WO2016040508A1 - Substituted pyrrolidine carboxamide compounds - Google Patents

Substituted pyrrolidine carboxamide compounds Download PDF

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Publication number
WO2016040508A1
WO2016040508A1 PCT/US2015/049225 US2015049225W WO2016040508A1 WO 2016040508 A1 WO2016040508 A1 WO 2016040508A1 US 2015049225 W US2015049225 W US 2015049225W WO 2016040508 A1 WO2016040508 A1 WO 2016040508A1
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cancer
cell
tumor
carcinoma
lymphoma
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French (fr)
Inventor
Megan Alene Cloonan FOLEY
Kevin Wayne Kuntz
Lorna Helen Mitchell
Michael John Munchhof
Darren Martin Harvey
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Epizyme Inc
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Epizyme Inc
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Priority to US15/510,583 priority Critical patent/US20170247326A1/en
Priority to AU2015315177A priority patent/AU2015315177A1/en
Priority to JP2017512983A priority patent/JP2017528461A/en
Priority to CA2960277A priority patent/CA2960277A1/en
Priority to EP15839644.0A priority patent/EP3193609A4/en
Publication of WO2016040508A1 publication Critical patent/WO2016040508A1/en
Anticipated expiration legal-status Critical
Priority to US16/423,618 priority patent/US20200048195A1/en
Ceased legal-status Critical Current

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    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
    • C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
    • C07D207/04—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
    • C07D207/10—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no 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, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
    • C07D207/14—Nitrogen atoms not forming part of a nitro radical
    • 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
    • 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
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D403/00—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00
    • C07D403/02—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings
    • C07D403/12—Heterocyclic compounds containing two or more hetero rings, having nitrogen atoms as the only ring hetero atoms, not provided for by group C07D401/00 containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D405/00—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom
    • C07D405/02—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings
    • C07D405/12—Heterocyclic compounds containing both one or more hetero rings having oxygen atoms as the only ring hetero atoms, and one or more rings having nitrogen as the only ring hetero atom containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D413/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms
    • C07D413/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings
    • C07D413/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and oxygen atoms as the only ring hetero atoms containing two hetero rings linked by a chain containing hetero atoms as chain links
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07D—HETEROCYCLIC COMPOUNDS
    • C07D417/00—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00
    • C07D417/02—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings
    • C07D417/12—Heterocyclic compounds containing two or more hetero rings, at least one ring having nitrogen and sulfur atoms as the only ring hetero atoms, not provided for by group C07D415/00 containing two hetero rings linked by a chain containing hetero atoms as chain links

Definitions

  • the present disclosure provides substituted pyrrolidine carboxamides as SMYD protein inhibitors, such as SMYD3 and SMYD2 inhibitors, and therapeutic methods of treating conditions and diseases wherein inhibition of SMYD proteins such as SMYD3 and SMYD2 provides a benefit.
  • Epigenetic regulation of gene expression is an important biological determinant of protein production and cellular differentiation and plays a significant pathogenic role in a number of human diseases.
  • Epigenetic regulation involves heritable modification of genetic material without changing its nucleotide sequence.
  • epigenetic regulation is mediated by selective and reversible modification (e.g., methylation) of DNA and proteins (e.g., histones) that control the conformational transition between transcriptionally active and inactive states of chromatin.
  • methyltransferases e.g., SMYD proteins such as SMYD3 and SMYD2
  • the present disclosure provides a Compound of the Disclosure and one or more pharmaceutically acceptable carriers.
  • the present disclosure provides a method of inhibiting SMYD proteins, such as SMYD3 or SMYD2, or both, in a mammal, comprising administering to the mammal an effective amount of at least one Compound of the Disclosure.
  • SMYD proteins such as SMYD3 or SMYD2
  • the present disclosure provides a method of irreversibly inhibiting SMYD proteins, such as SMYD3 or SMYD2, or both, in a mammal, comprising administering to the mammal an effective amount of at least one Compound of the Disclosure.
  • the present disclosure provides methods for treating a disease, disorder, or condition, e.g., cancer, responsive to inhibition of SMYD proteins, such as SMYD3 or SMYD2, or both, comprising administering a therapeutically effective amount of a Compound of the Disclosure.
  • a disease, disorder, or condition e.g., cancer
  • SMYD proteins such as SMYD3 or SMYD2
  • the present disclosure provides the use of Compounds of the Disclosure as inhibitors of SMYD3.
  • the present disclosure provides the use of Compounds of the Disclosure as inhibitors of SMYD2.
  • the present disclosure provides the use of Compounds of the Dicslosure as inhibitors of SMYD proteins.
  • the present disclosure provides a pharmaceutical composition for treating a disease, disorder, or condition responsive to inhibition of SMYD proteins, such as SMYD3 or SMYD2, or both, wherein the pharmaceutical composition comprises a therapeutically effective amount of a Compound of the Disclosure in a mixture with one or more pharmaceutically acceptable carriers.
  • the present disclosure provides Compounds of the Disclosure for use in treating cancer in a mammal, e.g., breast, cervical, colon, kidney, liver, head and neck, skin, pancreatic, ovary, esophageal, lung, and prostate cancer.
  • a mammal e.g., breast, cervical, colon, kidney, liver, head and neck, skin, pancreatic, ovary, esophageal, lung, and prostate cancer.
  • the present disclosure provides a Compound of the Disclosure for use in the ma dicament for treating cancer in a mammal.
  • the p ovides kit comprising a Compound of the Disclosure.
  • One aspect of the present disclosure is based on the use of Compounds of the Disclosure as inhibitors of SMYD3.
  • the Compounds of the Disclosure are useful for treating diseases, disorders, or conditions, e.g., cancer, responsive to inhibition of SMYD3.
  • One aspect of the present disclosure is based on the use of Compounds of the Disclosure as inhibitors of SMYD2.
  • the Compounds of the Disclosure are useful for treating diseases, disorders, or conditions, e.g., cancer, responsive to inhibition of SMYD2.
  • Compounds of the Disclosure are compounds having Formula I:
  • A is selected from the group consisting of 1,2,3-triazolyl, 1,2,4-triazolyl, 1,3,4- oxadiazole, 1,3,4-thiadiazole, 1-imidazolyl, 1-isoquinolinyl, 1-pyrazolyl, 2-(1,2,3,4- tetrahydroquinolinyl), 2-benzo[ nzo[d]thiazolyl, 2-chromenyl-4-one, 2-furanyl, 2-imidazo[1,2-b]pyr olyl, 2-indolyl, 2-naphthalenyl, 2- pyrazinyl, 2-pyridyl, 2-pyrim nyl, 2-pyrrolyl, 2-quinolinyl, 2- quinoxalinyl, 2-thiazolo[5,4-c azolyl, 2-thiophenyl, 3-(1,2,3,4- tetrahydroisoquinoline), 3-(1,2,4-oxadiazolyl), 3-imidazo[1,2-a
  • Y is -C(R 1a )(R 1b )-; or Y is absent, i.e., A forms a bond with the carbonyl carbon atom;
  • R 1s and R 1b are independently selected from the group consisting of hydrogen and C 1-4 alkyl;
  • R 5 is selected from the group consisting of hydrogen and C 1-4 alkyl
  • R 6 is selected from the group consisting of hydrogen and C 1-4 alkyl
  • R 7 is selected from the group consisting of hydrogen, C 1-4 alkyl, amino, alkylamino, dialkylamino, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, and hydroxyalkyl;
  • R 8a is selected from the group consisting of C 1-6 alkyl, haloalkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally substituted C 6-14 aryl, optionally substituted 4- to 14-membered heterocyclo, optionally substituted 5- to 14-membered heteroaryl, optionally substituted C 3-12 cycloalkyl, aralkyl, and heteroaralkyl; and
  • R 8b is selected from the group consisting of C 1-6 alkyl, haloalkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally aryl, optionally substituted C 4-14 heterocyclo, optionally subs membered heteroaryl, optionally substituted C 3-12 cycloalkyl, ara kyl.
  • e Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein A is selected from the group consisting of 1,2,3-triazolyl, 1,2,4-triazolyl, 1-imidazolyl, 1-isoquinolinyl, 1-pyrazolyl, 2-(1,2,3,4- tetrahydroquinolinyl), 2-benzo[d]imidazolyl, 2-benzo[d]thiazolyl, 2-chromenyl-4-one, 2-furanyl, 2-imidazo[1,2-b]pyridazinyl, 2-imidazolyl, 2-indolyl, 2-naphthalenyl, 2- pyrazinyl, 2-pyridyl, 2-pyrimidinyl, 2-pyrrolidinyl, 2-pyrrolyl, 2-quinolinyl, 2- quinoxalinyl, 2-thiazolo[5,4-c]pyridinyl, 2-
  • Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein B is selected from the group consisting of: .
  • Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein B is selected from the group consisting of:
  • R 4a is selected from the group consisting of hydroxy, C 1-6 alkyl, alkoxy, hydroxyalkyl, C 3-12 cycloalkyl, optionally substituted C 6-14 aryl, alkoxyalkyl, and aralkyl. In another embodiment, R 4a is C 1-4 alkyl. In another embodiment, R 4a is methyl.
  • X is absent.
  • Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein Z is selected from the group consisting of optionally substituted C 1-6 alkyl, hydroxyalkyl, (a lamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (hetero ally substituted C 6-14 aryl, optionally substituted 4- to 14-membered nally substituted 5- to 14-membered heteroaryl, optionally substituted C 3-12 cycloalkyl, aralkyl, and heteroaralkyl.
  • Z is selected from the group consisting of optionally substituted C 1-6 alkyl, hydroxyalkyl, (a lamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (hetero ally substituted C 6-14 aryl, optionally substituted 4- to 14-membere
  • Z is selected from the group consisting of C 1-6 alkyl and optionally substituted C 3-12 cycloalkyl. In another embodiment, Z is cyclopropyl.
  • Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein Y is -CH 2 -.
  • Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein Y is absent.
  • Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein A is selected from the group consisting of 5-indolinyl-2-one, 6-benzo[b][1,4]oxazinyl-3-one, 5-pyrazolyl, 3-isothiazolyl, 6-benzo[d]oxazolyl-2(3H)- one, 1,2,3-triazolyl, phenyl, and 5-(1,3-dihydro-2H-benzo[d]imidazolyl-2-one).
  • A is 5-indolinyl-2-one.
  • Compounds of the Disclosure are compounds of Tables 1, 3, and 4, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, or different pharmaceutically acceptable salt thereof.
  • the terms used in connection with A have the chemical structures set forth in Table 2, each of which may be optionally substituted with one or more substituents, e.g., 1, 2, 3, 4, or 5 substituents, depending on the nature of the group and the number of available positions.
  • substituents e.g., 1, 2, 3, 4, or 5 substituents, depending on the nature of the group and the number of available positions.
  • A 2-furanyl there are three carbon atoms for available for substitution.
  • a is 2-naphthalenyl there are seven carbon atoms available for substitution. Substitution may occur at any available carbon or nitrogen atom.
  • Optional substituents include, but are not limited to, halo, hydroxy, alkoxy, amino, alkylamino, dialkylamino, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, C 1-6 alkyl, haloalkyl, hydroxyalkyl, (carboxamido)alkyl, (cycloalkyl)alkyl, optionally substituted C 3-12 cycloalkyl, optionally substituted C 6-14 aryl, optionally substituted 5- to 14-membered heteroaryl, optionally substituted 4- to 14-membered heterocyclo, or aralkyl.
  • alkyl refers to a straight- or branched-chain aliphatic hydrocarbon containing one to twelve carbon atoms (i.e., C 1-12 alkyl) or the number of carbon atoms designated (i.e., a C 1 alkyl such as methyl, a C 2 alkyl such as ethyl, a C 3 alkyl such as propyl or isopropyl, etc.).
  • the alkyl group is chosen from a straight chain C 1-10 alkyl group.
  • the alkyl group is chosen from a branched chain C 3-10 alkyl group.
  • the alkyl group is chosen from a straight chain C 1-6 alkyl group iment, the alkyl group is chosen from a branched chain C 3-6 alkyl gro bodiment, the alkyl group is chosen from a straight chain C 1-4 alkyl mbodiment, the alkyl group is chosen from a branched chain C 3-4 alky g p her embodiment, the alkyl group is chosen from a straight or branched chain C 3-4 alkyl group.
  • the alkyl group is partially or completely deuterated, i.e., one or more hydrogen atoms of the alkyl group are replaced with deuterium atoms.
  • Non-limiting exemplary C 1-10 alkyl groups include methyl (including -CD 3 ), ethyl, propyl, isopropyl, butyl, sec-butyl, tert-butyl, iso-butyl, 3-pentyl, hexyl, heptyl, octyl, nonyl, and decyl.
  • Non-limiting exemplary C 1-4 alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, tert-butyl, and iso-butyl.
  • the term "optionally substituted alkyl" as used by itself or as part of another group means that the alkyl as defined above is either unsubstituted or substituted with one, two, or three substituents independently chosen from nitro, haloalkoxy, aryloxy, aralkyloxy, alkylthio, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, alkoxycarbonyl, and carboxyalkyl.
  • the alkyl is a C 1-4 alkyl.
  • the optionally substituted alkyl is substituted with two substituents. In another embodiment, the optionally substituted alkyl is substituted with one substituent.
  • Non-limiting exemplary optionally substituted alkyl groups include -CH 2 CH 2 NO 2 , -CH 2 CH 2 CO 2 H, -CH 2 CH 2 SO 2 CH 3 , -CH 2 CH 2 COPh, and -CH 2 C 6 H 11 .
  • cycloalkyl refers to saturated and partially unsaturated (containing one or two double bonds) cyclic aliphatic hydrocarbons containing one to three rings having from three to twelve carbon atoms (i.e., C 3-12 cycloalkyl) or the number of carbons designated.
  • the cycloalkyl group has two rings.
  • the cycloalkyl group has one ring.
  • the cycloalkyl group is chosen from a C 3-8 cycloalkyl group.
  • the cycloalkyl group is chosen from a C 3-6 cycloalkyl group.
  • Non-limiting exemplary cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, norbornyl, decalin, adamantyl, cyclohexenyl, and spiro[3.3]heptane.
  • the term "optionally substituted cycloalkyl" as used by itself or as part of another group means that the cycloalkyl as defined above is either unsubsti d with one, two, or three substituents independently chosen from no, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydrox oalkoxy, aryloxy, aralkyl, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclo, alkoxyalkyl, (a
  • the optionally substituted cycloalkyl is substituted with two substituents. In another embodiment, the optionally substituted cycloalkyl is substituted with one substituent. In one embodiment, the optionally substituted cycloalkyl is substituted with at least one amino, alkylamino, or dialkylamino group.
  • Non-limiting exemplary optionally substituted c cloalk l rou s include:
  • cycloalkenyl as used by itself or part of another group refers aturated cycloalkyl group as defined above.
  • the cycloalkenyl has one carbon-to-carbon double bond.
  • the cycloalkenyl group is chosen from a C 4-8 cycloalkenyl group.
  • Exemplary cycloalkenyl groups include cyclopentenyl and cyclohexenyl.
  • the term "optionally substituted cycloalkenyl" as used by itself or as part of another group means that the cycloalkenyl as defined above is either unsubstituted or substituted with one, two, or three substituents independently chosen from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, monohydroxyalkyl, dihydroxyalkyl, alkoxy, haloalkoxy, aryloxy, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl, optional
  • the optionally substituted cycloalkenyl is substituted with two substituents. In another embodiment, the optionally substituted cycloalkenyl is substituted with one substituent. In another embodiment, the cycloalkenyl is unsubstituted.
  • alkenyl refers to an alkyl group as defined above containing one, two or three carbon-to-carbon double bonds.
  • the alkenyl group is chosen from a C 2-6 alkenyl group.
  • the alkenyl group is chosen from a C 2-4 alkenyl group.
  • Non-limiting exemplary alkenyl groups include ethenyl, propenyl, isopropenyl, butenyl, sec-butenyl, pentenyl, and hexenyl.
  • the term "optionally substituted alkenyl” as used herein by itself or as part of another group means the alkenyl as defined above is either unsubstituted or substituted with one, two or three substituents independently chosen from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aryloxy, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino carboxy carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, opt aryl, optionally substituted heteroaryl, or optionally substituted heteroc
  • alkynyl refers to an alkyl group as defined above containing one to three carbon-to-carbon triple bonds.
  • the alkynyl has one carbon-to-carbon triple bond.
  • the alkynyl group is chosen from a C 2-6 alkynyl group.
  • the alkynyl group is chosen from a C 2-4 alkynyl group.
  • Non-limiting exemplary alkynyl groups include ethynyl, propynyl, butynyl, 2-butynyl, pentynyl, and hexynyl groups.
  • the term "optionally substituted alkynyl" as used herein by itself or as part of another group means the alkynyl as defined above is either unsubstituted or substituted with one, two or three substituents independently chosen from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aryloxy, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl, optionally substituted heteroaryl, or optionally
  • haloalkyl refers to an alkyl group substituted by one or more fluorine, chlorine, bromine and/or iodine atoms.
  • the alkyl group is substituted by one, two, or three fluorine and/or chlorine atoms.
  • the haloalkyl group is chosen from a C 1-4 haloalkyl group.
  • Non-limiting exemplary haloalkyl groups include fluoromethyl, difluoromethyl, trifluoromethyl, pentafluoroethyl, 1,1-difluoroethyl, 2,2-difluoroethyl, 2,2,2-trifluoroethyl, 3,3,3-trifluoropropyl, 4,4,4-trifluorobutyl, and trichloromethyl groups.
  • hydroxyalkyl refers to an alkyl group substituted with one or more, e.g., one, two, or three, hydroxy groups.
  • the hydroxyalkyl group is a monohydroxyalkyl group, i.e., substituted with one hydroxy group.
  • the hydroxyalkyl group is a dihydroxyalkyl group, i.e., substituted with two hydroxy groups.
  • the hydroxyalkyl group is chosen from a C 1-4 hydroxyalkyl group.
  • N lary hydroxyalkyl groups include hydroxymethyl, hydroxyethyl, nd hydroxybutyl groups, such as 1-hydroxyethyl, 2-hydroxyethyl, l, 2-hydroxypropyl, 3-hydroxypropyl, 3-hydroxybutyl, 4-hydroxybutyl, 2-hydroxy-1-methylpropyl, and 1,3-dihydroxyprop-2- yl.
  • alkoxy refers to an optionally substituted alkyl, optionally substituted cycloalkyl, optionally substituted alkenyl or optionally substituted alkynyl attached to a terminal oxygen atom.
  • the alkoxy group is chosen from a C 1-4 alkoxy group.
  • the alkoxy group is chosen from a C 1-4 alkyl attached to a terminal oxygen atom, e.g., methoxy, ethoxy, and tert-butoxy.
  • alkylthio refers to a sulfur atom substituted by an optionally substituted alkyl group.
  • the alkylthio group is chosen from a C 1-4 alkylthio group.
  • Non-limiting exemplary alkylthio groups include -SCH 3 , and -SCH 2 CH 3 .
  • alkoxyalkyl refers to an alkyl group substituted with an alkoxy group.
  • Non-limiting exemplary alkoxyalkyl groups include methoxymethyl, methoxyethyl, methoxypropyl, methoxybutyl, ethoxymethyl, ethoxyethyl, ethoxypropyl, ethoxybutyl, propoxymethyl, iso-propoxymethyl, propoxyethyl, propoxypropyl, butoxymethyl, tert- butoxymethyl, isobutoxymethyl, sec-butoxymethyl, and pentyloxymethyl.
  • haloalkoxy as used by itself or as part of another group refers to a haloalkyl attached to a terminal oxygen atom.
  • Non-limiting exemplary haloalkoxy groups include fluoromethoxy, difluoromethoxy, trifluoromethoxy, and 2,2,2-trifluoroethoxy.
