t, 2H), 2.94 (t, 2H), 2.36 (s, 3H) ppm; LC-MS (ES+): 391 [MH]+ m/e.
EXAMPLE 20
Thioacetic acid S-{2-[4-(l-acetyl-2,3-dihydro-lH-indole-5-sulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000041_0002
The compound thioacetic acid S-{2-[4-(l-acetyl-2,3-dihydro-lH-indole-5-sulfonylamino)-phenyl]- 2-oxo-ethyl} ester was synthesized according to the procedure described in EXAMPLE 18, Step 7 using l-acetyl-2,3-dihydro-lH-indole-5-sulfonic acid [4-(2-bromo-acetyl)-phenyl]-amide obtained from EXAMPLE 19, Step 3. 1H-NMR (DMSO): 10.83 (s, IH), 8.10 (d, IH) 7.88 (d, 2H), 7.67 (d, 2H), 7.21 (d, 2H), 4.40 (s, 2H), 4.11 (t, 2H), 3.16 (t, 2H), 2.35 (s, 3H), 2.15 (s, 3H); LC-MS (ES+): 433 [MH]+ m/e.
EXAMPLE 21
Thioacetic acid S-{2-[4-(l-acetyl-2,3-dihydro-lH-indole-6-sulfonylamiπo)-phenyl]-2-oxo-ethyl} ester
Figure imgf000041_0003
Step 1: l-(5-Bromo-2,3-dihydro-indol-l-yl)-ethanone
Figure imgf000042_0001
Bromine (1.8 mL, 34 mmol) was added to l-(2,3-dihydro-indol-l-yl)-ethanone (5.45g, 34 mmol) dissolved in Acetic acid (50 mL) and the resulting mixture was stirred at room temperature for 45 min. The reaction mixture was poured into EtOAc (300 mL) and it was washed with water (2x) followed by saturated NaHCO3 (2x), dried over MgSO4, and evaporated to dryness to afford the desired compound as a crude. Pure l-(5-bromo-2,3-dihydro-indol-l-yl)-ethanone (6.6g) was obtained as a white solid by recrystallization (EtOAc:Hexane). Step 2: l-Acetyl-5-bromo-2,3-dihydro-lH-indole-6-sulfonyl chloride
Figure imgf000042_0002
l-Acetyl-5-bromo-2,3-dihydro-lH-indole-6-sulfonyl chloride was synthesized as described in EXAMPLE 19, Step 2 using l-(5-bromo-2,3-dihydro-indol-l-yl)-ethanone as starting material.
Step 3: l-Acetyl-5-bromo-2,3-dihydro-lH-indole-6-sulfonic acid (4-acetyl-phenyl)-amide
Figure imgf000042_0003
The compound l-acetyl-5-bromo-2,3-dihydro-lH-indole-6-sulfonic acid (4-acetyl-phenyl)-amide was synthesized according to the procedures described in EXAMPLE 18, Step 5 using l-acetyl-5- bromo-2,3-dihydro-lH-indole-6-sulfonyl chloride and l-(4-amino-phenyl)-ethanone as starting materials.
Step 4: l-Acetyl-2,3-dihydro-lH-indole-6-sulfonic acid (4-acetyl-phenyl)-amide
Figure imgf000042_0004
To the solution of l-acetyl-54>romo-2,3-dihydro4H4ndole-6-sulfonic acid (4-acetyl-phenyl)-amide (5.61 g, 12.8 mmol) obtained from step 1 in DMF (100 mL) was added Et3N (2.2 mL, 15.4 mmol) and a catalytic amount of 10 % Pd/C. The reaction was stirred under a H2 atmosphere overnight. The resulting reacting mixture was filtered to remove Pd/C, then it is concentrated. The crude residue was washed with 50 % aqueous MeOH to remove Et3N.HBr, filtered and dried under vacuum to give a white solid (3.91 g, 85 %).