  • heteroalkyl refers to a stable straight or branched chain hydrocarbon radical containing 1 to 10 carbon atoms and at least two heteroatoms, which can be the same or different, selected from O, N, or S, wherein: 1) the nitrogen atom(s) and sulfur atom(s) can optionally be oxidized; and/or 2) the nitrogen atom(s) can optionally be quaternized.
  • the heteroatoms can be placed at any interior position of the heteroalkyl group or at a position at which the heteroalkyl group is attached to the remainder of the molecule.
  • the heteroalkyl group contains two oxygen atoms.
  • the heteroalkyl contains one ox ogen atom. In one embodiment, the heteroalkyl contains two nitrog miting exemplary heteroalkyl groups include -CH 2 OCH 2 CH 2 OCH 3 , CH 2 OCH 3 , -CH 2 NHCH 2 CH 2 OCH 2 , -OCH 2 CH 2 NH 2 , -NHCH 2 CH 2 N(H)CH 3 , -NHCH 2 CH 2 OCH 3 and -OCH 2 CH 2 OCH 3 .
  • aryl refers to a monocyclic or bicyclic aromatic ring system having from six to fourteen carbon atoms (i.e., C 6-14 aryl).
  • Non-limiting exemplary aryl groups include phenyl (abbreviated as "Ph"), naphthyl, phenanthryl, anthracyl, indenyl, azulenyl, biphenyl, biphenylenyl, and fluorenyl groups.
  • the aryl group is chosen from phenyl or naphthyl.
  • the aryl group is phenyl.
  • the term "optionally substituted aryl" as used herein by itself or as part of another group means that the aryl as defined above is either unsubstituted or substituted with one to five substituents independently selected from the group consisting of halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aryloxy, heteroaryloxy, aralkyl, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl,
  • the optionally substituted aryl is an optionally substituted phenyl. In one embodiment, the optionally substituted phenyl has four substituents. In another embodiment, the optionally substituted phenyl has three substituents. In another embodiment, the optionally substituted phenyl has two substituents. In another embodiment, the optionally substituted phenyl has one substituent. In another embodiment, the optionally substituted phenyl has one amino, alkylamino, dialkylamino, (amino)alkyl, (alkylamino)alkyl, or (dialkylamino)alkyl substituent.
  • Non-limiting exemplary substituted aryl groups include 2-methylphenyl, 2-methoxyphenyl, 2-fluorophenyl 2 chlorophenyl, 2-bromophenyl, 3-methylphenyl, 3-methoxyphenyl, 3-fluorophe yl, 4-methylphenyl, 4-ethylphenyl, 4-methoxyphenyl, 4-fluoroph nyl, 2,6-di-fluorophenyl, 2,6-di- chlorophenyl, 2-methyl, 3-m -ethyl, 3-methoxyphenyl, 3,4-di- methoxyphenyl, 3,5-di-fluorophenyl 3,5-di-methylphenyl, 3,5-dimethoxy, 4-methylphenyl, 2-fluoro-3-chlorophenyl, 3-chloro-4-fluorophenyl, and 2-phenylpropan- 2-amine.
  • the term optionally substituted aryl is meant to include groups having fused optionally substitute
  • aryloxy as used by itself or as part of another group refers to an optionally substituted aryl attached to a terminal oxygen atom.
  • a non-limiting exemplary aryloxy group is PhO-.
  • heteroaryloxy refers to an optionally substituted heteroaryl attached to a terminal oxygen atom.
  • aralkyloxy or "arylalkyloxy” as used by itself or as part of another group refers to an aralkyl group attached to a terminal oxygen atom.
  • a non-limiting exemplary aralkyloxy group is PhCH 2 O-.
  • heteroaryl refers to monocyclic and bicyclic aromatic ring systems having 5 to 14 ring atoms (i.e., a 5- to 14-membered heteroaryl) and 1, 2, 3, or 4 heteroatoms independently chosen from oxygen, nitrogen or sulfur.
  • the heteroaryl has three heteroatoms.
  • the heteroaryl has two heteroatoms.
  • the heteroaryl has one heteroatom.
  • the heteroaryl is a 5- to 10-membered heteroaryl.
  • the heteroaryl has 5 ring atoms, e.g., thienyl, a 5-membered heteroaryl having four carbon atoms and one sulfur atom.
  • the heteroaryl has 6 ring atoms, e.g., pyridyl, a 6-membered heteroaryl having five carbon atoms and one nitrogen atom.
  • Non- limiting exemplary heteroaryl groups include thienyl, benzo[b]thienyl, naphtho[2,3- b]thienyl, thianthrenyl, furyl, benzofuryl, pyranyl, isobenzofuranyl, benzooxazonyl, chromenyl, xanthenyl, 2H-pyrrolyl pyrrolyl imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, isoind dolyl, indazolyl, purinyl, isoquinolyl, quinolyl, phthalazinyl, naphth yl, quinazolinyl, pteridinyl, 4aH- carbazolyl, carbazolyl, ⁇ -ca thridinyl, acridinyl, pyrimidinyl, phenanthrolinyl
  • the heteroaryl is chosen from thienyl (e.g., thien- 2-yl and thien-3-yl), furyl (e.g., 2-furyl and 3-furyl), pyrrolyl (e.g., 1H-pyrrol-2-yl and 1H-pyrrol-3-yl), imidazolyl (e.g., 2H-imidazol-2-yl and 2H-imidazol-4-yl), pyrazolyl (e.g., 1H-pyrazol-3-yl, 1H-pyrazol-4-yl, and 1H-pyrazol-5-yl), pyridyl (e.g., pyridin-2-yl, pyridin-3-yl, and pyridin-4-yl), pyrimidinyl (e.g., pyrimidin-2-yl, pyrimidin-4-yl, and pyrimidin-5-yl), thiazolyl (e.
  • the term "optionally substituted heteroaryl" as used by itself or as part of another group means that the heteroaryl as defined above is either unsubstituted or substituted with one to four substituents, e.g., one or two substituents, independently chosen from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aralkyl, aryloxy, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted ary
  • the optionally substituted heteroaryl has one substituent.
  • the optionally substituted is an optionally substituted pyridyl, i.e 2 3 or 4 pyridyl. Any available carbon or nitrogen atom can be substituted.
  • heterocycle or “heterocyclo” as used by itself or as part of an to saturated and partially unsaturated (e.g., containing one or two double bonds) cyclic groups containing one, two, or three rings having from three to fourteen ring members (i.e., a 3- to 14-membered heterocyclo) and at least one heteroatom.
  • Each heteroatom is independently selected from the group consisting of oxygen, sulfur, including sulfoxide and sulfone, and/or nitrogen atoms, which can be quaternized.
  • heterocyclo is meant to include cyclic ureido groups such as imidazolidinyl-2-one, cyclic amide groups such as ⁇ -lactam, ⁇ -lactam, ⁇ -lactam and ⁇ -lactam, and cyclic carbamate groups such as oxazolidinyl-2-one.
  • heterocyclo is also meant to include groups having fused optionally substituted aryl groups, e.g., indolinyl, indolinyl-2-one, benzo[d]oxazolyl-2(3H)-one.
  • the heterocyclo group is chosen from a 4-, 5-, 6-, 7- or 8-membered cyclic group containing one ring and one or two oxygen and/or nitrogen atoms. In one embodiment, the heterocyclo group is chosen from a 5- or 6-membered cyclic group containing one ring and one or two nitrogen atoms. In one embodiment, the heterocyclo group is chosen from a 8-, 9-, 10-, 11-, or 12-membered cyclic group containing two rings and one or two nitrogen atoms. The heterocyclo can be optionally linked to the rest of the molecule through a carbon or nitrogen atom.
  • Non-limiting exemplary heterocyclo groups include 2-oxopyrrolidin-3-yl, 2-imidazolidinone, piperidinyl, morpholinyl, piperazinyl, pyrrolidinyl, 8-azabicyclo[3.2.1]octane (nortropane), 6-azaspiro[2.5]octane, 6-azaspiro[3.4]octane, indolinyl, indolinyl-2-one, 1,3-dihydro-2H-benzo[d]imidazol-2- one
  • the term "optionally substituted heterocyclo" as used herein by itself or part of another group means the heterocyclo as defined above is either unsubstituted or substituted with one to four substituents independently selected from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aryloxy, aralkyl aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclo, alkoxyalkyl, (amino)al
  • Substitution may occur on any available carbon or nitrogen ato spirocycle.
  • the optionally substituted heterocyc th at least one amino, alkylamino, or dialkylamino group.
  • Non-limit onally substituted heterocyclo groups include:
  • amino as used by itself or as part of another group refers to -NH 2 .
  • alkylamino as used by itself or as part of another group refers to -NHR 22 , wherein R 22 is C 1-6 alkyl. In one embodiment, R 22 is C 1-4 alkyl.
  • Non-limiting exemplary alkylamino groups include -N(H)CH 3 and -N(H)CH 2 CH 3 .
  • dialkylamino as used by itself or as part of another group refers to -NR 23a R 23b , wherein R 23a and R 23b are each independently C 1-6 alkyl. In o 23a and R 23b are each independently C 1-4 alkyl.
  • Non-limiting exe no groups include -N(CH 3 ) 2 and -N(CH 3 )CH 2 CH(CH 3 ) 2 .
  • hydroxyalkylamino as used by itself or as part of another group refers to -NHR 24 , wherein R 24 is hydroxyalkyl.
  • cycloalkylamino as used by itself or as part of another group refers to -NR 25a R 25b , wherein R 25a is optionally substituted cycloalkyl and R 25b is hydrogen or C 1-4 alkyl.
  • aralkylamino as used by itself or as part of another group refers to -NR 26a R 26b , wherein R 26a is aralkyl and R 26b is hydrogen or C 1-4 alkyl.
  • Non-limiting exemplary aralkylamino groups include -N(H)CH 2 Ph and -N(CH 3 )CH 2 Ph.
  • (amino)alkyl refers to an alkyl group substituted with an amino group.
  • the alkyl is a C 1-4 alkyl.
  • Non-limiting exemplary (amino)alkyl groups include -CH 2 NH 2 , -C(NH 2 )(H)CH 3 , -CH 2 CH 2 NH 2 , -CH 2 C(NH 2 )(H)CH 3 , -CH 2 CH 2 CH 2 NH 2 , -CH 2 CH 2 CH 2 CH 2 NH 2 , and -CH 2 C(CH 3 ) 2 CH 2 NH 2
  • (alkylamino)alkyl refers to an alkyl group substituted with an alkylamino group.
  • the alkyl is a C 1-4 alkyl.
  • a non-limiting exemplary (alkylamino)alkyl group is -CH 2 CH 2 N(H)CH 3 .
  • dialkylamino)alkyl refers to an alkyl group substituted by a dialkylamino group.
  • the alkyl is a C 1-4 alkyl.
  • Non-limiting exemplary (dialkylamino)alkyl groups are -CH 2 CH 2 N(CH 3 ) 2 .
  • (cycloalkylamino)alkyl refers to an alkyl group substituted by a cycloalkylamino group.
  • the alkyl is a C 1-4 alkyl.
  • Non-limiting exemplary (cycloalkylamino)alkyl groups include -CH 2 N(H)cyclopropyl, -CH 2 N(H)cyclobutyl, and -CH 2 N(H)cyclohexyl.
  • (aralkylamino)alkyl refers to an alkyl group substituted with an aralkylamino group.
  • the alkyl is a C 1-4 alkyl.
  • (cyano)alkyl as used by itself or as part of another group refe substituted with one or more cyano, e.g., -CN, groups.
  • the alkyl is a C 1-4 alkyl.
  • Non-limiting exemplary (cyano)alkyl groups include -CH 2 CH 2 CN, -CH 2 CH 2 CH 2 CN, and -CH 2 CH 2 CH 2 CH 2 CN.
  • (amino)(hydroxy)alkyl refers to an alkyl group substituted with one amino, alkylamino, or dialkylamino group and one hydroxy group.
  • the alkyl is a C 1-6 alkyl.
  • the alkyl is a C 1-4 alkyl.
  • Non-limiting exemplary (amino)(hydroxy)alkyl groups include:
  • (amino)(aryl)alkyl refers to an alkyl group substituted with one amino, alkylamino, or dialkylamino group and one optionally substituted aryl group.
  • the alkyl is a C 1-6 alkyl.
  • the optionally substituted aryl group is an optionally substituted phenyl.
  • Non-limiting exemplary (amino)(aryl)alkyl groups include:
  • (cycloalkyl)alkyl refers to an alkyl group substituted with one optionally substituted cycloalkyl group.
  • the alkyl is a C 1-4 alkyl.
  • the cycloalkyl is a C 3-6 cycloalkyl.
  • the optionally substituted cycloalkyl group is substituted with an amino or (amino)alkyl group.
  • Non-limiting exemplary (cycloalk l) lk l i clude:
  • (hydroxy)(aryl)alkyl refers to an alkyl group substituted with one hydroxy group and one optionally substituted aryl group.
  • the alkyl is a C- 1 -6 alkyl.
  • the optionally substituted aryl group is an optionally substituted hen l.
  • Non-limitin exem lar h drox ar l alk l rou s include:
  • Non-limiting exemplary carboxamido groups include -CONH 2 , -CON(H)CH 3 , CON(CH 3 ) 2 , and -CON(H)Ph.
  • (carboxamido)alkyl as used by itself or as part of anothe an alkyl group substituted with a carboxamido group.
  • Non-lim (carboxamido)alkyl groups include -CH 2 CONH 2 , -C(H)CH 3 -CONH )CH 3 .
  • sulfonamido refers to a radical of the formula -SO 2 NR 27a R 27b , wherein R 27a and R 27b are each independently hydrogen, optionally substituted alkyl, or optionally substituted aryl, or R 27a and R 27b taken together with the nitrogen to which they are attached from a 3- to 8-membered heterocyclo group.
  • Non-limiting exemplary sulfonamido groups include -SO 2 NH 2 , -SO 2 N(H)CH 3 , and -SO 2 N(H)Ph.
  • a non-limiting exemplary alkylcarbonyl group is -COCH 3 .
  • a non-limiting exemplary arylcarbonyl group is -COPh.
  • alkylsulfonyl as used by itself or as part of another group refers to a sulfonyl group, i.e., -SO 2 -, substituted by any of the above-mentioned optionally substituted alkyl groups.
  • a non-limiting exemplary alkylsulfonyl group is -SO 2 CH 3 .
  • arylsulfonyl as used by itself or as part of another group refers to a sulfonyl group, i.e., -SO 2 -, substituted by any of the above-mentioned optionally substituted aryl groups.
  • a non-limiting exemplary arylsulfonyl group is -SO 2 Ph.
  • mercaptoalkyl as used by itself or as part of another group refers to any of the above-mentioned alkyl groups substituted by a–SH group.
  • carboxy as used by itself or as part of another group refers to a radical of the formula -COOH.
  • carboxyalkyl as used by itself or as part of another group refers to any of the above-mentioned alkyl groups substituted with a -COOH.
  • a non limiting exemplary carboxyalkyl group is -CH 2 CO 2 H.
  • Non-limiti xycarbonyl groups are -CO 2 Me and -CO 2 Et.
  • aralkyl or“arylalkyl” as used by itself or as part of another group refers to an alkyl group substituted with one, two, or three optionally substituted aryl groups.
  • the aralkyl group is a C 1-4 alkyl substituted with one optionally substituted aryl group.
  • Non-limiting exemplary aralkyl groups include benzyl, phenethyl, -CHPh 2 , -CH 2 (4-OH-Ph), and -CH(4-F-Ph) 2 .
  • the term "(heterocyclo)alkyl” as used by itself or as part of another group refers to an alkyl group substituted with one, two, or three optionally substituted heterocyclo groups.
  • the (heterocyclo)alkyl is a C 1-4 alkyl substituted with one optionally substituted heterocyclo group.
  • the heterocyclo can be linked to the alkyl group through a carbon or nitrogen atom.
  • Non-limiting exemplary (heterocyclo)alkyl groups include:
  • (heteroaryl)alkyl refers to an alkyl group substituted with one, two, or three optionally substituted heteroaryl groups.
  • the (heteroaryl)alkyl group is a C 1-4 alkyl substituted with one optionally substituted heteroaryl group.
  • Non-limiting exemplary (heteroaryl)alkyl groups include: ,
  • alkylcarbonylamino as used by itself or as part of another group refers to an alkylcarbonyl group attached to an amino.
  • a non-limiting exemplary alkylcarbonylamino group is -NHCOCH 3 .
  • the present disclosure encompasses any of the Compounds of the Disclosure being isotopically-labelled (i.e., radiolabeled) by having one or more atoms replaced by an atom having a different atomic mass or mass number.
  • isotopes that can be incorporated into the disclosed compounds include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine and chlorine, such as 2 H (or deuterium (D)), 3 H, 1 1 C, 13 C, 14 C, 15 N, 18 O, 17 O, 31 P, 32 P, 35 S, 18 F, and 36 Cl, respectively, e.g., 3 H, 11 C, and 14 C.
  • compositions wherein substantially all of the atoms at a position within the Compound of the Disclosure are replaced by an atom having a different atomic mass or mass number.
  • Isotopically-labelled Compounds of the Disclosure can be prepared by methods known in the art.
  • Compounds of the Disclosure may contain one or more asymmetric centers and may thus give rise to enantiomers, diastereomers, and other stereoisomeric forms.
  • the present disclosure is meant to encompass the use of all such possible forms, as well as their racemic and resolved form eof.
  • the individual enantiomers can be separated according to meth art in view of the present disclosure.
  • the compounds described finic double bonds or other centers of geometric asymmetry, and unle p wise it is intended that they include both E and Z geometric isomers. All tautomers are intended to be encompassed by the present disclosure as well.
  • stereoisomers is a general term for all isomers of individual molecules that differ only in the orientation of their atoms in space. It includes enantiomers and isomers of compounds with more than one chiral center that are not mirror images of one another (diastereomers).
  • chiral center or "asymmetric carbon atom” refers to a carbon atom to which four different groups are attached.
  • enantiomer and “enantiomeric” refer to a molecule that cannot be superimposed on its mirror image and hence is optically active wherein the enantiomer rotates the plane of polarized light in one direction and its mirror image compound rotates the plane of polarized light in the opposite direction.
  • racemic refers to a mixture of equal parts of enantiomers and which mixture is optically inactive.
  • absolute configuration refers to the spatial arrangement of the atoms of a chiral molecular entity (or group) and its stereochemical description, e.g., R or S.
  • enantiomeric excess refers to a measure for how much of one enantiomer is present compared to the other.
  • the percent enantiomeric excess is defined as ([D] obs /[D] max )*100, where [D] obs is the optical rotation of the mixture of enantiomers and [D] max is the optical rotation of the pure enantiomer. Determination of enantiomeric excess is possible using a variety of analytical techniques, including NMR spectroscopy, chiral column chromatography or optical polarimetry.
  • enantiomeric ll refers to a sample of a chiral substance all of whose molecule of detection) have the same chirality sense.
  • enantiomeric nantioenriched refer to a sample of a chiral substance whose enantiomeric ratio is greater than 50:50. Enantiomerically enriched compounds may be enantiomerically pure.
  • the term "about,” as used herein, includes the recited number ⁇ 10%. Thus, “about 10” means 9 to 11.
  • the present disclosure encompasses the preparation and use of salts of the Compounds of the Disclosure, including non-toxic pharmaceutically acceptable salts. Examples of pharmaceutically acceptable addition salts include inorganic and organic acid addition salts and basic salts.
  • the pharmaceutically acceptable salts include, but are not limited to, metal salts such as sodium salt, potassium salt, cesium salt and the like; alkaline earth metals such as calcium salt, magnesium salt and the like; organic amine salts such as triethylamine salt, pyridine salt, picoline salt, ethanolamine salt, triethanolamine salt, dicyclohexylamine salt, N,N'-dibenzylethylenediamine salt and the like; inorganic acid salts such as hydrochloride, hydrobromide, phosphate, sulphate and the like; organic acid salts such as citrate, lactate, tartrate, maleate, fumarate, mandelate, acetate, dichloroacetate, trifluoroacetate, oxalate, formate and the like; sulfonates such as methanesulfonate, benzenesulfonate, p-toluenesulfonate and the like; and amino acid salts such as arginate, as
  • pharmaceutically acceptable salt refers to any salt, e.g., obtained by reaction with an acid or a base, of a Compound of the Disclosure that is physiologically tolerated in the target patient (e.g., a mammal, e.g., a human).
  • Acid addition salts can be formed by mixing a solution of the particular Compound of the Disclosure with a solution of a pharmaceutically acceptable non-toxic acid such as hydrochloric acid, fumaric acid, maleic acid, succinic acid, acetic acid, citric acid, tartaric acid, carbonic acid, phosphoric acid, oxalic acid, dichloroacetic acid, or the like.
  • Basic salts can be formed by mixing a solution of the compound of the present disclosure with a solution of a pharmaceutically acceptable non-toxic base such as sodium hydroxide, potassium hydroxide, choline hydroxide, sodium carbonate and the like.
  • the present disclosure encompasses the preparation and use of solvates of Compounds of the Disclosure ally do not significantly alter the physiological activity or toxic nds, and as such may function as pharmacological equivalents.
  • " as used herein is a combination, physical association and/or solvation of a compound of the present disclosure with a solvent molecule such as, e.g. a disolvate, monosolvate or hemisolvate, where the ratio of solvent molecule to compound of the present disclosure is about 2:1, about 1:1 or about 1:2, respectively.
  • This physical association involves varying degrees of ionic and covalent bonding, including hydrogen bonding.
  • solvate can be isolated, such as when one or more solvent molecules are incorporated into the crystal lattice of a crystalline solid.
  • solvate encompasses both solution-phase and isolatable solvates.
  • Compounds of the Disclosure can be present as solvated forms with a pharmaceutically acceptable solvent, such as water, methanol, ethanol, and the like, and it is intended that the disclosure includes both solvated and unsolvated forms of Compounds of the Disclosure.
  • a pharmaceutically acceptable solvent such as water, methanol, ethanol, and the like
  • solvate is a hydrate.
  • a "hydrate” relates to a particular subgroup of solvates where the solvent molecule is water.
  • Solvates typically can function as pharmacological equivalents. Preparation of solvates is known in the art. See, for example, M.
  • a typical, non-limiting, process of preparing a solvate would involve dissolving a Compound of the Disclosure in a desired solvent (organic, water, or a mixture thereof) at temperatures above 20°C to about 25°C, then cooling the solution at a rate sufficient to form crystals, and isolating the crystals by known methods, e.g., filtration.
  • Analytical techniques such as infrared spectroscopy can be used to confirm the presence of the solvent in a crystal of the solvate.
  • Compounds of the Disclosure are inhibitors of SMYD proteins, such as SMYD3 and SMYD2, a number of diseases, conditions, or disorders mediated by SMYD proteins, such as SMYD3 and SMYD2, can be treated by employing these compounds.
  • the present disclosure is thus directed generally to a method for treating a disease, condition, or disorder responsive to the inhibition of SMYD proteins, such as SMYD3 and SMYD2, in an animal suffering from, or at risk of suffering from, the disorder, the method comprising administering to the animal an effective amount of one or more Compounds of the Disclosure.
  • the present disclosure to a method of inhibiting SMYD proteins in an animal in need the omprising administering to the animal a therapeutically effective amount of at least one Compound of the Disclosure.
  • the present disclosure is further directed to a method of inhibiting SMYD3 in an animal in need thereof, the method comprising administering to the animal a therapeutically effective amount of at least one Compound of the Disclosure.
  • the present disclosure is further directed to a method of inhibiting SMYD2 in an animal in need thereof, the method comprising administering to the animal a therapeutically effective amount of at least one Compound of the Disclosure.
  • the terms “treat,” “treating,” “treatment,” and the like refer to eliminating, reducing, or ameliorating a disease or condition, and/or symptoms associated therewith. Although not precluded, treating a disease or condition does not require that the disease, condition, or symptoms associated therewith be completely eliminated.
  • the terms “treat,” “treating,” “treatment,” and the like may include “prophylactic treatment,” which refers to reducing the probability of redeveloping a disease or condition, or of a recurrence of a previously-controlled disease or condition, in a subject who does not have, but is at risk of or is susceptible to, redeveloping a disease or condition or a recurrence of the disease or condition.
  • the term “treat” and synonyms contemplate administering a therapeutically effective amount of a Compound of the Disclosure to an individual in need of such treatment.
  • treatment also includes relapse prophylaxis or phase prophylaxis, as well as the treatment of acute or chronic signs, symptoms and/or malfunctions.
  • the treatment can be orientated symptomatically, for example, to suppress symptoms. It can be effected over a short period, be oriented over a medium term, or can be a long-term treatment, for example within the context of a maintenance therapy.
  • terapéuticaally effective amount refers to an amount of the active ingredient(s) that is(are) sufficient, when administered by a method of the disclosure, to efficaciously deliver the active ingredient(s) for the treatment of condition or disease of interest to an individual in need thereof.
  • the therapeutically effective amount of the agent may reduce (i.e., retard d preferably stop) unwanted cellular proliferation; reduce the number uce the tumor size; inhibit (i.e., retard to some extent and preferably st tration into peripheral organs; inhibit (i.e., retard to some extent and preferably stop) tumor metastasis; inhibit, to some extent, tumor growth; modulate protein methylation in the target cells; and/or relieve, to some extent, one or more of the symptoms associated with the cancer.
  • the administered compound or composition prevents growth and/or kills existing cancer cells, it may be cytostatic and/or cytotoxic.
  • the term "container" means any receptacle and closure therefore suitable for storing, shipping, dispensing, and/or handling a pharmaceutical product.
  • insert means information accompanying a pharmaceutical product that provides a description of how to administer the product, along with the safety and efficacy data required to allow the physician, pharmacist, and patient to make an informed decision regarding use of the product.
  • the package insert generally is regarded as the "label" for a pharmaceutical product.
  • SMYD proteins such as SMYD3 and SMYD2
  • SMYD3 and SMYD2 can be used in treating diseases and conditions such as proliferative diseases, wherein inhibition of SMYD proteins, such as SMYD3 and SMYD2 provides a benefit.
  • the Compounds of the Disclosure can be used to treat a "SMYD protein mediated disorder" (e.g., a SMYD3-mediated disorder or a SMYD2-mediated disorder).
  • a SMYD protein mediated disorder is any pathological condition in which a SMYD protein is know to play a role.
  • a SMYD-mediated disorder is a proliferative disease.
  • inhibiting SMYD proteins is the inhibition of the activity of one or more activities of SMYD proteins such as SMYD3 and SMYD2.
  • the activity of the SMYD proteins such as SMYD3 and SMYD2 is the ability of the SMYD protein such as SMYD3 or SMYD2 to transfer a methyl group to a target protein (e.g., histone). It should be appreciated that the activity of the one or more SMYD proteins such as SMYD3 and SMYD2 may be inhibited in vitro or in vivo.
  • SMYD proteins such as SMYD lude at least 10% inhibition, at least 20% inhibition, at least 30% inh inhibition, at least 50% inhibition, at least 60% inhibition, at least 70% inhibition, at least 80% inhibition, at least 90% inhibition, and up to 100% inhibition.
  • the family of human SMYD proteins consists of SMYD1, SMYD2, SMYD3, SMYD4 and SMYD5.
  • SMYD1, SMYD2, and SMYD3 share a high degree of sequence homology and, with the exception of SMYD5, human SMYD proteins harbor at least one C-terminal tetratrico peptide repeat (TPR) domain.
  • TPR C-terminal tetratrico peptide repeat
  • SMYD proteins have been found to be linked to various cancers (See e.g., Hamamoto et al. Nat Cell. Biol. 2004, 6: 731-740), Hu et al. Canncer Research 2009, 4067-4072, and Komatsu et al. Carcinogenesis 2009, 301139-1146.)
  • SMYD3 is a protein methyltransferase found to be expressed at high levels in a number of different cancers (Hamamoto, R., et al., Nat. Cell Biol., 6(8):731-40 (2004)). SMYD3 likely plays a role in the regulation of gene transcription and signal transduction pathways critical for survival of breast, liver, prostate and lung cancer cell lines (Hamamoto, R., et al., Nat.
  • RNAi-based technologies have shown that ablation of SMYD3 in hepatocellular carcinoma cell lines greatly reduces cell viability and that its pro-survival role is dependent on its catalytic activity (Hamamoto, R., et al., Nat. Cell Biol., 6(8):731-40 (2004); Van Aller, G.S., et al., Epigenetics, 7(4):340-3 (2012)).
  • SMYD3 has also been shown to be a critical mediator of transformation resulting from gain of function mutations in the oncogene, KRAS for both pancreatic and l a in mouse models.
  • SMYD3 function has also been shown to play a role in immunology and development. For instance, de Almeida reported that SMYD3 plays a role in generation of inducible regulatory T cells (iTreg) cells.
  • iTreg inducible regulatory T cells
  • SMYD3-/- mice demonstrated exacerbation of RSV- induced disease related to enhanced proinflammatory responses and worsened pathogenesis within the lung (de Almeida et al. Mucosal Immunol. 2015 Feb 11. doi: 10.1038/mi.2015.4. [Epub ahead of print]).
  • SMYD2 (SET and MYND domain-containing protein 2) was first characterized as protein that is a member of a sub-family of SET domain containing proteins which catalyze the site-specific transfer of methyl groups onto substrate proteins.
  • SMYD2 was initially shown to have methyltransferase activity towards lysine 36 on histone H3 (H3K36) but has subsequently been shown to have both histone and non-histone methyltrasferase activity.
  • SMYD2 has been implicated in the pathogenesis of multiple cancers. It has been shown to be over-expressed, compared to matched normal samples, in tumors of the breast, cervix, colon, kidney, liver, head and neck, skin, pancreas, ovary, esophagus and prostate, as well as hematologic malignancies such as AML, B- and T-ALL, CLL and MCL, suggesting a role for SMYD2 in the biology of these cancers. More specifically, studies using genetic knock-down of SMYD2 have demonstrated anti-proliferative effects in esophageal squamous cell carcinoma (ESCC), bladder carcinoma and cervical carcinoma cell lines.
  • ESCC esophageal squamous cell carcinoma
  • SMYD2 has also been implicated in immunology. For instance, Xu et al. have shown that SMYD2 is a negative regulator of macrophage activation by suppressing Interleukin-6 and TNF-alpha production. (Xu et al., J Biol Chem. 2015 Feb 27;290(9):5414-23).
  • the present disclosure provides a method of treating cancer in a patient comprising administering a therapeutically effective amount of a Compound of the Disclosure. While not being limited to a specific mechanism, in some embodiemtns, Compounds of the Disclorure can treat cancer by inhibiting SMYD proteins, such as SMYD3 and SMYD2.
  • treatable cancers include, but are not limited to, adrenal cancer, acinic cell carcinoma, acoustic neuroma, acral lentigious melanoma, acrospiroma, acute eosinophilic leukemia, acute erythroid leukemia, acute lymphoblastic leukemia, acute megakaryoblastic leukemia, acute monocytic leukemia, acute promyelocytic leukemia, adenocarcinoma, adenoid cystic carcinoma, adenoma, adenomatoid odontogenic tumor, adenosquamous carcinoma, adipose tissue neoplasm, adrenocortical carcinoma, adult T-cell leukemia/lymphoma, aggressive NK-cell leukemia, AIDS-related lymphoma, alveolar rhabdomyosarcoma, alveolar soft part sarcoma, ameloblastic fibroma, anaplastic large cell
  • the cancer is breast, cervix, colon, kidney, liver, head and neck, skin, pancreas, ovary, esophagus, or prostate cancer.
  • the cancer is a hematologic malignancy such as acute myeloid leukemia (AML), B- and T-acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), or mantle cell lymphoma (MCL).
  • AML acute myeloid leukemia
  • ALL B- and T-acute lymphoblastic leukemia
  • CLL chronic lymphocytic leukemia
  • MCL mantle cell lymphoma
  • the cancer is esophageal squamous cell carcinoma (ESCC), bladder carcinoma, or cervical carcinoma.
  • ESCC esophageal squamous cell carcinoma
  • bladder carcinoma esophageal squamous cell carcinoma
  • cervical carcinoma esophageal squamous cell carcinoma
  • the cancer is a leukemia, for example a leukemia selected from acute monocytic leukemia, acute myelogenous leukemia, chronic myelogenous leukemia, chronic lymphocytic leukemia and mixed lineage leukemia (MLL).
  • the cancer is NUT-midline carcinoma.
  • the cancer is multiple myeloma.
  • the cancer is a lung cancer such as small cell lung cancer (SCLC).
  • SCLC small cell lung cancer
  • the cancer is a neuroblastoma.
  • the cancer is Burkitt's lymphoma.
  • the cancer is cervical cancer.
  • the cancer is esophageal cancer.
  • the cancer is ovarian cancer.
  • the cancer is colorectal cancer.
  • the cancer is prostate cancer.
  • the cancer is breast cancer.
  • the present disclosure provides a therapeutic method of modulating protein methylation, gene expression, cell proliferation, cell differentiation and/or apoptosis in vivo in the cancers mentioned above by administering a therapeutically effective amount of a Compound of the Disclosure to a subject in need of such therapy.
  • Compounds of the Disclosure can be administered to a mammal in the form of a raw chemical without any other components present.
  • Compounds of the Disclosure can also be administered to a mammal as part of a pharmaceutical composition containing the compound combined with a suitable pharmaceutically acceptable carrier.
  • a suitable pharmaceutically acceptable carrier can be selected from pharmaceu excipients and auxiliaries.
  • pharmaceutically acceptable armaceutically acceptable vehicle encompasses any of the stand al carriers, solvents, surfactants, or vehicles. Suitable pharmaceutically acceptable vehicles include aqueous vehicles and nonaqueous vehicles. Standard pharmaceutical carriers and their formulations are described in Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA, 19th ed. 1995.
  • compositions within the scope of the present disclosure include all compositions where a Compound of the Disclosure is combined with one or more pharmaceutically acceptable carriers.
  • the Compound of the Disclosure is present in the composition in an amount that is effective to achieve its intended therapeutic purpose. While individual needs may vary, a determination of optimal ranges of effective amounts of each compound is within the skill of the art.
  • a Compound of the Disclosure can be administered to a mammal, e.g., a human, orally at a dose of from about 0.0025 to about 1500 mg per kg body weight of the mammal, or an equivalent amount of a pharmaceutically acceptable salt or solvate thereof, per day to treat the particular disorder.
  • a useful oral dose of a Compound of the Disclosure administered to a mammal is from about 0.0025 to about 50 mg per kg body weight of the mammal, or an equivalent amount of the pharmaceutically acceptable salt or solvate thereof.
  • the dose is typically about one-half of the oral dose.
  • a unit oral dose may comprise from about 0.01 mg to about 1 g of the Compound of the Disclosure, e.g., about 0.01 mg to about 500 mg, about 0.01 mg to about 250 mg, about 0.01 mg to about 100 mg, 0.01 mg to about 50 mg, e.g., about 0.1 mg to about 10 mg, of the compound.
  • the unit dose can be administered one or more times daily, e.g., as one or more tablets or capsules, each containing from about 0.01 mg to about 1 g of the compound, or an equivalent amount of a pharmaceutically acceptable salt or solvate thereof.
  • a pharmaceutical composition of the present disclosure can be administered to any patient that may experience the beneficial effects of a Compound of the Disclosure.
  • mammals e.g., humans and companion animals, although the disclosure is not intended to be so limited.
  • the patient is a human.
  • a pharmaceutical compo nt disclosure can be administered by any means that achieves its inten xample, administration can be by the oral, parenteral, subcutaneous, intravenous, intramuscular, intraperitoneal, transdermal, intranasal, transmucosal, rectal, intravaginal or buccal route, or by inhalation.
  • the dosage administered and route of administration will vary, depending upon the circumstances of the particular subject, and taking into account such factors as age, gender, health, and weight of the recipient, condition or disorder to be treated, kind of concurrent treatment, if any, frequency of treatment, and the nature of the effect desired.
  • a pharmaceutical composition of the present disclosure can be administered orally.
  • a pharmaceutical composition of the present disclosure can be administered orally and is formulated into tablets, dragees, capsules, or an oral liquid preparation.
  • the oral formulation comprises extruded multiparticulates comprising the Compound of the Disclosure.
  • a pharmaceutical composition of the present disclosure can be administered rectally, and is formulated in suppositories.
  • composition of the present disclosure can be administered by injection.
  • composition of the present disclosure can be administered transdermally.
  • composition of the present disclosure can be administered by inhalation or by intranasal or transmucosal administration.
  • composition of the present disclosure can be administered by the intravaginal route.
  • a pharmaceutical composition of the present disclosure can contain from about 0.01 to 99 percent by weight, e.g., from about 0.25 to 75 percent by weight, of a Compound of the Disclosure, e.g., about 1%, about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75% by weight of a Compound of the Disclosure.
  • a Compound of the Disclosure e.g., about 1%, about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75% by weight of a Compound of the Disclosure.
  • a pharmaceutical composition of the present disclosure is manufactured in a manner which itself will be known in view of the instant disclosure, for example, by means of conventional mixing ee-making, dissolving, extrusion, or lyophilizing processes.
  • p positions for oral use can be obtained by combining the active com excipients, optionally grinding the resulting mixture and processing the mixture of granules, after adding suitable auxiliaries, if desired or necessary, to obtain tablets or dragee cores.
  • Suitable excipients include fillers such as saccharides (for example, lactose, sucrose, mannitol or sorbitol), cellulose preparations, calcium phosphates (for example, tricalcium phosphate or calcium hydrogen phosphate), as well as binders such as starch paste (using, for example, maize starch, wheat starch, rice starch, or potato starch), gelatin, tragacanth, methyl cellulose, hydroxypropylmethylcellulose, sodium carboxymethylcellulose, and/or polyvinyl pyrrolidone.
  • saccharides for example, lactose, sucrose, mannitol or sorbitol
  • cellulose preparations for example, calcium phosphates (for example, tricalcium phosphate or calcium hydrogen phosphate)
  • binders such as starch paste (using, for example, maize starch, wheat starch, rice starch, or potato starch), gelatin, tragacanth, methyl cellulose, hydroxypropylmethyl
  • one or more disintegrating agents can be added, such as the above-mentioned starches and also carboxymethyl-starch, cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof, such as sodium alginate.
  • Auxiliaries are typically flow-regulating agents and lubricants such as, for example, silica, talc, stearic acid or salts thereof (e.g., magnesium stearate or calcium stearate), and polyethylene glycol.
  • Dragee cores are provided with suitable coatings that are resistant to gastric juices.
  • concentrated saccharide solutions can be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, polyethylene glycol and/or titanium dioxide, lacquer solutions and suitable organic solvents or solvent mixtures.
  • cellulose preparations such as acetylcellulose phthalate or hydroxypropylmethyl-cellulose phthalate
  • Dye stuffs or pigments can be added to the tablets or dragee coatings, for example, for identification or in order to characterize combinations of active compound doses.
  • Examples of other pharmaceutical preparations that can be used orally include push-fit capsules made of gelatin, or soft, sealed capsules made of gelatin and a plasticizer such as glycerol or sorbitol.
  • the push-fit capsules can contain a compound in the form of granules, which can be mixed with fillers such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers, or in the form of extruded multiparticulates.
  • the active compounds are preferably dissolved or suspended in suitable liquids, such as fatty oils or liquid paraffin.
  • stabilizers can be added
  • Possible pharmaceutica rectal administration include, for example, suppositories, which co ion of one or more active compounds with a suppository base.