Sep 5: Thioacetic acid S-{2-[4-(l-acetyl-2,3-dihydro-lH-indole-6-sulfonylamino)-phenyl]-2- oxo-ethyl} ester
Figure imgf000043_0001
The compound thioacetic acid S-{2-[4-(l-acetyl-2>3-dihydro-lH-indole-6-sulfonylamino)-phenyl]- 2-oxo-ethyl} ester was synthesized according to the procedures described in EXAMPLE 18, Steps 6 and 7 using l-acetyl-2,3-dihydro-lH-indole-6-sulfonic acid (4-acetyl-phenyl)-amide obtained from the previous step. 1H-NMR (DMSO): 10.99 (s, IH), 8.54 (s, IH) 7.88 (d, 2H), 7.48 (d, IH), 7.37 (d, IH), 7.21 (d, 2H), 4.40 (s, 2H), 4.11 (t, 2H), 3.16 (t, 2H), 2.35 (s, 3H), 2.15 (s, 3H) ppm; LC-MS (ES+): 433 [MH]+ m/e.
EXAMPLE 22
Thioacetic acid S-{2-[4-(lH-indole-6-sulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000043_0002
Step 1 : lH-Indole-6-sulfonic acid [4-(2-chloro-acetyl)-phenyl]-amide
Figure imgf000043_0003
2,3-Dihydro-lH-indole-6-sulfonic acid [4-(2-chloro-acetyl)-phenyl]-amide (0.300 g, 0.86 mmol) that was prepared in a similar fashion as in EXAMPLE 19, step 4, was dissolved in acetonitrile (13 mL). DDQ (0.204 g, 0.90 mmol) was added and the solution was stirred at room temperature for 3 hours. The reaction mixture was purified using a preparative HPLC to give a pale orange solid (0.197 g, 66 %)• Step 2: Thioacetic acid S-{2-[4-(lH-indole-6-sulfonylamlno)-phenyl]-2-oxo-ethyl} ester
Figure imgf000044_0001
The compound thioacetic acid S-{2-[4-(lH-indole-6-sulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized according the method described in EXAMPLE 18, Step 7 using lH-indole-6-sulfonic acid [4-(2-chloro-acetyl)-phenyl]-amide as starting material. 1H-NMR (DMSO): 11.63 (s, IH),
10.83 (s, IH) 7.94 (s, IH), 7.85 (d, 2H), 7.68 (d, IH), 7.62 (m, IH), 7.44 (dt, IH), 7.23 (d, 2H), 6.53 (s, IH), 4.37 (s, 2H), 2.33 (s, 3H) ppm; LC-MS (ES+): 389 [MH]+ vale.
EXAMPLE 23
Thioacetic acid S-{2-[4-(4-hydroxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000044_0002
Step 1 : Sodium 4-acetoxy-benzenesulfonate
Figure imgf000044_0003
Acetic anhydride (250 mL).and pyridine (16g, 202.28 mmol) were added to sodium 4- hydroxybenzenesulfonate (36g, 183.52 mmol) and the resulting solution was allowed to react, with stirring, overnight while the temperature was maintained at 60-70°C until done by TLC. The mixture was concentrated by evaporation under vacuum using a rotary evaporator. This resulted in 35g (80%) of sodium 4-acetoxybenzenesulfonate as a dark red solid.
Step 2: Acetic acid 4-chlorosulfonyl-phenyl ester
Figure imgf000044_0004
Phosphorus pentachloride (61 g, 292.93 mmol) was added to sodium 4-acetoxybenzenesulfonate (35 g, 146.94 mmol) and the resulting solution was allowed to react, with stirring, for 6h while the temperature was maintained at 6O0C. The reaction mixture was then quenched by the adding 300 ml of H2O/ice. The resulting solution was extracted (2x) with 300 ml of CH2Cl2 and the organic layers combined, dried over MgSO4, and concentrated by evaporation under vacuum using a rotary evaporator. This resulted in 35g (61%) of acetic acid 4-chlorosulfonyl-phenyl ester as yellow oil. Step 3: Acetic acid 4-(4-acetyl-pheπylsulfamoyl)-phenyl ester
Figure imgf000045_0001
Acetic acid 4-(4-acetyl-phenylsulfamoyl)-phenyl ester was synthesized as described in EXAMPLE 1, Step 1 using acetic acid 4-chlorosulfonyl-phenyl ester and l-(4-amino-phenyl)-ethanone as starting materials.