  • Sui bases include natural and synthetic triglycerides, and paraffin hydrocarbons, among others.
  • gelatin rectal capsules consisting of a combination of active compound with a base material such as, for example, a liquid triglyceride, polyethylene glycol, or paraffin hydrocarbon.
  • Suitable formulations for parenteral administration include aqueous solutions of the active compound in a water-soluble form such as, for example, a water-soluble salt, alkaline solution, or acidic solution.
  • a suspension of the active compound can be prepared as an oily suspension.
  • Suitable lipophilic solvents or vehicles for such as suspension may include fatty oils (for example, sesame oil), synthetic fatty acid esters (for example, ethyl oleate), triglycerides, or a polyethylene glycol such as polyethylene glycol-400 (PEG-400).
  • An aqueous suspension may contain one or more substances to increase the viscosity of the suspension, including, for example, sodium carboxymethyl cellulose, sorbitol, and/or dextran.
  • the suspension may optionally contain stabilizers.
  • kits which comprise a Compound of the Disclosure (or a composition comprising a Compound of the Disclosure) packaged in a manner that facilitates their use to practice methods of the present disclosure.
  • the kit includes a Compound of the Disclosure (or a composition comprising a Compound of the Disclosure) packaged in a container, such as a sealed bottle or vessel, with a label affixed to the container or included in the kit that describes use of the compound or composition to practice the method of the disclosure.
  • the compound or composition is packaged in a unit dosage form.
  • the kit further can include a device suitable for administering the composition according to the intended route of administration.
  • a suitable sulfonyl chloride Z-SO 2 Cl
  • a suitable acide chloride Z-COCl
  • a suitable base such as TEA or DIPEA
  • a suitable solvent such as dichloromethane, acetonitrile, or DMF
  • Z-CO 2 H a suitable coup
  • a suitable coupling reagent such as HATU and a suitable base such as TEA or DIPEA
  • a suitable solvent such as dichloromethane, acetonitrile, or DMF.
  • reactions were heated using conventional hotplate apparatus or heating mantle or microwave irradiation equipment. Reactions were conducted with or without stirring, under atmospheric or elevated pressure in either open or closed vessels. Reaction progress was monitored using conventional techniques such as TLC, HPLC, UPLC, or LCMS using instrumentation and methods described below. Reactions were quenched and crude compounds isolated using conventional methods as described in the specific examples provided. Solvent removal was carried out with or without heating, under atmospheric or reduced pressure, using either a rotary or centrifugal evaporator.
  • SAM S-adenosylmethionine
  • SAH S-adenosylhomocysteine
  • Tris Tween20, dimethylsulfoxide (DMSO), bovine skin gelatin (BSG), and Tris(2- carboxyethyl)phosphine hydrochloride solution (TCEP)
  • SAM S-adenosylmethionine
  • SAH S-adenosylhomocysteine
  • Tris Tween20
  • BSG bovine skin gelatin
  • TCEP Tris(2- carboxyethyl)phosphine hydrochloride solution
  • 3 H-SAM was purchase from American Radiolabeled Chemicals with a specific activity of 80 Ci/mmol.
  • 384-well opaque white OptiPlates and SPA beads (Perkin Elmer, catalog # RPNQ0013) were purchased from PerkinElmer. Substrates
  • N-terminally GST-tagged MEKK2 (MAP3K2) protein corresponding to reference sequence AAF63496.3 was purchased from Life Technologies (catalog # PV4010). This protein was expressed in High Five insect cells and purified to >85 % purity. Protein identity was confirmed by MS/MS analysis after proteolytic digestion. The protein sequence used was:
  • Full-length human SMYD3 isoform 1 (BAB86333) was inserted into a modified pET21b plasmid containing a His6 tag and TEV and SUMO cleavage sites. Because two common variants of SMYD3 exist in the population, site directed mutagenesis was subsequently performed to change amino acid 13 from an asparagine to a lysine, resulting in plasmid pEPZ533. A lysine at position 13 conforms to the more commonly occurring sequence (NP_001161212). Protein Expression
  • E. coli (BL21 codonplus RIL strain, Stratagene) were transformed with plasmid pEPZ553 by mixing competent cells and plasmid DNA and incubating on ice for 30 minutes followed by heat shock at 42°C for 1 minute and cooling on ice for 2 minutes. Transformed cells were grown and selected on LB agar with 100 ⁇ g/mL ampicillin and 17 ⁇ g/mL chloramphenicol at 37°C overnight.
  • a single clone was used to inoculate 200 mL of LB medium with 100 ⁇ g/mL ampicillin and 17 ⁇ g/mL chloramphenicol and incubated at 37°C on an orbital h k t 180 Once in log growth, the culture was diluted 1:100 into 2 L of LB m ntil OD 600 was about 0.3 after which the culture was incubated at 15 Once OD 600 reached about 0.4, IPTG was added to a final concentratio e cells were grown overnight at 15°C and 160 rpm. Cells were harvested by centrifugation at 8000 rpm, for 4 minutes at 4°C and stored at -80 o C for purification.
  • SMYD3 was eluted in Buffer A and further purified using an S200 size-exclusion column equilibrated with Buffer A. SMYD3 was concentrated to 2 mg/mL with a final purity of 89%.
  • the assays were all perfo onsisting of 25 mM Tris-Cl pH 8.0, 1 mM TCEP, 0.005% BSG, and 0.005% Tween 20, prepared on the day of use.
  • Compounds in 100% DMSO (1ul) were spotted into a 384-well white opaque OptiPlate using a Bravo automated liquid handling platform outfitted with a 384-channel head (Agilent Technologies).
  • DMSO (1ul) was added to Columns 11, 12, 23, 24, rows A-H for the maximum signal control and 1ul of SAH, a known product and inhibitor of SMYD3, was added to columns 11, 12, 23, 24, rows I-P for the minimum signal control.
  • the assays were stopped by the addition of non-radiolabeled SAM (10ul) to a final concentration of 100 uM, which dilutes the 3 H-SAM to a level where its incorporation into MEKK2 is no longer detectable.
  • Radiolabeled MEKK2 was detected using a scintillation proximity assay (SPA). 10 uL of a 10 mg/mL solution of SPA beads in 0.5 M citric acid was added and the plates centrifuged at 600 rpm for 1 min to precipitate the radiolabeled MEKK2 onto the SPA beads. The plates were then read in a PerkinElmer TopCount plate reader to measure the quantity of 3 H-labeled MEKK2 as disintegrations per minute (dpm) or alternatively, referred to as counts per minute (cpm). % inhibition calculation
  • top and bottom ar wed to float may be fixed at 100 or 0 respectively in a 3-paramet fficient normally allowed to float but may also be fixed at 1 in a 3-par % inhibition and X is the compound concentration.
  • 293T/17 adherent cells were purchased from ATCC (American Type Culture Collection), Manassas, VA, USA.
  • MEM/Glutamax medium, Optimem Reduced Serum medium, penicillin-streptomycin, 0.05% trypsin and 1x D-PBS were purchased from Life Technologies, Grand Island, NY, USA.
  • PBS-10X was purchased from Ambion, Life Technologies, Grand Island, New York, USA.
  • PBS with Tween 20 (PBST (10x)) was purchased from KPL, Gaithersburg, Maryland,USA.
  • Tet System FBS- approved FBS US Source was purchased from Clontech, Mountain View, California, USA.
  • Licor Biosciences Lincoln, NE, USA.
  • Tri-methyl-Lysine [A260]-MEKK2 antibody, MEKK2 and SMYD3 plasmids were made at Epizyme.
  • Anti-flag monoclonal mouse antibody was purchased from Sigma, St. Louis, MO, USA.
  • Methanol was purchased from VWR, Franklin, MA, USA.
  • 10% Tween 20 was purchased from KPL, Inc., Gaithersburg, Maryland, USA. Fugene was purchased from Promega, Madison, WI, USA.
  • the Biotek ELx405 was purchased from BioTek, Winooski, Vermont, USA.
  • the multidrop combi was purchased from Thermo Scientific, Waltham, Massachusetts, USA.
  • Plasmids were prepared for delivery to cells by first mixing 1350 ⁇ L Opti-MEM with Fugene (81 ⁇ L) in a sterile Eppendorf and incubated for five minutes at room temperature (RT).
  • MEKK2-flag (13.6 ug/T150)
  • SMYD3 p3XFlag-CMV-14 without C-3XFlag plasmids were aliquotted to a 1.7 mL sterile microfuge tube.
  • the gene ID for MEKK2 and SMYD3 is NM_006609.3 and Q9H7B4, respectively.
  • Entire volume of Opti-MEM/Fugene mixture was then added to a microfuge tube containing DNA plasmid, mixed and then incubated x 15 minutes at RT.
  • the medium on the 293T/17 cells was refreshed, and the DNA/Fugene complex is added aseptically to each flask, rocked gently, and incubated at 37 C for 5 hours. Medium was then removed, and cells were washed once with PBS in the flask. Trypsin 0.05% (3mL) was added and cells incubated for three minutes. Room temperature MEM+10% Tet system FBS was added and cells were mixed gently, and counted using the Vi-cell. Cells were seeded at 100,000 cells/mL in 50 ⁇ L MEM/10%Tet FBS/Pen/Strep to a 384 well black/clear poly-D-lysine coated plate containing test agent diluted in DMSO.
  • test compound was 40 ⁇ M.
  • the total concentration of DMSO did not exceed 0.2% (v/v).
  • Plates were incubated x 30 minutes at RT in low-airflow area, followed by incubation at 37 °C under 5% CO 2 for 24 hours. Medium was aspirated from all wells of assay plates prior to fixation and permeabilization with ice cold (-20 oC) methanol (90 ⁇ L/well) for ten minutes. Plates were rinsed with PBS three times on BioTek ELx405. PBS was removed with a final aspiration, and Odyssey blocking buffer (50 ⁇ L/well) was added to each well and incubated for one hour at RT.
  • Detection antibody solution (IRDye 800 CW goat anti-rabbit IgG diluted 1:400 in diluent (Odyssey Blocking buffer + 0.1% Tween 20), plus IRDye 680CW goat anti-mouse IgG at 1:500 in diluent (Odyssey Blocking buffer + 0.1% Tween 20) was added (20 ⁇ L/well) and incubated in dark for one hour at RT. Plat ed four times with PBS-T (1X) on ELx405. A final rinse with wa (115 ⁇ L/well x three washes on the ELx405). Plates were then ce own, on paper towel, at 200 x g to remove excess reagent.
  • Each plate included fourteen control wells of DMSO only treatment (Minimum Inhibition) as well as fourteen control wells for maximum inhibition (Background). The average of the ratio values for each control type was calculated and used to determine the percent inhibition for each test well in the plate. Reference compound was serially diluted two-fold in DMSO for a total of nine test concentrations, beginning at 40 ⁇ M. Percent inhibition was calculated below . P
  • SAM S-adenosylmethionine
  • SAH S-adenosylhomocysteine
  • bicine Tween20
  • dimethylsulfoxide DMSO
  • bovine skin gelatin BSG
  • Tris(2- carboxyethyl)phosphine hydrochloride TCEP
  • Peptide was synthesized with a N-terminal linker-affinity tag motif and a C- terminal amide cap by 21 st Century Biochemicals.
  • the peptide was high high-perfomance liquid chromatography (HPLC) purified to greater than 95% purity and confirmed by liquid chromatography mass spectrometry (LC-MS).
  • HPLC high-perfomance liquid chromatography
  • LC-MS liquid chromatography mass spectrometry
  • SMYD2 (NP_064582.2) was cloned into a pFastbac-Htb-lic vector with an N-terminal His6 tag and FLAG tag, preceded by a TEV protease cleavage site.
  • the protein was expressed in Sf9 insect cells. Cells were resuspended in lysis buffer (25 mM HEPES-NaOH, pH 7.5, 200 mM NaCl, 5% glycerol, and 5 mM ⁇ -ME) and lysed by sonication.
  • lysis buffer 25 mM HEPES-NaOH, pH 7.5, 200 mM NaCl, 5% glycerol, and 5 mM ⁇ -ME
  • the protein was purified by Ni-NTA (Qiagen), followed by TEV cleavage to remove the His6 tag, subtractive Ni-NTA (Qiagen), and gel filtration chromatography using an S200 column (GE Healthcare). Purified protein was stored in 20 mM Tris-HCl, pH 8.0, 100 mM NaCl, and 1 mM TCEP.
  • Compounds in 100% DMSO (1ul) were spotted into a polypropylene 384-well V- bottom plates (Greiner) using a Platemate Plus outfitted with a 384-channel head (Thermo Scientific).
  • DMSO (1ul) was added to Columns 11, 12, 23, 24, rows A-H for the maximum signal control and 1ul of SAH, a known product and inhibitor of SMYD2, was added to columns 11, 12, 23, 24, rows I-P for the minimum signal control.
  • the assays were stopped by the addition of no (10ul) to a final concentration of 600uM, which dilutes the 3 H-S ere its incorporation into the peptide substrate is no longer detectable n in the 384-well polypropylene plate was then transferred to a 384-well Flashplate and the biotinylated peptides were allowed to bind to the streptavidin surface for at least 1 hour before being washed three times with 0.1%Tween20 in a Biotek ELx405 plate washer.

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Abstract

The present disclosure provides pyrrolidine carboxamide compounds having Formula I and the pharmaceutically acceptable salts and solvates thereof, wherein A, B, X, Y, and Z are defined as set forth in the specification. The present disclosure is also directed to the use of compounds of Formula I to treat a disorder responsive to the blockade of SMYD proteins such as SMYD3 or SMYD2. Compounds of the present disclosure are especially useful for treating cancer.

Description

SUBSTITUTED PYRROLIDINE CARBOXAMIDE COMPOUNDS BACKGROUND OF THE INVENTION Field of the Invention
[0001] The present disclosure provides substituted pyrrolidine carboxamides as SMYD protein inhibitors, such as SMYD3 and SMYD2 inhibitors, and therapeutic methods of treating conditions and diseases wherein inhibition of SMYD proteins such as SMYD3 and SMYD2 provides a benefit. Background
[0002] Epigenetic regulation of gene expression is an important biological determinant of protein production and cellular differentiation and plays a significant pathogenic role in a number of human diseases. Epigenetic regulation involves heritable modification of genetic material without changing its nucleotide sequence. Typically, epigenetic regulation is mediated by selective and reversible modification (e.g., methylation) of DNA and proteins (e.g., histones) that control the conformational transition between transcriptionally active and inactive states of chromatin. These covalent modifications can be controlled by enzymes such as methyltransferases (e.g., SMYD proteins such as SMYD3 and SMYD2), many of which are associated with genetic alterations that can cause human disease, such as proliferative disorders. Thus, there is a need for the development of small molecules that are capable of inhibiting the activity of SMYD proteins such as SMYD3 and SMYD2. BRIEF SUMMARY OF THE INVENTION [0003] In one aspect, the provides substituted pyrrolidine carboxamide compounds repre I below, and the pharmaceutically acceptable salts and solvates th eferred to herein as "Compounds of the Disclosure."
[0004] In another aspect, the present disclosure provides a Compound of the Disclosure and one or more pharmaceutically acceptable carriers.
[0005] In another aspect, the present disclosure provides a method of inhibiting SMYD proteins, such as SMYD3 or SMYD2, or both, in a mammal, comprising administering to the mammal an effective amount of at least one Compound of the Disclosure.
[0006] In another aspect, the present disclosure provides a method of irreversibly inhibiting SMYD proteins, such as SMYD3 or SMYD2, or both, in a mammal, comprising administering to the mammal an effective amount of at least one Compound of the Disclosure.
[0007] In another aspect, the present disclosure provides methods for treating a disease, disorder, or condition, e.g., cancer, responsive to inhibition of SMYD proteins, such as SMYD3 or SMYD2, or both, comprising administering a therapeutically effective amount of a Compound of the Disclosure.
[0008] In another aspect, the present disclosure provides the use of Compounds of the Disclosure as inhibitors of SMYD3.
[0009] In another aspect, the present disclosure provides the use of Compounds of the Disclosure as inhibitors of SMYD2.
[0010] In another aspect, the present disclosure provides the use of Compounds of the Dicslosure as inhibitors of SMYD proteins.
[0011] In another aspect, the present disclosure provides a pharmaceutical composition for treating a disease, disorder, or condition responsive to inhibition of SMYD proteins, such as SMYD3 or SMYD2, or both, wherein the pharmaceutical composition comprises a therapeutically effective amount of a Compound of the Disclosure in a mixture with one or more pharmaceutically acceptable carriers.
[0012] In another aspect, the present disclosure provides Compounds of the Disclosure for use in treating cancer in a mammal, e.g., breast, cervical, colon, kidney, liver, head and neck, skin, pancreatic, ovary, esophageal, lung, and prostate cancer.
[0013] In another aspect, the present disclosure provides a Compound of the Disclosure for use in the ma dicament for treating cancer in a mammal.
[0014] In another aspect, the p ovides kit comprising a Compound of the Disclosure.
[0015] Additional embodiments and advantages of the disclosure will be set forth, in part, in the description that follows, and will flow from the description, or can be learned by practice of the disclosure. The embodiments and advantages of the disclosure will be realized and attained by means of the elements and combinations particularly pointed out in the appended claims. [0016] It is to be understood that both the foregoing summary and the following detailed description are exemplary and explanatory only, and are not restrictive of the invention as claimed. DETAILED DESCRIPTION OF THE INVENTION [0017] One aspect of the present disclosure is based on the use of Compounds of the Disclosure as inhibitors of SMYD proteins. In view of this property, the Compounds of the Disclosure are useful for treating diseases, disorders, or conditions, e.g., cancer, responsive to inhibition of SMYD proteins.
[0018] One aspect of the present disclosure is based on the use of Compounds of the Disclosure as inhibitors of SMYD3. In view of this property, the Compounds of the Disclosure are useful for treating diseases, disorders, or conditions, e.g., cancer, responsive to inhibition of SMYD3.
[0019] One aspect of the present disclosure is based on the use of Compounds of the Disclosure as inhibitors of SMYD2. In view of this property, the Compounds of the Disclosure are useful for treating diseases, disorders, or conditions, e.g., cancer, responsive to inhibition of SMYD2.
[0020] In one embodiment, Compounds of the Disclosure are compounds having Formula I:
Figure imgf000004_0001
and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein:
[0021] A is selected from the group consisting of 1,2,3-triazolyl, 1,2,4-triazolyl, 1,3,4- oxadiazole, 1,3,4-thiadiazole, 1-imidazolyl, 1-isoquinolinyl, 1-pyrazolyl, 2-(1,2,3,4- tetrahydroquinolinyl), 2-benzo[ nzo[d]thiazolyl, 2-chromenyl-4-one, 2-furanyl, 2-imidazo[1,2-b]pyr olyl, 2-indolyl, 2-naphthalenyl, 2- pyrazinyl, 2-pyridyl, 2-pyrim nyl, 2-pyrrolyl, 2-quinolinyl, 2- quinoxalinyl, 2-thiazolo[5,4-c azolyl, 2-thiophenyl, 3-(1,2,3,4- tetrahydroisoquinoline), 3-(1,2,4-oxadiazolyl), 3-imidazo[1,2-a]pyrimidinyl, 3-indazolyl, 3-indolyl, 3-isothiazolyl, 3-pyrazolyl, 3-pyridazinyl, 3-pyridinyl-2-one, 3-pyridyl, 3-pyrrolo[3,2-b]pyridinyl, 3-quinolinyl, 4-(2,2- difluorobenzo[d][1,3]dioxolyl), 4-cyclohexanyl-1-amine, 4-imidazolyl, 4-indolinyl-2- one, 4-indolyl, 4-isothiazolyl, 4-oxazolyl, 4-piperidinyl, 4-pyrazolyl, 4-pyridyl, 4- quinolinyl, 5-(1,3-dihydro-2H-benzo[d]imidazolyl-2-one), 5-(1,3-dihydro-2H- pyrrolo[2,3-b]pyridinyl-2-one), 5-(1,3-dihydro-2H-pyrrolo[2,3-c]pyridinyl-2-one), 5-(2,2-difluorobenzo[d][1,3]dioxolyl), 5-(2,4-dihydro-3H-1,2,4-triazolyl-3-one), 5-4H- furo[3,2-b]pyrrolyl, 5-benzo[c][1,2,5]oxadiazolyl, 5-benzo[d][1,3]dioxolyl, 5-benzo[d]oxazolyl-2(3H)-one, 5-bicyclo[2.2.1]heptyl-2-ene, 5-indolinyl-2,3-dione, 5-indolinyl-2-one, 5-indolyl, 5-isoindolinyl-1-one, 5-isoxazolyl, 5-pyrazolo[3,4- c]pyridinyl, 5-pyrazolyl, 5-pyrimidinyl, 5-thiazolyl, 6-(1,2,3,4- tetrahydronaphthalenyl), 6-(3,4-dihydroquinolinyl-2(1H)-one), 6-(3,4- dihydroquinoxalinyl-2(1H)-one), 6-(4,5-dihydropyridazinyl-3(2H)-one), 6-benzo[b][1,4]oxazinyl-3-one, 6-benzo[d]imidazolyl, 6-benzo[d]oxazolyl-2(3H)-one, 6-benzo[d]thiazolyl, 6-chromenyl-2-one, 6-imidazo[2,1-b]thiazole, 6-indazolyl, 6-indolinyl-2-one, 6-indolyl, 6-isoquinolinyl, 6-quinolinyl, 6-quinoxalinyl, 6-quinoxalinyl-2(1H)-one, 7-(3,4-dihydroquinolinyl-2(1H)-one), 7-(3,4- dihydroquinoxalin-2(1H)-one), 7-benzo[b][1,4]oxazinyl-3-one, 7-indolinyl-2-one, 7- quinolinyl, 8-benzo[b][1,4]oxazinyl-3-one, cyclopropanyl, phenyl, 4-(prop-1-en-1-yl)- imidazole, 1-butanyl-imidazole, sec-butylcyclopropane, 2-(ethylsulfonyl)propanyl, 1-isobutylpyrrolidine, 4-pyridyl 1-oxide, and 5-benzo[c][1,2,5]oxadiazolyl 1-oxide, each of which is optionally substituted with one, two, or three substituents independently selected from the group consisting of halo, hydroxy, alkoxy, amino, alkylamino, dialkylamino, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, C1-6 alkyl, haloalkyl, hydroxyalkyl, (carboxamido)alkyl, (cycloalkyl)alkyl, optionally substituted C3-12 cycloalkyl, optionally substituted C6-14 aryl, optionally substituted 5- to 14-membered heteroaryl, optionally substituted 4- to 14-membered heterocyclo, and aralkyl;
[0022] Y is -C(R1a)(R1b)-; or Y is absent, i.e., A forms a bond with the carbonyl carbon atom;
[0023] B is:
Figure imgf000005_0001
;
[0024] X is selected from the group consisting of -S(=O)2-, -S(=O)2N(R6)-, -S(=O)2C(R7)(H)-, -C(=O)-, -C(=O)N(R6)-, -C(=O)O-, and -C(=O)C(R7)(H)-; or X is absent, i.e., Z is forms a bond with the nitrogen atom; [0025] Z is selected from the group consisting of hydrogen, optionally substituted C1-6 alkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, (amino)(hydroxy)alkyl, (amino)(aryl)alkyl, (hydroxy)(aryl)alkyl, (aralkylamino)alkyl, alkoxyalkyl, optionally substituted C6-14 aryl, optionally substituted 4- to 14-membered heterocyclo, optionally substituted 5- to 14-membered heteroaryl, optionally substituted C3-12 cycloalkyl, aralkyl, and heteroaralkyl;
[0026] R1s and R1b are independently selected from the group consisting of hydrogen and C1-4 alkyl;
[0027] R2a, R2b, R3a, R3b, R4a, and R4b are each independently selected from the group consisting of hydrogen, amino, alkylamino, dialkylamino, cycloalkylamino, halo, hydroxy, C1-6 alkyl, alkoxy, haloalkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally substituted C3-12 cycloalkyl, optionally substituted C6-14 aryl, optionally substituted 4- to 14-membered heterocyclo, optionally substituted 5- to 14-membered heteroaryl, alkoxyalkyl, aralkyl, alkoxycarbonyl, sulfonamido, carboxamido, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or
[0028] R2a and R2b taken together with the carbon atom to which they are attached form a carbonyl; and R3a, R3b, R4a, and R4b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, aralkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or
[0029] R3a and R3b taken together with the carbon atom to which they are attached form a carbonyl; and R2a, R2b, R4a, and R4b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally sub alkoxyalkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or
[0030] R4a and R4b taken toge n atom to which they are attached form a carbonyl; and R2a, R2b, ch independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, -N(H)C(=O)R8a; and - CH2N(H)C(=O)R8b; or
[0031] R2a and R2b taken together with the carbon atom to which they are attached form a C3-6 cycloalkyl or C3-6 heterocyclo; and R3a, R3b, R4a, and R4b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, aralkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or
[0032] R3a and R3b taken together with the carbon atom to which they are attached form a C3-6 cycloalkyl or C3-6 heterocyclo; and R2a, R2b, R4a, and R4b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or
[0033] R4a and R4b taken together with the carbon atom to which they are attached form a C3-6 cycloalkyl or C3-6 heterocyclo; and R2a, R2b, R3a, and R3b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, - N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b;
[0034] R5 is selected from the group consisting of hydrogen and C1-4 alkyl;
[0035] R6 is selected from the group consisting of hydrogen and C1-4 alkyl;
[0036] R7 is selected from the group consisting of hydrogen, C1-4 alkyl, amino, alkylamino, dialkylamino, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, and hydroxyalkyl;
[0037] R8a is selected from the group consisting of C1-6 alkyl, haloalkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally substituted C6-14 aryl, optionally substituted 4- to 14-membered heterocyclo, optionally substituted 5- to 14-membered heteroaryl, optionally substituted C3-12 cycloalkyl, aralkyl, and heteroaralkyl; and
[0038] R8b is selected from the group consisting of C1-6 alkyl, haloalkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally aryl, optionally substituted C4-14 heterocyclo, optionally subs membered heteroaryl, optionally substituted C3-12 cycloalkyl, ara kyl.
[0039] In another embodiment e Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein A is selected from the group consisting of 1,2,3-triazolyl, 1,2,4-triazolyl, 1-imidazolyl, 1-isoquinolinyl, 1-pyrazolyl, 2-(1,2,3,4- tetrahydroquinolinyl), 2-benzo[d]imidazolyl, 2-benzo[d]thiazolyl, 2-chromenyl-4-one, 2-furanyl, 2-imidazo[1,2-b]pyridazinyl, 2-imidazolyl, 2-indolyl, 2-naphthalenyl, 2- pyrazinyl, 2-pyridyl, 2-pyrimidinyl, 2-pyrrolidinyl, 2-pyrrolyl, 2-quinolinyl, 2- quinoxalinyl, 2-thiazolo[5,4-c]pyridinyl, 2-thiazolyl, 2-thiophenyl, 3-(1,2,3,4- tetrahydroisoquinoline), 3-(1,2,4-oxadiazolyl), 3-imidazo[1,2-a]pyrimidinyl, 3-indazolyl, 3-indolyl, 3-isothiazolyl, 3-pyrazolyl, 3-pyridazinyl, 3-pyridinyl-2-one, 3-pyridyl, 3-pyrrolo[3,2-b]pyridinyl, 3-quinolinyl, 4-(2,2- difluorobenzo[d][1,3]dioxolyl), 4-cyclohexanyl-1-amine, 4-imidazolyl, 4-indolinyl-2- one, 4-indolyl, 4-isothiazolyl, 4-oxazolyl, 4-piperidinyl, 4-pyrazolyl, 4-pyridyl, 4- quinolinyl, 5-(1,3-dihydro-2H-benzo[d]imidazolyl-2-one), 5-(1,3-dihydro-2H- pyrrolo[2,3-b]pyridinyl-2-one), 5-(1,3-dihydro-2H-pyrrolo[2,3-c]pyridinyl-2-one), 5-(2,2-difluorobenzo[d][1,3]dioxolyl), 5-(2,4-dihydro-3H-1,2,4-triazolyl-3-one), 5-4H- furo[3,2-b]pyrrolyl, 5-benzo[c][1,2,5]oxadiazolyl, 5-benzo[d][1,3]dioxolyl, 5-benzo[d]oxazolyl-2(3H)-one, 5-bicyclo[2.2.1]heptyl-2-ene, 5-indolinyl-2,3-dione, 5-indolinyl-2-one, 5-indolyl, 5-isoindolinyl-1-one, 5-isoxazolyl, 5-pyrazolo[3,4- c]pyridinyl, 5-pyrazolyl, 5-pyrimidinyl, 5-thiazolyl, 6-(1,2,3,4- tetrahydronaphthalenyl), 6-(3,4-dihydroquinolinyl-2(1H)-one), 6-(3,4- dihydroquinoxalinyl-2(1H)-one), 6-(4,5-dihydropyridazinyl-3(2H)-one), 6-benzo[b][1,4]oxazinyl-3-one, 6-benzo[d]imidazolyl, 6-benzo[d]oxazolyl-2(3H)-one, 6-benzo[d]thiazolyl, 6-chromenyl-2-one, 6-imidazo[2,1-b]thiazole, 6-indazolyl, 6-indolinyl-2-one, 6-indolyl, 6-isoquinolinyl, 6-quinolinyl, 6-quinoxalinyl, 6-quinoxalinyl-2(1H)-one, 7-(3,4-dihydroquinolinyl-2(1H)-one), 7-(3,4- dihydroquinoxalin-2(1H)-one), 7-benzo[b][1,4]oxazinyl-3-one, 7-indolinyl-2-one, 7- quinolinyl, 8-benzo[b][1,4]oxazinyl-3-one, cyclopropanyl, phenyl, 4-(prop-1-en-1-yl)- imidazole, 1-butanyl-imidazole, sec-butylcyclopropane, 2-(ethylsulfonyl)propanyl, 1-isobutylpyrrolidine, 4-pyridyl 1-oxide, and 5-benzo[c][1,2,5]oxadiazolyl 1-oxide, each of which is optionally substituted with one, two, or three substituents independently selected from th of halo, hydroxy, alkoxy, amino, alkylamino, dialkylamino, (am no)alkyl, (dialkylamino)alkyl, C1-6 alkyl, haloalkyl, hydroxyalky lkyl, (cycloalkyl)alkyl, optionally substituted C3-12 cycloalkyl, op C6-14 aryl, optionally substituted 5- to 14-membered heteroaryl, optionally substituted 4- to 14-membered heterocyclo, and aralkyl.
[0040] In another embodiment, Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein B is selected from the group consisting of:
Figure imgf000009_0001
. [0041] In another embodiment, Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein B is selected from the group consisting of:
Figure imgf000009_0002
Figure imgf000009_0003
[0042] R4a is selected from the group consisting of hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, and aralkyl. In another embodiment, R4a is C1-4 alkyl. In another embodiment, R4a is methyl.
[0043] In another embodiment, Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein X is selected from the group consisting of -S(=O)2- and -C(=O)-; or X is absent. In another embodiment, X is -S(=O)2-. In another embodiment, X is - C(=O)-. In another embodiment, X is absent.
[0044] In another embodiment, Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein Z is selected from the group consisting of optionally substituted C1-6 alkyl, hydroxyalkyl, (a lamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (hetero ally substituted C6-14 aryl, optionally substituted 4- to 14-membered nally substituted 5- to 14-membered heteroaryl, optionally substituted C3-12 cycloalkyl, aralkyl, and heteroaralkyl. In another embodiment, Z is selected from the group consisting of C1-6 alkyl and optionally substituted C3-12 cycloalkyl. In another embodiment, Z is cyclopropyl. [0045] In another embodiment, Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein Y is -CH2-.
[0046] In another embodiment, Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein Y is absent.
[0047] In another embodiment, Compounds of the Disclosure are compounds having Formula I, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, wherein A is selected from the group consisting of 5-indolinyl-2-one, 6-benzo[b][1,4]oxazinyl-3-one, 5-pyrazolyl, 3-isothiazolyl, 6-benzo[d]oxazolyl-2(3H)- one, 1,2,3-triazolyl, phenyl, and 5-(1,3-dihydro-2H-benzo[d]imidazolyl-2-one). In another embodiment, A is 5-indolinyl-2-one.
[0048] In another embodiment, Compounds of the Disclosure are compounds of Tables 1, 3, and 4, and the pharmaceutically acceptable salts or solvates, e.g., hydrates, thereof, or different pharmaceutically acceptable salt thereof.
[0049] It should be appreciated that the Compounds of the Disclosure in certain embodiments are the free base, various salts, and hydrate forms, and are not limited to the particular salt listed in Tables 1, 3, and 4.
Figure imgf000011_0001
Figure imgf000012_0001
Figure imgf000013_0001
Figure imgf000014_0001
Figure imgf000015_0001
Figure imgf000016_0001
Figure imgf000017_0001
Figure imgf000018_0001
Figure imgf000019_0001
Figure imgf000020_0001
Figure imgf000021_0001
Figure imgf000022_0001
Figure imgf000023_0001
Figure imgf000024_0001
Figure imgf000025_0001
Figure imgf000026_0001
Figure imgf000027_0001
Figure imgf000028_0001
Figure imgf000029_0001
Figure imgf000030_0001
Figure imgf000031_0001
Figure imgf000032_0001
Figure imgf000033_0001
Definitions
[0050] For the purpose of the present discosure, the terms used in connection with A have the chemical structures set forth in Table 2, each of which may be optionally substituted with one or more substituents, e.g., 1, 2, 3, 4, or 5 substituents, depending on the nature of the group and the number of available positions. For example, when A is 2-furanyl there are three carbon atoms for available for substitution. When A is 2-naphthalenyl there are seven carbon atoms available for substitution. Substitution may occur at any available carbon or nitrogen atom. Optional substituents include, but are not limited to, halo, hydroxy, alkoxy, amino, alkylamino, dialkylamino, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, C1-6 alkyl, haloalkyl, hydroxyalkyl, (carboxamido)alkyl, (cycloalkyl)alkyl, optionally substituted C3-12 cycloalkyl, optionally substituted C6-14 aryl, optionally substituted 5- to 14-membered heteroaryl, optionally substituted 4- to 14-membered heterocyclo, or aralkyl.
Figure imgf000034_0001
Figure imgf000035_0001
Figure imgf000036_0001
Figure imgf000037_0001
Figure imgf000038_0001
Figure imgf000039_0001
Figure imgf000040_0001
Figure imgf000041_0001
[0051] For the purpose of the present disclosure, the term "alkyl" as used by itself or as part of another group refers to a straight- or branched-chain aliphatic hydrocarbon containing one to twelve carbon atoms (i.e., C1-12 alkyl) or the number of carbon atoms designated (i.e., a C1 alkyl such as methyl, a C2 alkyl such as ethyl, a C3 alkyl such as propyl or isopropyl, etc.). In one embodiment, the alkyl group is chosen from a straight chain C1-10 alkyl group. In another embodiment, the alkyl group is chosen from a branched chain C3-10 alkyl group. In another embodiment, the alkyl group is chosen from a straight chain C1-6 alkyl group iment, the alkyl group is chosen from a branched chain C3-6 alkyl gro bodiment, the alkyl group is chosen from a straight chain C1-4 alkyl mbodiment, the alkyl group is chosen from a branched chain C3-4 alky g p her embodiment, the alkyl group is chosen from a straight or branched chain C3-4 alkyl group. In another embodiment, the alkyl group is partially or completely deuterated, i.e., one or more hydrogen atoms of the alkyl group are replaced with deuterium atoms. Non-limiting exemplary C1-10 alkyl groups include methyl (including -CD3), ethyl, propyl, isopropyl, butyl, sec-butyl, tert-butyl, iso-butyl, 3-pentyl, hexyl, heptyl, octyl, nonyl, and decyl. Non-limiting exemplary C1-4 alkyl groups include methyl, ethyl, propyl, isopropyl, butyl, sec-butyl, tert-butyl, and iso-butyl.
[0052] For the purpose of the present disclosure, the term "optionally substituted alkyl" as used by itself or as part of another group means that the alkyl as defined above is either unsubstituted or substituted with one, two, or three substituents independently chosen from nitro, haloalkoxy, aryloxy, aralkyloxy, alkylthio, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, alkoxycarbonyl, and carboxyalkyl. In one embodiment, the alkyl is a C1-4 alkyl. In one embodiment, the optionally substituted alkyl is substituted with two substituents. In another embodiment, the optionally substituted alkyl is substituted with one substituent. Non-limiting exemplary optionally substituted alkyl groups include -CH2CH2NO2, -CH2CH2CO2H, -CH2CH2SO2CH3, -CH2CH2COPh, and -CH2C6H11.
[0053] For the purpose of the present disclosure, the term "cycloalkyl" as used by itself or as part of another group refers to saturated and partially unsaturated (containing one or two double bonds) cyclic aliphatic hydrocarbons containing one to three rings having from three to twelve carbon atoms (i.e., C3-12 cycloalkyl) or the number of carbons designated. In one embodiment, the cycloalkyl group has two rings. In one embodiment, the cycloalkyl group has one ring. In another embodiment, the cycloalkyl group is chosen from a C3-8 cycloalkyl group. In another embodiment, the cycloalkyl group is chosen from a C3-6 cycloalkyl group. Non-limiting exemplary cycloalkyl groups include cyclopropyl, cyclobutyl, cyclopentyl, cyclohexyl, cycloheptyl, cyclooctyl, norbornyl, decalin, adamantyl, cyclohexenyl, and spiro[3.3]heptane.
[0054] For the purpose of the present disclosure, the term "optionally substituted cycloalkyl" as used by itself or as part of another group means that the cycloalkyl as defined above is either unsubsti d with one, two, or three substituents independently chosen from no, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydrox oalkoxy, aryloxy, aralkyl, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclo, alkoxyalkyl, (amino)alkyl, hydroxyalkylamino, (alkylamino)alkyl, (dialkylamino)alkyl, (cyano)alkyl, (carboxamido)alkyl, mercaptoalkyl, (heterocyclo)alkyl, or (heteroaryl)alkyl. In one embodiment, the optionally substituted cycloalkyl is substituted with two substituents. In another embodiment, the optionally substituted cycloalkyl is substituted with one substituent. In one embodiment, the optionally substituted cycloalkyl is substituted with at least one amino, alkylamino, or dialkylamino group. Non-limiting exemplary optionally substituted c cloalk l rou s include:
Figure imgf000043_0001
[0055] For the purpose of the pr term "cycloalkenyl" as used by itself or part of another group refers aturated cycloalkyl group as defined above. In one embodiment, the cycloalkenyl has one carbon-to-carbon double bond. In another embodiment, the cycloalkenyl group is chosen from a C4-8 cycloalkenyl group. Exemplary cycloalkenyl groups include cyclopentenyl and cyclohexenyl.
[0056] For the purpose of the present disclosure, the term "optionally substituted cycloalkenyl" as used by itself or as part of another group means that the cycloalkenyl as defined above is either unsubstituted or substituted with one, two, or three substituents independently chosen from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, monohydroxyalkyl, dihydroxyalkyl, alkoxy, haloalkoxy, aryloxy, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclo, alkoxyalkyl, (amino)alkyl, hydroxyalkylamino, (alkylamino)alkyl, (dialkylamino)alkyl, (cyano)alkyl, (carboxamido)alkyl, mercaptoalkyl, (heterocyclo)alkyl, and (heteroaryl)alkyl. In one embodiment, the optionally substituted cycloalkenyl is substituted with two substituents. In another embodiment, the optionally substituted cycloalkenyl is substituted with one substituent. In another embodiment, the cycloalkenyl is unsubstituted.