Step 4 : N-(4-acetyl-phenyl)-4-hydroxy-beπzenesulfonamide
Figure imgf000045_0002
Into a 500 ml 3-necked roundbottom flask, was placed a solution of acetic acid 4-(4-acetyl- phenylsulfamoyl)-phenyl ester (8 g, 24.00 mmol) in EtOH (100 ml). This was followed by the addition of a solution of potassium hydroxide (1.48 g, 26.38 mmol) in EL2O (10 ml). The resulting solution was allowed to react, with stirring, for 2.5 hours while the temperature was maintained at reflux. The reaction progress was monitored by TLC (CH2Cl2/Me0H = 10:1). The reaction mixture was then quenched by the adding 200 ml of H2O/ice. Adjustment of the pH to 8 was accomplished by the addition of Na2COs (2-3%). The resulting solution was extracted three times with 200 ml of CH2Cl2 and one time with 200 ml of EtOAc, the aqueous layers combined. Adjustment of the pH to 6 was accomplished by the addition of HCl (2-3%). The resulting solution was extracted three times with 200 ml of CH2Cl2 and the organic layers combined, dried over MgSO4 and concentrated by evaporation under vacuum using a rotary evaporator. This resulted in 3 g (43%) of N- (4- acetylphenyl)-4-hydroxybenzenesulfonamide as a light gray solid.
Step 5: Thioacetic acid S-{2-[4-(4-hydroxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000045_0003
Thioacetic acid S-{2-[4-(4-hydroxy-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1, Step 2 and 3 using N-(4-acetyl-phenyl)-4-hydroxy- benzenesulfonamide. 1H-NMR (DMSO): 10.50 (bs, 2H), 7.87 (d, 2H) 7.67 (d, 2H), 7.19 (d, 2H), 6.87 (d, 2H), 4.40 (s, 2H), 2.35 (s, 3H) ppm; LC-MS (ES+): 366 [MH]+ m/e. EXAMPLE 24
Thioacetic acid S-{2-[2-(4-methoxy-benzenesulfonylamino)-pyrimidin-5-yl]-2-oxo-ethyl} ester
Figure imgf000046_0001
Step 1: N-(5-Acetyl-pyrimidin-2-yl)-4-methoxy-benzenesuIfonamide
Figure imgf000046_0002
l-(2-Ammo-pyrimidin-5-yl)-ethanone (265 mg, 1.93 mmol) was dissolved in THF (15 ml) under a nitrogen atmosphere. NaOH (60 % dispersion - 309 mg, 7.72 mmol) was then added, and the resulting mixture was stirred at 60 C for 15 minutes, until gas evolution had ceased. 4- Methoxybenzenesulfonyl chloride was then added as a THF solution (638 mg, 3.09 mmol, in 3 ml THF) over a period of 10 minutes. The reaction was then quenched by addition of acetic acid (1 ml). The volatiles were then removed, leaving a yellow solid which was purified by column chromatography (acetonitrile/dichloromethane). The desired compound was isolated as a white solid (120 mg, 0.39 mmol). 1H-NMR (DMSO): 12.22 (bs, IH), 8.96 (s, 2H), 7.93 (d, 2H), 7.09 (d, 2H), 3.81 (s, 3H), 2.48 (s, 3H).
Step 2: Thioacetic acid S-{2-[2-(4-methoxy-beπzenesulfonylamino)-pyrimidin-5-yl]-2-oxo- ethyl} ester
Figure imgf000046_0003
Thioacetic acid S-{2-[2-(4-methoxy-benzenesulfonylamino)-pyrimidin-5-yl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1, Step 2 and 3 using N-(5-acetyl-pyrimidin-2-yl)-4- methoxy-benzenesulfonamide as starting material. 1H-NMR (DMSO): 12.32 (bs, IH), 9.02 (s, 2H), 7.93 (d, 2H), 7.08 (d, 2H), 4.42 (s, 2H), 3.80 (s, 3H), 2.34 (s, 3H). LC-MS (ES+): 382 [MH]+ m/e.
EXAMPLE 25
This example is intentionally left blank. EXAMPLE 26
This example is intentionally left blank. EXAMPLE 27
This example is intentionally left blank.
EXAMPLE 28
Thioacetic acid S-(2-{6-[4-(3-hydroxy-propoxy)-benzenesuIfonylamino]-pyridin-3-yl}-2-oxo- ethyl) ester
Figure imgf000047_0001
Thioacetic acid S-(2-{6-[4-(3-hydroxy-ρropoxy)-benzenesulfonylamino]-pyridin-3-yl}-2-oxo-ethyl) ester was synthesized as described in EXAMPLE 14 using N-(5-acetyl-ρyridin-2-yl)-4-iodo- benzenesulfonamide andpropane-l,3-diol as starting materials. 1HNMR(400 MHz, CD3OD) δ 8.73 (s, IH), 8.17 (d, IH), 7.91 (d, 2H), 7.20 (d, IH), 7.03 (d, 2H), 4.33 (s, 2H), 4.13 (t, 2H), 3.70 (t, 2H), 2.35 (s, 3H), 1.97 (m, 2H). LCMS: 425 (MH)+.
EXAMPLE 29
Thioacetic acid S-(2-{6-[4-(3-hydroxy-3-methyl-butoxy)-benzeπesulfonylamino]-pyridin-3-yl}- 2-oxo-ethyl) ester
Figure imgf000047_0002
Thioacetic acid S-(2- {6-[4-(3 -hydroxy-3-methyl-butoxy)-benzenesulfonylamino]-pyridin-3 -yl } -2- oxo-ethyl) ester was synthesized as described in EXAMPLE 14 using N-(5-acetyl-pyridm-2-yl)-4- iodo-benzenesulfonamide and 3-methyl-butane-l,3-diol as starting materials. 1H NMR (400 MHz, DMSOd6) δ 8.75 (s, IH), 8.16 (d, IH), 7.86 (d, 2H), 7.17 (d, IH), 7.06 (d, 2H), 4.41 (s, 2H), 4.12 (t, 2H), 2.36 (s, 2H), 1.82 (t, 2H), 1.13 (s, 6H). LCMS: 453 (M+l)+. RXAMPLE 30
Thioacetlc acid S-{2-[4-(4-methanesuIfoπyl-benzenesulfonyIamino)-phenyl]-2-oxo-ethyl}
Figure imgf000048_0001
Thioacetic acid S-{2-[4-(4-methanesulfonyl-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 4-methanesulfonyl-benzenesulfonyl chloride and 1- (4-amino-phenyl)-ethanone as starting materials. 1H-NMR (400 MHz CDCl3) δ 8.06(d, 2H), 8.04(d, 2H), 7.91(d, 2H), 7.20(d, 2H), 4.29(s, 2H), 3.06(s, 3H), 2.40(s, 3H). LCMS: 428 (M+l)+.
EXAMPLE 31
Thioacetic acid S-{2-[4-(4-acetylamino-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester
Figure imgf000048_0002
Thioacetic acid S-{2-[4-(4-acetylamino-benzenesulfonylamino)-phenyl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using 4-acetylamino-benzenesulfonyl chloride and l-(4- amino-phenyl)-ethanone as starting materials. 1HNMR (400 MHz, DMSO-d6) δ 10.85 (s, IH), 10.33 (s, IH), 7.88 (d, 2H), 7.78 (d, 2H), 7.72 (d, 2H), 7.20 (d, 2H), 4.40 (s, 2H), 2.35 (s, 3H), 2.05 (s, 3H). LCMS: 407 (M+l)+.