[0057] For the purpose of the present disclosure, the term "alkenyl" as used by itself or as part of another group refers to an alkyl group as defined above containing one, two or three carbon-to-carbon double bonds. In one embodiment, the alkenyl group is chosen from a C2-6 alkenyl group. In another embodiment, the alkenyl group is chosen from a C2-4 alkenyl group. Non-limiting exemplary alkenyl groups include ethenyl, propenyl, isopropenyl, butenyl, sec-butenyl, pentenyl, and hexenyl.
[0058] For the purpose of the present disclosure, the term "optionally substituted alkenyl" as used herein by itself or as part of another group means the alkenyl as defined above is either unsubstituted or substituted with one, two or three substituents independently chosen from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aryloxy, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino carboxy carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, opt aryl, optionally substituted heteroaryl, or optionally substituted heteroc
[0059] For the purpose of the p he term "alkynyl" as used by itself or as part of another group refers to an alkyl group as defined above containing one to three carbon-to-carbon triple bonds. In one embodiment, the alkynyl has one carbon-to-carbon triple bond. In one embodiment, the alkynyl group is chosen from a C2-6 alkynyl group. In another embodiment, the alkynyl group is chosen from a C2-4 alkynyl group. Non-limiting exemplary alkynyl groups include ethynyl, propynyl, butynyl, 2-butynyl, pentynyl, and hexynyl groups.
[0060] For the purpose of the present disclosure, the term "optionally substituted alkynyl" as used herein by itself or as part of another group means the alkynyl as defined above is either unsubstituted or substituted with one, two or three substituents independently chosen from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aryloxy, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl, optionally substituted heteroaryl, or optionally substituted heterocyclo.
[0061] For the purpose of the present disclosure, the term "haloalkyl" as used by itself or as part of another group refers to an alkyl group substituted by one or more fluorine, chlorine, bromine and/or iodine atoms. In one embodiment, the alkyl group is substituted by one, two, or three fluorine and/or chlorine atoms. In another embodiment, the haloalkyl group is chosen from a C1-4 haloalkyl group. Non-limiting exemplary haloalkyl groups include fluoromethyl, difluoromethyl, trifluoromethyl, pentafluoroethyl, 1,1-difluoroethyl, 2,2-difluoroethyl, 2,2,2-trifluoroethyl, 3,3,3-trifluoropropyl, 4,4,4-trifluorobutyl, and trichloromethyl groups.
[0062] For the purpose of the present disclosure, the term "hydroxyalkyl" as used by itself or as part of another group refers to an alkyl group substituted with one or more, e.g., one, two, or three, hydroxy groups. In one embodiment, the hydroxyalkyl group is a monohydroxyalkyl group, i.e., substituted with one hydroxy group. In another embodiment, the hydroxyalkyl group is a dihydroxyalkyl group, i.e., substituted with two hydroxy groups. In another embodiment the hydroxyalkyl group is chosen from a C1-4 hydroxyalkyl group. N lary hydroxyalkyl groups include hydroxymethyl, hydroxyethyl, nd hydroxybutyl groups, such as 1-hydroxyethyl, 2-hydroxyethyl, l, 2-hydroxypropyl, 3-hydroxypropyl, 3-hydroxybutyl, 4-hydroxybutyl, 2-hydroxy-1-methylpropyl, and 1,3-dihydroxyprop-2- yl.
[0063] For the purpose of the present disclosure, the term "alkoxy" as used by itself or as part of another group refers to an optionally substituted alkyl, optionally substituted cycloalkyl, optionally substituted alkenyl or optionally substituted alkynyl attached to a terminal oxygen atom. In one embodiment, the alkoxy group is chosen from a C1-4 alkoxy group. In another embodiment, the alkoxy group is chosen from a C1-4 alkyl attached to a terminal oxygen atom, e.g., methoxy, ethoxy, and tert-butoxy.
[0064] For the purpose of the present disclosure, the term "alkylthio" as used by itself or as part of another group refers to a sulfur atom substituted by an optionally substituted alkyl group. In one embodiment, the alkylthio group is chosen from a C1-4 alkylthio group. Non-limiting exemplary alkylthio groups include -SCH3, and -SCH2CH3.
[0065] For the purpose of the present disclosure, the term "alkoxyalkyl" as used by itself or as part of another group refers to an alkyl group substituted with an alkoxy group. Non-limiting exemplary alkoxyalkyl groups include methoxymethyl, methoxyethyl, methoxypropyl, methoxybutyl, ethoxymethyl, ethoxyethyl, ethoxypropyl, ethoxybutyl, propoxymethyl, iso-propoxymethyl, propoxyethyl, propoxypropyl, butoxymethyl, tert- butoxymethyl, isobutoxymethyl, sec-butoxymethyl, and pentyloxymethyl.
[0066] For the purpose of the present disclosure, the term "haloalkoxy" as used by itself or as part of another group refers to a haloalkyl attached to a terminal oxygen atom. Non-limiting exemplary haloalkoxy groups include fluoromethoxy, difluoromethoxy, trifluoromethoxy, and 2,2,2-trifluoroethoxy.
[0067] For the purpose of the present disclosure, the term "heteroalkyl" as used by itself or part of another group refers to a stable straight or branched chain hydrocarbon radical containing 1 to 10 carbon atoms and at least two heteroatoms, which can be the same or different, selected from O, N, or S, wherein: 1) the nitrogen atom(s) and sulfur atom(s) can optionally be oxidized; and/or 2) the nitrogen atom(s) can optionally be quaternized. The heteroatoms can be placed at any interior position of the heteroalkyl group or at a position at which the heteroalkyl group is attached to the remainder of the molecule. In one embodiment, the heteroalkyl group contains two oxygen atoms. In one embodiment, the heteroalkyl contains one ox ogen atom. In one embodiment, the heteroalkyl contains two nitrog miting exemplary heteroalkyl groups include -CH2OCH2CH2OCH3, CH2OCH3, -CH2NHCH2CH2OCH2, -OCH2CH2NH2, -NHCH2CH2N(H)CH3, -NHCH2CH2OCH3 and -OCH2CH2OCH3.
[0068] For the purpose of the present disclosure, the term "aryl" as used by itself or as part of another group refers to a monocyclic or bicyclic aromatic ring system having from six to fourteen carbon atoms (i.e., C6-14 aryl). Non-limiting exemplary aryl groups include phenyl (abbreviated as "Ph"), naphthyl, phenanthryl, anthracyl, indenyl, azulenyl, biphenyl, biphenylenyl, and fluorenyl groups. In one embodiment, the aryl group is chosen from phenyl or naphthyl. In one embodiment, the aryl group is phenyl.
[0069] For the purpose of the present disclosure, the term "optionally substituted aryl" as used herein by itself or as part of another group means that the aryl as defined above is either unsubstituted or substituted with one to five substituents independently selected from the group consisting of halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aryloxy, heteroaryloxy, aralkyl, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclo, alkoxyalkyl, (amino)alkyl, hydroxyalkylamino, (alkylamino)alkyl, (dialkylamino)alkyl, (cyano)alkyl, (carboxamido)alkyl, mercaptoalkyl, (heterocyclo)alkyl, (cycloalkylamino)alkyl, (C1-4 haloalkoxy)alkyl, (heteroaryl)alkyl, -N(R43)(R44), and -N(H)C(=O)-R45, wherein R43 is hydrogen or C1-4 alkyl; R44 is alkoxyalkyl, (heterocyclo)alkyl, (amino)alkyl, (alkylamino)alkyl, or (dialkylamino)alkyl; and R45 is alkyl, optionally substituted aryl or optionally substituted heteroaryl. In one embodiment, the optionally substituted aryl is an optionally substituted phenyl. In one embodiment, the optionally substituted phenyl has four substituents. In another embodiment, the optionally substituted phenyl has three substituents. In another embodiment, the optionally substituted phenyl has two substituents. In another embodiment, the optionally substituted phenyl has one substituent. In another embodiment, the optionally substituted phenyl has one amino, alkylamino, dialkylamino, (amino)alkyl, (alkylamino)alkyl, or (dialkylamino)alkyl substituent. Non-limiting exemplary substituted aryl groups include 2-methylphenyl, 2-methoxyphenyl, 2-fluorophenyl 2 chlorophenyl, 2-bromophenyl, 3-methylphenyl, 3-methoxyphenyl, 3-fluorophe yl, 4-methylphenyl, 4-ethylphenyl, 4-methoxyphenyl, 4-fluoroph nyl, 2,6-di-fluorophenyl, 2,6-di- chlorophenyl, 2-methyl, 3-m -ethyl, 3-methoxyphenyl, 3,4-di- methoxyphenyl, 3,5-di-fluorophenyl 3,5-di-methylphenyl, 3,5-dimethoxy, 4-methylphenyl, 2-fluoro-3-chlorophenyl, 3-chloro-4-fluorophenyl, and 2-phenylpropan- 2-amine. The term optionally substituted aryl is meant to include groups having fused optionally substituted cycloalkyl and fused optionally substituted heterocyclo rings. Examples include:
Figure imgf000048_0001
.
[0070] For the purpose of the present disclosure, the term "aryloxy" as used by itself or as part of another group refers to an optionally substituted aryl attached to a terminal oxygen atom. A non-limiting exemplary aryloxy group is PhO-.
[0071] For the purpose of the present disclosure, the term "heteroaryloxy" as used by itself or as part of another group refers to an optionally substituted heteroaryl attached to a terminal oxygen atom.
[0072] For the purpose of the present disclosure, the term "aralkyloxy" or "arylalkyloxy" as used by itself or as part of another group refers to an aralkyl group attached to a terminal oxygen atom. A non-limiting exemplary aralkyloxy group is PhCH2O-.
[0073] For the purpose of the present disclosure, the term "heteroaryl" or "heteroaromatic" refers to monocyclic and bicyclic aromatic ring systems having 5 to 14 ring atoms (i.e., a 5- to 14-membered heteroaryl) and 1, 2, 3, or 4 heteroatoms independently chosen from oxygen, nitrogen or sulfur. In one embodiment, the heteroaryl has three heteroatoms. In another embodiment, the heteroaryl has two heteroatoms. In another embodiment, the heteroaryl has one heteroatom. In another embodiment, the heteroaryl is a 5- to 10-membered heteroaryl. In one embodiment, the heteroaryl has 5 ring atoms, e.g., thienyl, a 5-membered heteroaryl having four carbon atoms and one sulfur atom. In another embodiment, the heteroaryl has 6 ring atoms, e.g., pyridyl, a 6-membered heteroaryl having five carbon atoms and one nitrogen atom. Non- limiting exemplary heteroaryl groups include thienyl, benzo[b]thienyl, naphtho[2,3- b]thienyl, thianthrenyl, furyl, benzofuryl, pyranyl, isobenzofuranyl, benzooxazonyl, chromenyl, xanthenyl, 2H-pyrrolyl pyrrolyl imidazolyl, pyrazolyl, pyridyl, pyrazinyl, pyrimidinyl, pyridazinyl, isoind dolyl, indazolyl, purinyl, isoquinolyl, quinolyl, phthalazinyl, naphth yl, quinazolinyl, pteridinyl, 4aH- carbazolyl, carbazolyl, ȕ-ca thridinyl, acridinyl, pyrimidinyl, phenanthrolinyl, phenazinyl, thiazolyl, isothiazolyl, phenothiazolyl, isoxazolyl, furazanyl, and phenoxazinyl. In one embodiment, the heteroaryl is chosen from thienyl (e.g., thien- 2-yl and thien-3-yl), furyl (e.g., 2-furyl and 3-furyl), pyrrolyl (e.g., 1H-pyrrol-2-yl and 1H-pyrrol-3-yl), imidazolyl (e.g., 2H-imidazol-2-yl and 2H-imidazol-4-yl), pyrazolyl (e.g., 1H-pyrazol-3-yl, 1H-pyrazol-4-yl, and 1H-pyrazol-5-yl), pyridyl (e.g., pyridin-2-yl, pyridin-3-yl, and pyridin-4-yl), pyrimidinyl (e.g., pyrimidin-2-yl, pyrimidin-4-yl, and pyrimidin-5-yl), thiazolyl (e.g., thiazol-2-yl, thiazol-4-yl, and thiazol-5-yl), isothiazolyl (e.g., isothiazol-3-yl, isothiazol-4-yl, and isothiazol-5-yl), oxazolyl (e.g., oxazol-2-yl, oxazol-4-yl, and oxazol-5-yl) and isoxazolyl (e.g., isoxazol-3-yl, isoxazol-4-yl, and isoxazol-5-yl). The term "heteroaryl" is also meant to include possible N-oxides. Exemplary N-oxides include pyridyl N-oxide.
[0074] For the purpose of the present disclosure, the term "optionally substituted heteroaryl" as used by itself or as part of another group means that the heteroaryl as defined above is either unsubstituted or substituted with one to four substituents, e.g., one or two substituents, independently chosen from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aralkyl, aryloxy, aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, optionally substituted cycloalkyl, alkenyl, alkynyl, optionally substituted aryl, optionally substituted heteroaryl, optionally substituted heterocyclo, alkoxyalkyl, (amino)alkyl, hydroxyalkylamino, (alkylamino)alkyl, (dialkylamino)alkyl, (cyano)alkyl, (carboxamido)alkyl, mercaptoalkyl, (heterocyclo)alkyl, (heteroaryl)alkyl, -N(R43)(R44), or -N(H)C(=O)-R45, wherein R43 is hydrogen or C1-4 alkyl; R44 is alkoxyalkyl, (heterocyclo)alkyl, (amino)alkyl, (alkylamino)alkyl, or (dialkylamino)alkyl; and R45 is alkyl, optionally substituted aryl, or optionally substituted heteroaryl. In one embodiment, the optionally substituted heteroaryl has one substituent. In one embodiment, the substituent is amino, alkylamino, dialkylamino, (amino)alkyl, hydroxyalkylamino, (alkylamino)alkyl, (dialkylamino)alkyl, (heterocyclo)alkyl, -N(R43)(R44), or -N(H)C(=O)-R45. In one embodiment, the optionally substituted is an optionally substituted pyridyl, i.e 2 3 or 4 pyridyl. Any available carbon or nitrogen atom can be substituted.
[0075] For the purpose of the pr e term "heterocycle" or "heterocyclo" as used by itself or as part of an to saturated and partially unsaturated (e.g., containing one or two double bonds) cyclic groups containing one, two, or three rings having from three to fourteen ring members (i.e., a 3- to 14-membered heterocyclo) and at least one heteroatom. Each heteroatom is independently selected from the group consisting of oxygen, sulfur, including sulfoxide and sulfone, and/or nitrogen atoms, which can be quaternized. The term "heterocyclo" is meant to include cyclic ureido groups such as imidazolidinyl-2-one, cyclic amide groups such as ȕ-lactam, Ȗ-lactam, į-lactam and İ-lactam, and cyclic carbamate groups such as oxazolidinyl-2-one. The term "heterocyclo" is also meant to include groups having fused optionally substituted aryl groups, e.g., indolinyl, indolinyl-2-one, benzo[d]oxazolyl-2(3H)-one. In one embodiment, the heterocyclo group is chosen from a 4-, 5-, 6-, 7- or 8-membered cyclic group containing one ring and one or two oxygen and/or nitrogen atoms. In one embodiment, the heterocyclo group is chosen from a 5- or 6-membered cyclic group containing one ring and one or two nitrogen atoms. In one embodiment, the heterocyclo group is chosen from a 8-, 9-, 10-, 11-, or 12-membered cyclic group containing two rings and one or two nitrogen atoms. The heterocyclo can be optionally linked to the rest of the molecule through a carbon or nitrogen atom. Non-limiting exemplary heterocyclo groups include 2-oxopyrrolidin-3-yl, 2-imidazolidinone, piperidinyl, morpholinyl, piperazinyl, pyrrolidinyl, 8-azabicyclo[3.2.1]octane (nortropane), 6-azaspiro[2.5]octane, 6-azaspiro[3.4]octane, indolinyl, indolinyl-2-one, 1,3-dihydro-2H-benzo[d]imidazol-2- one
[0076] For the purpose of the present disclosure, the term "optionally substituted heterocyclo" as used herein by itself or part of another group means the heterocyclo as defined above is either unsubstituted or substituted with one to four substituents independently selected from halo, nitro, cyano, hydroxy, amino, alkylamino, dialkylamino, haloalkyl, hydroxyalkyl, alkoxy, haloalkoxy, aryloxy, aralkyl aralkyloxy, alkylthio, carboxamido, sulfonamido, alkylcarbonyl, arylcarbonyl, alkylsulfonyl, arylsulfonyl, ureido, guanidino, carboxy, carboxyalkyl, alkyl, cycloalkyl, alkenyl, alkynyl, aryl, heteroaryl, heterocyclo, alkoxyalkyl, (amino)alkyl, hydroxyalkylamino, (alkylamino)alkyl, (dialkylamino)alkyl, (cyano)alkyl, (carboxamido)alkyl, mercaptoalkyl, (heterocyclo)alkyl and (heteroaryl)alkyl. Substitution may occur on any available carbon or nitrogen ato spirocycle. In one embodiment, the optionally substituted heterocyc th at least one amino, alkylamino, or dialkylamino group. Non-limit onally substituted heterocyclo groups include:
Figure imgf000051_0001
.
[0077] For the purpose of the present disclosure, the term "amino" as used by itself or as part of another group refers to -NH2.
[0078] For the purpose of the present disclosure, the term "alkylamino" as used by itself or as part of another group refers to -NHR22, wherein R22 is C1-6 alkyl. In one embodiment, R22 is C1-4 alkyl. Non-limiting exemplary alkylamino groups include -N(H)CH3 and -N(H)CH2CH3.
[0079] For the purpose of the present disclosure, the term "dialkylamino" as used by itself or as part of another group refers to -NR23aR23b, wherein R23a and R23b are each independently C1-6 alkyl. In o 23a and R23b are each independently C1-4 alkyl. Non-limiting exe no groups include -N(CH3)2 and -N(CH3)CH2CH(CH3)2.
[0080] For the purpose of the present disclosure, the term "hydroxyalkylamino" as used by itself or as part of another group refers to -NHR24, wherein R24 is hydroxyalkyl.
[0081] For the purpose of the present disclosure, the term "cycloalkylamino" as used by itself or as part of another group refers to -NR25aR25b, wherein R25a is optionally substituted cycloalkyl and R25b is hydrogen or C1-4 alkyl. [0082] For the purpose of the present disclosure, the term "aralkylamino" as used by itself or as part of another group refers to -NR26aR26b, wherein R26a is aralkyl and R26b is hydrogen or C1-4 alkyl. Non-limiting exemplary aralkylamino groups include -N(H)CH2Ph and -N(CH3)CH2Ph.
[0083] For the purpose of the present disclosure, the term "(amino)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with an amino group. In one embodiment, the alkyl is a C1-4 alkyl. Non-limiting exemplary (amino)alkyl groups include -CH2NH2, -C(NH2)(H)CH3, -CH2CH2NH2, -CH2C(NH2)(H)CH3, -CH2CH2CH2NH2, -CH2CH2CH2CH2NH2, and -CH2C(CH3)2CH2NH2
[0084] For the purpose of the present disclosure, the term "(alkylamino)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with an alkylamino group. In one embodiment, the alkyl is a C1-4 alkyl. A non-limiting exemplary (alkylamino)alkyl group is -CH2CH2N(H)CH3.
[0085] For the purpose of the present disclosure, the term "(dialkylamino)alkyl" as used by itself or as part of another group refers to an alkyl group substituted by a dialkylamino group. In one embodiment, the alkyl is a C1-4 alkyl. Non-limiting exemplary (dialkylamino)alkyl groups are -CH2CH2N(CH3)2.