EXAMPLE 32
Thioacetic acid S-{2-[6-(4-acetylamino-benzenesulfonylamino)-pyridin-3-yl]-2-oxo-ethyl} ester
Figure imgf000048_0003
Thioacetic acid S-{2-[6-(4-acetylamino-benzenesulfonylamino)-pyridin-3-yl]-2-oxo-ethyl} ester was synthesized as described in EXAMPLE 1 using N-{4-[5-(2-bromo-acetyl)-pyridin-2-ylsulfamoyl]- phenylj-acetamide and l-(6-amino-pyridin-3-yl)-ethanone (J. Med. Chem. 1973, 16 (8), 959-961) as starting materials. 1HNMR (400 MHz, DMSOd6) β 10.31 (s, IH), 8.75 (s, IH), 8.15 (d, IH), 7.87 (d, 2H), 7.73 (d, 2H), 7.17 (d, IH), 4.42 (s, 2H), 2.35 (s, 3H), 2.06 (s, 3H). LCMS: 408 (M+l)+.
EXAMPLE 33
N-{4-[5-(2-Acetylsulfanyl-acetyl)-pyridin-2-ylsulfamoyl]-phenyl}-succinamic acid methyl ester
Figure imgf000049_0001
Step 1 : N-(5-Acetyl-pyridin-2-yl)-4-amino-benzenesulfonamide
Figure imgf000049_0002
N-[4-(5-acetyl-pyridine-2-ylsulfamoyl)-phenyl]-acetamide (1.33 g, 3.98 mmol) was dissolved in a dioxane HCl solution (10 ml, 4 M). Water (20 ml) was added, and the resulting tan solution was heated to reflux for 1 hour. The volatiles were removed, and the resulting tan solid was purified by HPLC to leave the desired amine as a tan solid (0.78 g, 67 %). 1H-NMR (400 MHz, DMSOd6) δ 10.50 (bs, 1 H), 8.10 (dd, IH), 7.44 (d, 2H), 7.18 (d, IH), 6.53 (d, 2H), 6.05 (bs, 2H), 2.47. LCMS: 292 (M+l)+.
Step 2: N-[4-(5-Acetyl-pyridin-2-ylsulfamoyl)-phenyl]-succinamic acid methyl ester
Figure imgf000049_0003
N-(5-acetyl-pyridin-2-yl)-4-amino-benzenesulfonamide (149 mg, 0.51 mmol) was suspended in THFVCH2Cl2 (1:1, 4 ml) before triethylamine (0.21 ml, 1.53 mmol) and DMF (0.5 ml) were added to afford a tan solution. Methyl 4-chloro-4-oxo-butyrate (0.07 ml, 0.56 mmol) was then added as a liquid. The resulting yellow mixture was then stirred at room temperature for 15 minutes, before being filtered, and the mother liquor evaporated to leave a yellow oil which was purified by HPLC. The desired compound was isolated as a white powder (126 mg, 0.31 mmol, 61 %) 1H-NMR (400 MHz, DMSO-d6) β 10.38 (s, IH), 8.64 (bs, IH), 8.12 (dd, IH), 7.85 (d, 2H), 7.72 (d, 2H), 7.16 (d, IH), 3.58 (s, 3H), 2.61 (m, 4H), 2.48 (s, 3H). LCMS: 406 (M+l)+.
Step 3: N-{4-[5-(2-Bromo-acetyl)-pyridin-2-ylsulfamoyl]-phenyl}-succinamic acid methyl ester
Figure imgf000050_0001
N-{4-[5-(2-Bromo-acetyl)-pyridin-2-ylsulfamoyl]-phenyl}-succinamic acid methyl ester was synthesized as described in EXAMPLE 1, Step 2, using N-[4-(5-Acetyl-ρyridin-2-ylsulfamoyl)- phenyl]-succinamic acid methyl ester as starting material Step 4: N-{4-[5-(2-Acetylsulfanyl-acetyl)-pyridin-2-ylsulfamoyl]-phenyl}-succinamic acid methyl ester
Figure imgf000050_0002
N-{4-[5-(2-Acetylsulfanyl-acetyl)-pyridin-2-ylsulfamoyl]-ρhenyl}-succinamic acid methyl ester was synthesized as described in EXAMPLE 1, Step 3, using N-{4-[5-(2-Bromo-acetyl)-ρyridin-2- ylsulfamoylj-phenylj-succinamic acid methyl ester as starting material. 1H-NMR (400 MHz,
DMSOd6) δ 10.38 (s, IH), 8.76 (bs, IH), 8.15 (dd, IH), 7.86 (bd, 2H), 7.73 (d, 2H), 7.18 (bs, IH), 4.41 (s, 2H), 3.58 (s, 3H), 2.61 (m, 4H), 2.36 (s, 3H). LCMS: 480 (M+l)+.
The following compounds can generally be made using the methods described above. It is expected that these compounds when made will have activity similar to those that have been made in the examples above.
Figure imgf000050_0003
Figure imgf000051_0001
The activity of the compounds of the preceding examples as HDAC inhibitors has been shown by the following assays.
Inhibition Assays
Y) In vitro HDAC-inhibition Assay:
This assay measures a compound's ability to inhibit acetyl-lysine deacetylation in vitro and was used as both a primary screening method as well as for IC50 determinations of confirmed inhibitors. The assay is performed in vitro using an HDAC enzyme source {e.g. partially purified nuclear extract or immunopurified HDAC complexes) and a proprietary fluorescent
substrate/developer system (HDAC Quantizyme Fluor de Lys Fluorescent Activity Assay, BIOMOL). The assay is run in 1,536-well Greiner white-bottom plates using the following volumes and order of addition:
Step 1: Enzyme (2.5 μL) source added to plate (from refrigerated container)
Step 2: Compounds (50 nL) added with pin transfer device
Step 3: Fluor de Lys (2.5 μL) substrate added, incubate at RT, 30 minutes
Step 4: Developer (5 μL) solution is added (containing TSA), to stop reaction
Step 5: Plate Reader - data collection
The deacetylated fluorophore is excited with 360 nm light and the emitted light
(460 nm) is detected on an automated fluorometric plate reader (Aquest, Molecular Devices).
2) Cellular Histone Hvperacetylation Assays:
These two secondary assays evaluate a compound's ability to inhibit HDAC in cells by measuring cellular histone acetylation levels. The cytoblot facilitates quantitative EC50 information for cellular HDAC inhibition. Transformed cell lines (e.g. HeLa, A549, MCF-7) are cultured under standard media and culture conditions prior to plating.
a) For Cytoblot:
Cells (approx.2,500/well) are allowed to adhere 10-24 hours to wells of a 384-well Greiner PS assay plate in media containing 1-5% serum. Cells are treated with appropriate compound and specific concentrations for 0 to 24 hours. Cells are washed once with PBS (60 μL) and then fixed (95% ethanol, 5% acetic acid or 2% PFA) for 1 minute at RT (30 μL). Cells are blocked with 1% BSA for 1 hour and washed and stained with antibody (e.g. anti-Acetylated Histone H3, Upstate Biotechnology), followed by washing and incubation with an appropriate secondary antibody conjugated to HRP or fluorophore. For luminescence assays, signal is generated using Luminol substrate (Santa Cruz Biotechnology) and detected using an Aquest plate reader (Molecular Devices).
b~) For Immunoblot:
Cells (4 x 105/well) are plated into Corning 6-well dish and allowed to adhere overnight. Cells are treated with compound at appropriate concentration for 12-18 hours at 37 degrees. Cells are washed with PBS on ice. Cells are dislodged with rubber policeman and lysed in buffer containing 25 mM Tris, pH7.6; 150 mM NaCl, 25 mM MgCl2, 1% Tween-20, and nuclei collected by centrifugation (750Og). Nuclei are washed once in 25 mM Tris, pH7.6; 10 mM EDTA, collected by centrifugation (750Og). Supernatant is removed and histones are extracted using 0.4 M HCl. Samples are centrifuged at 1400Og and supernatants are precipitated in 1 ml cold acetone. The histone pellet is dissolved in water and histones are separated and analyzed by SDS-PAGE
Coomassie and immunoblotting (anti-acetylated histone antibodies, Upstate Biotechnology) using standard techniques.