[0086] For the purpose of the present disclosure, the term "(cycloalkylamino)alkyl" as used by itself or as part of another group refers to an alkyl group substituted by a cycloalkylamino group. In one embodiment, the alkyl is a C1-4 alkyl. Non-limiting exemplary (cycloalkylamino)alkyl groups include -CH2N(H)cyclopropyl, -CH2N(H)cyclobutyl, and -CH2N(H)cyclohexyl.
[0087] For the purpose of the present disclosure, the term "(aralkylamino)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with an aralkylamino group. In one embodiment the alkyl is a C1-4 alkyl. A non-limiting exemplary (aralkylamino)alkyl g H2N(H)CH2Ph.
[0088] For the purpose of the pr term "(cyano)alkyl" as used by itself or as part of another group refe substituted with one or more cyano, e.g., -CN, groups. In one embodiment, the alkyl is a C1-4 alkyl. Non-limiting exemplary (cyano)alkyl groups include -CH2CH2CN, -CH2CH2CH2CN, and -CH2CH2CH2CH2CN.
[0089] For the purpose of the present disclosure, the term "(amino)(hydroxy)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with one amino, alkylamino, or dialkylamino group and one hydroxy group. In one embodiment, the alkyl is a C1-6 alkyl. In another embodiment, the alkyl is a C1-4 alkyl. Non-limiting exemplary (amino)(hydroxy)alkyl groups include:
Figure imgf000053_0001
.
[0090] For the purpose of the present disclosure, the term "(amino)(aryl)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with one amino, alkylamino, or dialkylamino group and one optionally substituted aryl group. In one embodiment, the alkyl is a C1-6 alkyl. In one embodiment, the optionally substituted aryl group is an optionally substituted phenyl. Non-limiting exemplary (amino)(aryl)alkyl groups include:
Figure imgf000053_0002
.
[0091] For the purpose of the present disclosure, the term "(cycloalkyl)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with one optionally substituted cycloalkyl group. In one embodiment, the alkyl is a C1-4 alkyl. In one embodiment, the cycloalkyl is a C3-6 cycloalkyl. In one embodiment, the optionally substituted cycloalkyl group is substituted with an amino or (amino)alkyl group. Non-limiting exemplary (cycloalk l) lk l i clude:
Figure imgf000054_0001
[0092] For the purpose of the present disclosure, the term "(hydroxy)(aryl)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with one hydroxy group and one optionally substituted aryl group. In one embodiment, the alkyl is a C- 1-6 alkyl. In one embodiment, the optionally substituted aryl group is an optionally substituted hen l. Non-limitin exem lar h drox ar l alk l rou s include:
[0093] For the purpose of the present disclosure, the term "carboxamido" as used by itself or as part of another group refers to a radical of formula -C(=O)NR26aR26b, wherein R26a and R26b are each independently hydrogen, optionally substituted alkyl, optionally substituted aryl, or optionally substituted heteroaryl, or R26a and R26b taken together with the nitrogen to which they are attached from a 3- to 8-membered heterocyclo group. In one embodiment, R26a and R26b are each independently hydrogen or optionally substituted alkyl. Non-limiting exemplary carboxamido groups include -CONH2, -CON(H)CH3, CON(CH3)2, and -CON(H)Ph.
[0094] For the purpose of the p e term "(carboxamido)alkyl" as used by itself or as part of anothe an alkyl group substituted with a carboxamido group. Non-lim (carboxamido)alkyl groups include -CH2CONH2, -C(H)CH3-CONH )CH3.
[0095] For the purpose of the present disclosure, the term "sulfonamido" as used by itself or as part of another group refers to a radical of the formula -SO2NR27aR27b, wherein R27a and R27b are each independently hydrogen, optionally substituted alkyl, or optionally substituted aryl, or R27a and R27b taken together with the nitrogen to which they are attached from a 3- to 8-membered heterocyclo group. Non-limiting exemplary sulfonamido groups include -SO2NH2, -SO2N(H)CH3, and -SO2N(H)Ph.
[0096] For the purpose of the present disclosure, the term "alkylcarbonyl" as used by itself or as part of another group refers to a carbonyl group, i.e., -C(=O)-, substituted by an alkyl group. A non-limiting exemplary alkylcarbonyl group is -COCH3.
[0097] For the purpose of the present disclosure, the term "arylcarbonyl" as used by itself or as part of another group refers to a carbonyl group, i.e., -C(=O)-, substituted by an optionally substituted aryl group. A non-limiting exemplary arylcarbonyl group is -COPh.
[0098] For the purpose of the present disclosure, the term "alkylsulfonyl" as used by itself or as part of another group refers to a sulfonyl group, i.e., -SO2-, substituted by any of the above-mentioned optionally substituted alkyl groups. A non-limiting exemplary alkylsulfonyl group is -SO2CH3.
[0099] For the purpose of the present disclosure, the term "arylsulfonyl" as used by itself or as part of another group refers to a sulfonyl group, i.e., -SO2-, substituted by any of the above-mentioned optionally substituted aryl groups. A non-limiting exemplary arylsulfonyl group is -SO2Ph.
[0100] For the purpose of the present disclosure, the term "mercaptoalkyl" as used by itself or as part of another group refers to any of the above-mentioned alkyl groups substituted by a–SH group.
[0101] For the purpose of the present disclosure, the term "carboxy" as used by itself or as part of another group refers to a radical of the formula -COOH.
[0102] For the purpose of the present disclosure, the term "carboxyalkyl" as used by itself or as part of another group refers to any of the above-mentioned alkyl groups substituted with a -COOH. A non limiting exemplary carboxyalkyl group is -CH2CO2H.
[0103] For the purpose of the p he term "alkoxycarbonyl" as used by itself or as part of another group yl group, i.e., -C(=O)-, substituted by an alkoxy group. Non-limiti xycarbonyl groups are -CO2Me and -CO2Et.
[0104] For the purpose of the present disclosure, the term "aralkyl" or“arylalkyl” as used by itself or as part of another group refers to an alkyl group substituted with one, two, or three optionally substituted aryl groups. In one embodiment, the aralkyl group is a C1-4 alkyl substituted with one optionally substituted aryl group. Non-limiting exemplary aralkyl groups include benzyl, phenethyl, -CHPh2, -CH2(4-OH-Ph), and -CH(4-F-Ph)2.
[0105] For the purpose of the present disclosure, the term "ureido" as used by itself or as part of another group refers to a radical of the formula -NR30a-C(=O)-NR30bR30c, wherein R22a is hydrogen, alkyl, or optionally substituted aryl, and R30b and R30c are each independently hydrogen, alkyl, or optionally substituted aryl, or R30b and R30c taken together with the nitrogen to which they are attached form a 4- to 8-membered heterocyclo group. Non-limiting exemplary ureido groups include -NH-C(C=O)-NH2 and -NH-C(C=O)-NHCH3.
[0106] For the purpose of the present disclosure, the term "guanidino" as used by itself or as part of another group refers to a radical of the formula -NR28a-C(=NR29)-NR28bR28c, wherein R28a, R28b, and R28c are each independently hydrogen, alkyl, or optionally substituted aryl, and R29 is hydrogen, alkyl, cyano, alkylsulfonyl, alkylcarbonyl, carboxamido, or sulfonamido. Non-limiting exemplary guanidino groups include -NH- C(C=NH)-NH2, -NH-C(C=NCN)-NH2, and -NH-C(C=NH)-NHCH3.
[0107] For the purpose of the present disclosure, the term "(heterocyclo)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with one, two, or three optionally substituted heterocyclo groups. In one embodiment, the (heterocyclo)alkyl is a C1-4 alkyl substituted with one optionally substituted heterocyclo group. The heterocyclo can be linked to the alkyl group through a carbon or nitrogen atom. Non-limiting exemplary (heterocyclo)alkyl groups include:
Figure imgf000056_0001
.
[0108] For the purpose of the p e term "(heteroaryl)alkyl" as used by itself or as part of another group refers to an alkyl group substituted with one, two, or three optionally substituted heteroaryl groups. In one embodiment, the (heteroaryl)alkyl group is a C1-4 alkyl substituted with one optionally substituted heteroaryl group. Non-limiting exemplary (heteroaryl)alkyl groups include: ,
Figure imgf000057_0001
.
[0109] For the purpose of the present disclosure, the term "alkylcarbonylamino" as used by itself or as part of another group refers to an alkylcarbonyl group attached to an amino. A non-limiting exemplary alkylcarbonylamino group is -NHCOCH3.
[0110] The present disclosure encompasses any of the Compounds of the Disclosure being isotopically-labelled (i.e., radiolabeled) by having one or more atoms replaced by an atom having a different atomic mass or mass number. Examples of isotopes that can be incorporated into the disclosed compounds include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorous, fluorine and chlorine, such as 2H (or deuterium (D)), 3H, 11C, 13C, 14C, 15N, 18O, 17O, 31P, 32P, 35S, 18F, and 36Cl, respectively, e.g., 3H, 11C, and 14C. In one embodiment, provided is a composition wherein substantially all of the atoms at a position within the Compound of the Disclosure are replaced by an atom having a different atomic mass or mass number. In another embodiment, provided is a composition wherein a portion of the atoms at a position within the Compound of the disclosure are replaced, i.e., the Compound of the Disclosure is enriched at a position with an atom having a different atomic mass or mass number." Isotopically-labelled Compounds of the Disclosure can be prepared by methods known in the art.
[0111] Compounds of the Disclosure may contain one or more asymmetric centers and may thus give rise to enantiomers, diastereomers, and other stereoisomeric forms. The present disclosure is meant to encompass the use of all such possible forms, as well as their racemic and resolved form eof. The individual enantiomers can be separated according to meth art in view of the present disclosure. When the compounds described finic double bonds or other centers of geometric asymmetry, and unle p wise, it is intended that they include both E and Z geometric isomers. All tautomers are intended to be encompassed by the present disclosure as well.
[0112] As used herein, the term "stereoisomers" is a general term for all isomers of individual molecules that differ only in the orientation of their atoms in space. It includes enantiomers and isomers of compounds with more than one chiral center that are not mirror images of one another (diastereomers).
[0113] The term "chiral center" or "asymmetric carbon atom" refers to a carbon atom to which four different groups are attached.
[0114] The terms "enantiomer" and "enantiomeric" refer to a molecule that cannot be superimposed on its mirror image and hence is optically active wherein the enantiomer rotates the plane of polarized light in one direction and its mirror image compound rotates the plane of polarized light in the opposite direction.
[0115] The term "racemic" refers to a mixture of equal parts of enantiomers and which mixture is optically inactive.
[0116] The term "absolute configuration" refers to the spatial arrangement of the atoms of a chiral molecular entity (or group) and its stereochemical description, e.g., R or S.
[0117] The stereochemical terms and conventions used in the specification are meant to be consistent with those described in Pure & Appl. Chem 68:2193 (1996), unless otherwise indicated.
[0118] The term "enantiomeric excess" or "ee" refers to a measure for how much of one enantiomer is present compared to the other. For a mixture of R and S enantiomers, the percent enantiomeric excess is defined as ŇR - SŇ*100, where R and S are the respective mole or weight fractions of enantiomers in a mixture such that R + S = 1. With knowledge of the optical rotation of a chiral substance, the percent enantiomeric excess is defined as ([D]obs/[D]max)*100, where [D]obs is the optical rotation of the mixture of enantiomers and [D]max is the optical rotation of the pure enantiomer. Determination of enantiomeric excess is possible using a variety of analytical techniques, including NMR spectroscopy, chiral column chromatography or optical polarimetry.
[0119] The terms "enantiomeric ll " " tiopure" refer to a sample of a chiral substance all of whose molecule of detection) have the same chirality sense.
[0120] The terms "enantiomeric nantioenriched" refer to a sample of a chiral substance whose enantiomeric ratio is greater than 50:50. Enantiomerically enriched compounds may be enantiomerically pure.
[0121] The terms "a" and "an" refer to one or more.
[0122] The term "about," as used herein, includes the recited number ± 10%. Thus, "about 10" means 9 to 11. [0123] The present disclosure encompasses the preparation and use of salts of the Compounds of the Disclosure, including non-toxic pharmaceutically acceptable salts. Examples of pharmaceutically acceptable addition salts include inorganic and organic acid addition salts and basic salts. The pharmaceutically acceptable salts include, but are not limited to, metal salts such as sodium salt, potassium salt, cesium salt and the like; alkaline earth metals such as calcium salt, magnesium salt and the like; organic amine salts such as triethylamine salt, pyridine salt, picoline salt, ethanolamine salt, triethanolamine salt, dicyclohexylamine salt, N,N'-dibenzylethylenediamine salt and the like; inorganic acid salts such as hydrochloride, hydrobromide, phosphate, sulphate and the like; organic acid salts such as citrate, lactate, tartrate, maleate, fumarate, mandelate, acetate, dichloroacetate, trifluoroacetate, oxalate, formate and the like; sulfonates such as methanesulfonate, benzenesulfonate, p-toluenesulfonate and the like; and amino acid salts such as arginate, asparginate, glutamate and the like. The term "pharmaceutically acceptable salt" as used herein, refers to any salt, e.g., obtained by reaction with an acid or a base, of a Compound of the Disclosure that is physiologically tolerated in the target patient (e.g., a mammal, e.g., a human).
[0124] Acid addition salts can be formed by mixing a solution of the particular Compound of the Disclosure with a solution of a pharmaceutically acceptable non-toxic acid such as hydrochloric acid, fumaric acid, maleic acid, succinic acid, acetic acid, citric acid, tartaric acid, carbonic acid, phosphoric acid, oxalic acid, dichloroacetic acid, or the like. Basic salts can be formed by mixing a solution of the compound of the present disclosure with a solution of a pharmaceutically acceptable non-toxic base such as sodium hydroxide, potassium hydroxide, choline hydroxide, sodium carbonate and the like.
[0125] The present disclosure encompasses the preparation and use of solvates of Compounds of the Disclosure ally do not significantly alter the physiological activity or toxic nds, and as such may function as pharmacological equivalents. " as used herein is a combination, physical association and/or solvation of a compound of the present disclosure with a solvent molecule such as, e.g. a disolvate, monosolvate or hemisolvate, where the ratio of solvent molecule to compound of the present disclosure is about 2:1, about 1:1 or about 1:2, respectively. This physical association involves varying degrees of ionic and covalent bonding, including hydrogen bonding. In certain instances, the solvate can be isolated, such as when one or more solvent molecules are incorporated into the crystal lattice of a crystalline solid. Thus, "solvate" encompasses both solution-phase and isolatable solvates. Compounds of the Disclosure can be present as solvated forms with a pharmaceutically acceptable solvent, such as water, methanol, ethanol, and the like, and it is intended that the disclosure includes both solvated and unsolvated forms of Compounds of the Disclosure. One type of solvate is a hydrate. A "hydrate" relates to a particular subgroup of solvates where the solvent molecule is water. Solvates typically can function as pharmacological equivalents. Preparation of solvates is known in the art. See, for example, M. Caira et al, J. Pharmaceut. Sci., 93(3):601-611 (2004), which describes the preparation of solvates of fluconazole with ethyl acetate and with water. Similar preparation of solvates, hemisolvates, hydrates, and the like are described by E.C. van Tonder et al., AAPS Pharm. Sci. Tech., 5(1):Article 12 (2004), and A.L. Bingham et al., Chem. Commun. 603-604 (2001). A typical, non-limiting, process of preparing a solvate would involve dissolving a Compound of the Disclosure in a desired solvent (organic, water, or a mixture thereof) at temperatures above 20°C to about 25°C, then cooling the solution at a rate sufficient to form crystals, and isolating the crystals by known methods, e.g., filtration. Analytical techniques such as infrared spectroscopy can be used to confirm the presence of the solvent in a crystal of the solvate.
[0126] Since Compounds of the Disclosure are inhibitors of SMYD proteins, such as SMYD3 and SMYD2, a number of diseases, conditions, or disorders mediated by SMYD proteins, such as SMYD3 and SMYD2, can be treated by employing these compounds. The present disclosure is thus directed generally to a method for treating a disease, condition, or disorder responsive to the inhibition of SMYD proteins, such as SMYD3 and SMYD2, in an animal suffering from, or at risk of suffering from, the disorder, the method comprising administering to the animal an effective amount of one or more Compounds of the Disclosure.
[0127] The present disclosure to a method of inhibiting SMYD proteins in an animal in need the omprising administering to the animal a therapeutically effective amount of at least one Compound of the Disclosure.
[0128] The present disclosure is further directed to a method of inhibiting SMYD3 in an animal in need thereof, the method comprising administering to the animal a therapeutically effective amount of at least one Compound of the Disclosure. [0129] The present disclosure is further directed to a method of inhibiting SMYD2 in an animal in need thereof, the method comprising administering to the animal a therapeutically effective amount of at least one Compound of the Disclosure.
[0130] As used herein, the terms "treat," "treating," "treatment," and the like refer to eliminating, reducing, or ameliorating a disease or condition, and/or symptoms associated therewith. Although not precluded, treating a disease or condition does not require that the disease, condition, or symptoms associated therewith be completely eliminated. As used herein, the terms "treat," "treating," "treatment," and the like may include "prophylactic treatment," which refers to reducing the probability of redeveloping a disease or condition, or of a recurrence of a previously-controlled disease or condition, in a subject who does not have, but is at risk of or is susceptible to, redeveloping a disease or condition or a recurrence of the disease or condition. The term "treat" and synonyms contemplate administering a therapeutically effective amount of a Compound of the Disclosure to an individual in need of such treatment.
[0131] Within the meaning of the disclosure, "treatment" also includes relapse prophylaxis or phase prophylaxis, as well as the treatment of acute or chronic signs, symptoms and/or malfunctions. The treatment can be orientated symptomatically, for example, to suppress symptoms. It can be effected over a short period, be oriented over a medium term, or can be a long-term treatment, for example within the context of a maintenance therapy.
[0132] The term "therapeutically effective amount" or "effective dose" as used herein refers to an amount of the active ingredient(s) that is(are) sufficient, when administered by a method of the disclosure, to efficaciously deliver the active ingredient(s) for the treatment of condition or disease of interest to an individual in need thereof. In the case of a cancer or other proliferation disorder the therapeutically effective amount of the agent may reduce (i.e., retard d preferably stop) unwanted cellular proliferation; reduce the number uce the tumor size; inhibit (i.e., retard to some extent and preferably st tration into peripheral organs; inhibit (i.e., retard to some extent and preferably stop) tumor metastasis; inhibit, to some extent, tumor growth; modulate protein methylation in the target cells; and/or relieve, to some extent, one or more of the symptoms associated with the cancer. To the extent the administered compound or composition prevents growth and/or kills existing cancer cells, it may be cytostatic and/or cytotoxic. [0133] The term "container" means any receptacle and closure therefore suitable for storing, shipping, dispensing, and/or handling a pharmaceutical product.
[0134] The term "insert" means information accompanying a pharmaceutical product that provides a description of how to administer the product, along with the safety and efficacy data required to allow the physician, pharmacist, and patient to make an informed decision regarding use of the product. The package insert generally is regarded as the "label" for a pharmaceutical product.
[0135] The term "disease" or "condition" or "disorder" denotes disturbances and/or anomalies that as a rule are regarded as being pathological conditions or functions, and that can manifest themselves in the form of particular signs, symptoms, and/or malfunctions. As demonstrated below, Compounds of the Disclosure inhibit SMYD proteins, such as SMYD3 and SMYD2 and can be used in treating diseases and conditions such as proliferative diseases, wherein inhibition of SMYD proteins, such as SMYD3 and SMYD2 provides a benefit.
[0136] In some embodiments, the Compounds of the Disclosure can be used to treat a "SMYD protein mediated disorder" (e.g., a SMYD3-mediated disorder or a SMYD2-mediated disorder). A SMYD protein mediated disorder is any pathological condition in which a SMYD protein is know to play a role. In some embodiments, a SMYD-mediated disorder is a proliferative disease.