3) Differential Cytotoxicity Assay:
HDAC inhibitors display differential cytotoxicity toward certain transformed cell lines.
Cells are cultured according to standard ATCC recommended conditions that are appropriate to each cell type. Compounds were tested for their ability to kill different cell types (normal and transformed) using the ATPlite luminescence ATP detection assay system (Perkin Elmer). Assays are run in either 384-well or 1536-well Greiner PS plates. Cells (30 μL or 5 μL, respectively) are dispensed using either multichannel pipette for 384-well plates, or proprietary Kalypsys bulk liquid dispenser for 1536-well plates. Compounds added using proprietary pin-transfer device (500 nL or 5 nL) and incubated 5 to 30 hours prior to analysis. Luminescence is measured using Aquest plate reader (Molecular Devices).
Results are shown in Table 1 below.
Table 1
Figure imgf000053_0001
All references cited above are incorporated herein by reference in their entirety as if written herein.
From the foregoing description, one skilled in the art can easily ascertain the essential characteristics of this invention, and without departing from the spirit and scope thereof, can make various changes and modifications of the invention to adapt it to various usages and conditions.

Claims

CLAIMS WHAT IS CLAIMED IS:
1. A compound having a structural Formula (I)
Figure imgf000054_0001
or a pharmaceutically acceptable salt, ester, or prodrug thereof, wherein:
Gi is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, and optionally substituted 5 or 6 membered heteroaryl;
G2 is selected from the group consisting of an N-sulfonamide moiety having structure (II), an S-sulfonamide moiety having structure (III), an amide of the form -NR3C(O)-, and an amide of the form -C(O)NR3-:
Figure imgf000054_0002
G3 is selected from the group consisting of optionally substituted phenyl, optionally substituted 5 or 6 membered aryl, or optionally substituted 5 or 6 membered heteroaryl;
Ri and R2 are each independently selected from the group consisting of hydrogen, lower alkyl, halogen and perhaloalkyl, or Ri and R2 taken together may form an optionally substituted cycloalkyl or optionally substituted heterocycloalkyl;
R3 and R4 are each independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, and optionally substituted aryl;
T is selected from the group consisting Of-(Xi)nINR7C(O)(X2)H2Rs and-
Figure imgf000054_0003
nl is 0-5;
n2 is O-5;
R7 is selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted heteroalkyl, and optionally substituted lower alkoxy;
R8 is selected from the group consisting of optionally substituted lower alkyl, optionally substituted heteroalkyl, optionally substituted heterocyclyl, optionally substituted heteroaryl and -NR9Ri0;
R9 and Ri0 are each independently selected from the group consisting of hydrogen, optionally substituted lower alkyl, optionally substituted lower alkenyl, optionally substituted alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted cycloalkyl, optionally substituted heterocycloalkyl, optionally substituted cycloaklenyl, optionally substituted fused aryl, optionally substituted fused heteroaryl, optionally substituted fused heterocycloalkyl, and optionally substituted fused cycloalkyl; or R9 and Rio, taken together with the nitrogen to which they are attached, form an optionally substituted five-membered or six-membered heteroaromatic or heteroaliphatic ring; and
G6 is selected from the group consisting of hydrogen, optionally substituted acyl, optionally substituted aryl, optionally substituted alkyl, optionally substituted heteroaryl, optionally substituted alkylthio, optionally substituted arylthio and optionally substituted heteroarylthio.
2. The compound as recited in Claim 1 wherein Ge is selected from the group consisting of optionally substituted acyl and hydrogen.
3. The compound as recited in Claim 2 wherein Gi is optionally substituted 6 membered aryl or optionally substituted 6 membered heteroaryl.
4. The compound as recited in Claim 3 wherein G2 is N-sulfonamide.
5. The compound as recited in Claim 4 wherein G3 is phenyl.
6. The compound as recited in Claim 5 wherein:
Gi is pyridinyl; and
T is -CXi)HiNR7C(O)R8-; and
nl is 0.
7. The compound as recited in Claim 5 wherein:
Gi is phenyl;
T is -(Xi)nINR7C(O)R8-; and
nl is 0.
8. The compound as recited in Claim 1 wherein said compound is selected from the group consisting of Examples 30-33.
9. A compound selected from the group consisting of Examples 1-29.
10. A pharmaceutical composition comprising a compound as recited in Claim 1 together with at least one pharmaceutically acceptable carrier.
11. The compound as recited in Claim 1 wherein the compound inhibits the catalytic activity of histone deacetylase (HDAC).
12. A method of treatment of a HDAC-related disease in a patient in need thereof comprising the administration of the following in any order:
a therapeutically effective amount of a compound as recited in Claim 1 ; and b null or another chemotherapeutic agent.
13. The method as recited in Claim 12 wherein said chemotherapeutic agent is one selected from the group consisting of aromatase inhibitors, antiestrogen, anti -androgen, or a gonadorelin agonists, topoisomerase land 2 inhibitors, microtubule active agents, alkylating agents, antineoplastic antimetabolite, or platin containing compound, lipid or protein kinase targeting agents, protein or lipid phosphatase targeting agents, anti-angiogentic agents, agents that induce cell differentiation, bradykinin 1 receptor and angiotensin II antagonists, cyclooxygenase inhibitors, heparanase inhibitors, lymphokines or cytokine inhibitors, bisphosphanates, rapamycin derivatives, anti-apoptotic pathway inhibitors, apoptotic pathway agonists, PPAR agonists, inhibitors of Ras isoforms, telomerase inhibitors, protease inhibitors, metalloproteinase inhibitors, and aminopeptidase inhibitors.
14. The method as recited in Claim 13 wherein said chemotherapeutic agent is useful for the treatment of multiple myeloma and is selected from the group consisting of alkylating agents, anthracyclines , corticosteroids, IMiDs, protease inhibitors, IGF-I inhibitors, CD40 antibodies, Smac mimetics, FGF3 modulators, mTOR inhibitors, HDAC inhibitors, IKK inhibitors, P38MAPK inhibitors, HSP90 inhibitors, and akt inhitors.
15. The method as recited in Claim 14 wherein said chemotherapeutic agent is selected from the group consisting of melphalan, doxorubicin, dexamethasone, prednisone, thalidomide, lenalidomide, bortezomib, and NPI0052.
16. The method as recited in either Claim 15, wherein said disease is selected from the group consisting of a hyperproliferative condition, a neurological disorder, a cardiovascular condition, an autoimmune disease, a dermatologic disorder, and an ophthalmologic disorder.
17. The method as recited in Claim 16 wherein said hyperproliferative condition is selected from the group consisting of hematologic and norihematologic cancers.
18. The method as recited in Claim 17 wherein said hematologic cancer is selected from the group consisting of multiple myeloma, leukemias, and lymphomas.
19. The method as recited in Claim 18 wherein said hematologic cancer is multiple myeloma.
20. The method as recited in Claim 18 wherein said hematologic cancer is lymphomas.
21. The method as recited in Claim 20 wherein said lymphoma is selected from the group
consisting of cutaneous t-cell lymphoma and mantle cell lymphoma.
22. A method of inhibition of BQDAC comprising contacting HDAC with a compound as recited in Claim 1.
23. The compound as recited in Claim 1 for use in the manufacture of a medicament for the prevention or treatment of a disease or condition ameliorated by the modulation of histone deacetylase (HDAC).
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