[0137] In some embodiments inhibiting SMYD proteins, such as SMYD3 and SMYD2, is the inhibition of the activity of one or more activities of SMYD proteins such as SMYD3 and SMYD2. In some embodiments, the activity of the SMYD proteins such as SMYD3 and SMYD2 is the ability of the SMYD protein such as SMYD3 or SMYD2 to transfer a methyl group to a target protein (e.g., histone). It should be appreciated that the activity of the one or more SMYD proteins such as SMYD3 and SMYD2 may be inhibited in vitro or in vivo. Ex nhibition of the activity one or more SMYD proteins such as SMYD lude at least 10% inhibition, at least 20% inhibition, at least 30% inh inhibition, at least 50% inhibition, at least 60% inhibition, at least 70% inhibition, at least 80% inhibition, at least 90% inhibition, and up to 100% inhibition.
[0138] The SMYD (SET and MYND domain) family of lysine methyltransferases (KMTs) plays pivotal roles in various cellular processes, including gene expression regulation and DNA damage response. The family of human SMYD proteins consists of SMYD1, SMYD2, SMYD3, SMYD4 and SMYD5. SMYD1, SMYD2, and SMYD3 share a high degree of sequence homology and, with the exception of SMYD5, human SMYD proteins harbor at least one C-terminal tetratrico peptide repeat (TPR) domain. (See e.g., Abu-Farha et al. J Mol Cell Biol (2011) 3 (5) 301-308). The SMYD proteins have been found to be linked to various cancers (See e.g., Hamamoto et al. Nat Cell. Biol. 2004, 6: 731-740), Hu et al. Canncer Research 2009, 4067-4072, and Komatsu et al. Carcinogenesis 2009, 301139-1146.)
[0139] SMYD3 is a protein methyltransferase found to be expressed at high levels in a number of different cancers (Hamamoto, R., et al., Nat. Cell Biol., 6(8):731-40 (2004)). SMYD3 likely plays a role in the regulation of gene transcription and signal transduction pathways critical for survival of breast, liver, prostate and lung cancer cell lines (Hamamoto, R., et al., Nat. Cell Biol., 6(8):731-40 (2004); Hamamoto, R., et al., Cancer Sci., 97(2):113-8 (2006); Van Aller, G.S., et al., Epigenetics, 7(4):340-3 (2012); Liu, C., et al., J. Natl. Cancer Inst., 105(22):1719-28 (2013); Mazur, P.K., et al., Nature, 510(7504):283-7 (2014)).
[0140] Genetic knockdown of SMYD3 leads to a decrease in proliferation of a variety of cancer cell lines (Hamamoto, R., et al., Nat. Cell Biol., 6(8):731-40 (2004); Hamamoto, R., et al., Cancer Sci., 97(2):113-8 (2006); Van Aller, G.S., et al., Epigenetics, 7(4):340- 3 (2012); Liu, C., et al., J. Natl. Cancer Inst., 105(22):1719-28 (2013); Mazur, P.K., et al., Nature, 510(7504):283-7 (2014)). Several studies employing RNAi-based technologies have shown that ablation of SMYD3 in hepatocellular carcinoma cell lines greatly reduces cell viability and that its pro-survival role is dependent on its catalytic activity (Hamamoto, R., et al., Nat. Cell Biol., 6(8):731-40 (2004); Van Aller, G.S., et al., Epigenetics, 7(4):340-3 (2012)). Moreover, SMYD3 has also been shown to be a critical mediator of transformation resulting from gain of function mutations in the oncogene, KRAS for both pancreatic and l a in mouse models. The dependence of KRAS on SMYD3 was also dent on its catalytic activity (Mazur, P.K., et al., Nature, 510(7504):2 YD3 function has also been implicated in colerectal cancers and RNAi mediated knockdown of SMYD3 has been shown to impair colerectal cell proliferation. (Peserico et al., Cell Physiol. 2015 Feb 28. doi: 10.1002/jcp.24975. [Epub ahead of print]).
[0141] Furthermore, SMYD3 function has also been shown to play a role in immunology and development. For instance, de Almeida reported that SMYD3 plays a role in generation of inducible regulatory T cells (iTreg) cells. In a mouse model of respiratory syncytial virus (RSV) infection, a model in which iTreg cells have a critical role in regulating lung pathogenesis, SMYD3-/- mice demonstrated exacerbation of RSV- induced disease related to enhanced proinflammatory responses and worsened pathogenesis within the lung (de Almeida et al. Mucosal Immunol. 2015 Feb 11. doi: 10.1038/mi.2015.4. [Epub ahead of print]). In addition, as to development, Proserpio et al. have shown the importance of SMYD3 in the regulation of skeletal muscle atrophy (Proserpio et al. Genes Dev. 2013 Jun 1;27(11):1299-312), while Fujii et al. have elucidated the role of SMYD3 in cardiac and skeletal muscle development (Fujii et al. PLoS One. 2011;6(8):e23491).
[0142] SMYD2 (SET and MYND domain-containing protein 2) was first characterized as protein that is a member of a sub-family of SET domain containing proteins which catalyze the site-specific transfer of methyl groups onto substrate proteins. SMYD2 was initially shown to have methyltransferase activity towards lysine 36 on histone H3 (H3K36) but has subsequently been shown to have both histone and non-histone methyltrasferase activity.
[0143] SMYD2 has been implicated in the pathogenesis of multiple cancers. It has been shown to be over-expressed, compared to matched normal samples, in tumors of the breast, cervix, colon, kidney, liver, head and neck, skin, pancreas, ovary, esophagus and prostate, as well as hematologic malignancies such as AML, B- and T-ALL, CLL and MCL, suggesting a role for SMYD2 in the biology of these cancers. More specifically, studies using genetic knock-down of SMYD2 have demonstrated anti-proliferative effects in esophageal squamous cell carcinoma (ESCC), bladder carcinoma and cervical carcinoma cell lines. (See e.g., Komatsu et al., Carcinogenesis 2009, 30, 1139, and Cho et al., Neoplasia. 2012 Jun;14(6):476 86) Moreover, high expression of SMYD2 has been shown to be a poor progn ESCC and pediatric ALL. (See e.g., Komatsu et al. Br J Cancer. 201 7-64, and Sakamoto et al., Leuk Res. 2014 Apr;38(4):496-502). Rec ., have shown that a small molecule inhibitor of SMYD2 (LLY-507) inhibited the proliferation of several esophageal, liver and breast cancer cell lines in a dose-dependent manner. (Nguyen et al. J Biol Chem. 2015 Mar 30. pii: jbc.M114.626861. [Epub ahead of print]).
[0144] SMYD2 has also been implicated in immunology. For instance, Xu et al. have shown that SMYD2 is a negative regulator of macrophage activation by suppressing Interleukin-6 and TNF-alpha production. (Xu et al., J Biol Chem. 2015 Feb 27;290(9):5414-23).
[0145] In one aspect, the present disclosure provides a method of treating cancer in a patient comprising administering a therapeutically effective amount of a Compound of the Disclosure. While not being limited to a specific mechanism, in some embodiemtns, Compounds of the Disclorure can treat cancer by inhibiting SMYD proteins, such as SMYD3 and SMYD2. Examples of treatable cancers include, but are not limited to, adrenal cancer, acinic cell carcinoma, acoustic neuroma, acral lentigious melanoma, acrospiroma, acute eosinophilic leukemia, acute erythroid leukemia, acute lymphoblastic leukemia, acute megakaryoblastic leukemia, acute monocytic leukemia, acute promyelocytic leukemia, adenocarcinoma, adenoid cystic carcinoma, adenoma, adenomatoid odontogenic tumor, adenosquamous carcinoma, adipose tissue neoplasm, adrenocortical carcinoma, adult T-cell leukemia/lymphoma, aggressive NK-cell leukemia, AIDS-related lymphoma, alveolar rhabdomyosarcoma, alveolar soft part sarcoma, ameloblastic fibroma, anaplastic large cell lymphoma, anaplastic thyroid cancer, angioimmunoblastic T-cell lymphoma, angiomyolipoma, angiosarcoma, astrocytoma, atypical teratoid rhabdoid tumor, B-cell chronic lymphocytic leukemia, B- cell prolymphocytic leukemia, B-cell lymphoma, basal cell carcinoma, biliary tract cancer, bladder cancer, blastoma, bone cancer, Brenner tumor, Brown tumor, Burkitt's lymphoma, breast cancer, brain cancer, carcinoma, carcinoma in situ, carcinosarcoma, cartilage tumor, cementoma, myeloid sarcoma, chondroma, chordoma, choriocarcinoma, choroid plexus papilloma, clear-cell sarcoma of the kidney, craniopharyngioma, cutaneous T-cell lymphoma, cervical cancer, colorectal cancer, Degos disease, desmoplastic small round cell tumor, diffuse large B-cell lymphoma, dysembryoplastic neuroepithelial tumor, dysgerminoma embryonal carcinoma, endocrine gland neoplasm, endodermal sinus tumor, entero -cell lymphoma, esophageal cancer, fetus in fetu, fibroma, fibrosa ymphoma, follicular thyroid cancer, ganglioneuroma, gastrointestina tumor, gestational choriocarcinoma, giant cell fibroblastoma, giant cell tumor of the bone, glial tumor, glioblastoma multiforme, glioma, gliomatosis cerebri, glucagonoma, gonadoblastoma, granulosa cell tumor, gynandroblastoma, gallbladder cancer, gastric cancer, hairy cell leukemia, hemangioblastoma, head and neck cancer, hemangiopericytoma, hematological malignancy, hepatoblastoma, hepatosplenic T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, invasive lobular carcinoma, intestinal cancer, kidney cancer, laryngeal cancer, lentigo maligna, lethal midline carcinoma, leukemia, leydig cell tumor, liposarcoma, lung cancer, lymphangioma, lymphangiosarcoma, lymphoepithelioma, lymphoma, acute lymphocytic leukemia, acute myelogeous leukemia, chronic lymphocytic leukemia, liver cancer, small cell lung cancer, non-small cell lung cancer, MALT lymphoma, malignant fibrous histiocytoma, malignant peripheral nerve sheath tumor, malignant triton tumor, mantle cell lymphoma, marginal zone B-cell lymphoma, mast cell leukemia, mediastinal germ cell tumor, medullary carcinoma of the breast, medullary thyroid cancer, medulloblastoma, melanoma, meningioma, merkel cell cancer, mesothelioma, metastatic urothelial carcinoma, mixed Mullerian tumor, mucinous tumor, multiple myeloma, muscle tissue neoplasm, mycosis fungoides, myxoid liposarcoma, myxoma, myxosarcoma, nasopharyngeal carcinoma, neurinoma, neuroblastoma, neurofibroma, neuroma, nodular melanoma, ocular cancer, oligoastrocytoma, oligodendroglioma, oncocytoma, optic nerve sheath meningioma, optic nerve tumor, oral cancer, osteosarcoma, ovarian cancer, Pancoast tumor, papillary thyroid cancer, paraganglioma, pinealoblastoma, pineocytoma, pituicytoma, pituitary adenoma, pituitary tumor, plasmacytoma, polyembryoma, precursor T-lymphoblastic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, preimary peritoneal cancer, prostate cancer, pancreatic cancer, pharyngeal cancer, pseudomyxoma periotonei, renal cell carcinoma, renal medullary carcinoma, retinoblastoma, rhabdomyoma, rhabdomyosarcoma, Richter's transformation, rectal cancer, sarcoma, Schwannomatosis, seminoma, Sertoli cell tumor, sex cord-gonadal stromal tumor, signet ring cell carcinoma, skin cancer, small blue round cell tumors, small cell carcinoma, soft tissue sarcoma, somatostatinoma, soot wart, spinal tumor, splenic marginal zone lymphoma, squamous cell carcinoma, synovial sarcoma Sezary's disease, small intestine cancer, squamous carcinoma, stomach cancer, T-ce ular cancer, thecoma, thyroid cancer, transitional cell carcinoma, thro cancer, urogenital cancer, urothelial carcinoma, uveal melanoma, rrucous carcinoma, visual pathway glioma, vulvar cancer, vaginal cancer, Waldenstrom's macroglobulinemia, Warthin's tumor, and Wilms' tumor.
[0146] In another embodiment, the cancer is breast, cervix, colon, kidney, liver, head and neck, skin, pancreas, ovary, esophagus, or prostate cancer. [0147] In another embodiment, the cancer is a hematologic malignancy such as acute myeloid leukemia (AML), B- and T-acute lymphoblastic leukemia (ALL), chronic lymphocytic leukemia (CLL), or mantle cell lymphoma (MCL).
[0148] In another embodiment, the cancer is esophageal squamous cell carcinoma (ESCC), bladder carcinoma, or cervical carcinoma.
[0149] In another embodiment, the cancer is a leukemia, for example a leukemia selected from acute monocytic leukemia, acute myelogenous leukemia, chronic myelogenous leukemia, chronic lymphocytic leukemia and mixed lineage leukemia (MLL). In another embodiment the cancer is NUT-midline carcinoma. In another embodiment the cancer is multiple myeloma. In another embodiment the cancer is a lung cancer such as small cell lung cancer (SCLC). In another embodiment the cancer is a neuroblastoma. In another embodiment the cancer is Burkitt's lymphoma. In another embodiment the cancer is cervical cancer. In another embodiment the cancer is esophageal cancer. In another embodiment the cancer is ovarian cancer. In another embodiment the cancer is colorectal cancer. In another embodiment, the cancer is prostate cancer. In another embodiment, the cancer is breast cancer.
[0150] In another embodiment, the present disclosure provides a therapeutic method of modulating protein methylation, gene expression, cell proliferation, cell differentiation and/or apoptosis in vivo in the cancers mentioned above by administering a therapeutically effective amount of a Compound of the Disclosure to a subject in need of such therapy.
[0151] Compounds of the Disclosure can be administered to a mammal in the form of a raw chemical without any other components present. Compounds of the Disclosure can also be administered to a mammal as part of a pharmaceutical composition containing the compound combined with a suitable pharmaceutically acceptable carrier. Such a carrier can be selected from pharmaceu excipients and auxiliaries. The term "pharmaceutically acceptable armaceutically acceptable vehicle" encompasses any of the stand al carriers, solvents, surfactants, or vehicles. Suitable pharmaceutically acceptable vehicles include aqueous vehicles and nonaqueous vehicles. Standard pharmaceutical carriers and their formulations are described in Remington's Pharmaceutical Sciences, Mack Publishing Co., Easton, PA, 19th ed. 1995. [0152] Pharmaceutical compositions within the scope of the present disclosure include all compositions where a Compound of the Disclosure is combined with one or more pharmaceutically acceptable carriers. In one embodiment, the Compound of the Disclosure is present in the composition in an amount that is effective to achieve its intended therapeutic purpose. While individual needs may vary, a determination of optimal ranges of effective amounts of each compound is within the skill of the art. Typically, a Compound of the Disclosure can be administered to a mammal, e.g., a human, orally at a dose of from about 0.0025 to about 1500 mg per kg body weight of the mammal, or an equivalent amount of a pharmaceutically acceptable salt or solvate thereof, per day to treat the particular disorder. A useful oral dose of a Compound of the Disclosure administered to a mammal is from about 0.0025 to about 50 mg per kg body weight of the mammal, or an equivalent amount of the pharmaceutically acceptable salt or solvate thereof. For intramuscular injection, the dose is typically about one-half of the oral dose.
[0153] A unit oral dose may comprise from about 0.01 mg to about 1 g of the Compound of the Disclosure, e.g., about 0.01 mg to about 500 mg, about 0.01 mg to about 250 mg, about 0.01 mg to about 100 mg, 0.01 mg to about 50 mg, e.g., about 0.1 mg to about 10 mg, of the compound. The unit dose can be administered one or more times daily, e.g., as one or more tablets or capsules, each containing from about 0.01 mg to about 1 g of the compound, or an equivalent amount of a pharmaceutically acceptable salt or solvate thereof.
[0154] A pharmaceutical composition of the present disclosure can be administered to any patient that may experience the beneficial effects of a Compound of the Disclosure. Foremost among such patients are mammals, e.g., humans and companion animals, although the disclosure is not intended to be so limited. In one embodiment, the patient is a human.
[0155] A pharmaceutical compo nt disclosure can be administered by any means that achieves its inten xample, administration can be by the oral, parenteral, subcutaneous, intravenous, intramuscular, intraperitoneal, transdermal, intranasal, transmucosal, rectal, intravaginal or buccal route, or by inhalation. The dosage administered and route of administration will vary, depending upon the circumstances of the particular subject, and taking into account such factors as age, gender, health, and weight of the recipient, condition or disorder to be treated, kind of concurrent treatment, if any, frequency of treatment, and the nature of the effect desired.
[0156] In one embodiment, a pharmaceutical composition of the present disclosure can be administered orally. In another embodiment, a pharmaceutical composition of the present disclosure can be administered orally and is formulated into tablets, dragees, capsules, or an oral liquid preparation. In one embodiment, the oral formulation comprises extruded multiparticulates comprising the Compound of the Disclosure.
[0157] Alternatively, a pharmaceutical composition of the present disclosure can be administered rectally, and is formulated in suppositories.
[0158] Alternatively, a pharmaceutical composition of the present disclosure can be administered by injection.
[0159] Alternatively, a pharmaceutical composition of the present disclosure can be administered transdermally.
[0160] Alternatively, a pharmaceutical composition of the present disclosure can be administered by inhalation or by intranasal or transmucosal administration.
[0161] Alternatively, a pharmaceutical composition of the present disclosure can be administered by the intravaginal route.
[0162] A pharmaceutical composition of the present disclosure can contain from about 0.01 to 99 percent by weight, e.g., from about 0.25 to 75 percent by weight, of a Compound of the Disclosure, e.g., about 1%, about 5%, about 10%, about 15%, about 20%, about 25%, about 30%, about 35%, about 40%, about 45%, about 50%, about 55%, about 60%, about 65%, about 70%, or about 75% by weight of a Compound of the Disclosure.
[0163] A pharmaceutical composition of the present disclosure is manufactured in a manner which itself will be known in view of the instant disclosure, for example, by means of conventional mixing ee-making, dissolving, extrusion, or lyophilizing processes. Thus, p positions for oral use can be obtained by combining the active com excipients, optionally grinding the resulting mixture and processing the mixture of granules, after adding suitable auxiliaries, if desired or necessary, to obtain tablets or dragee cores.
[0164] Suitable excipients include fillers such as saccharides (for example, lactose, sucrose, mannitol or sorbitol), cellulose preparations, calcium phosphates (for example, tricalcium phosphate or calcium hydrogen phosphate), as well as binders such as starch paste (using, for example, maize starch, wheat starch, rice starch, or potato starch), gelatin, tragacanth, methyl cellulose, hydroxypropylmethylcellulose, sodium carboxymethylcellulose, and/or polyvinyl pyrrolidone. If desired, one or more disintegrating agents can be added, such as the above-mentioned starches and also carboxymethyl-starch, cross-linked polyvinyl pyrrolidone, agar, or alginic acid or a salt thereof, such as sodium alginate.
[0165] Auxiliaries are typically flow-regulating agents and lubricants such as, for example, silica, talc, stearic acid or salts thereof (e.g., magnesium stearate or calcium stearate), and polyethylene glycol. Dragee cores are provided with suitable coatings that are resistant to gastric juices. For this purpose, concentrated saccharide solutions can be used, which may optionally contain gum arabic, talc, polyvinyl pyrrolidone, polyethylene glycol and/or titanium dioxide, lacquer solutions and suitable organic solvents or solvent mixtures. In order to produce coatings resistant to gastric juices, solutions of suitable cellulose preparations such as acetylcellulose phthalate or hydroxypropylmethyl-cellulose phthalate can be used. Dye stuffs or pigments can be added to the tablets or dragee coatings, for example, for identification or in order to characterize combinations of active compound doses.
[0166] Examples of other pharmaceutical preparations that can be used orally include push-fit capsules made of gelatin, or soft, sealed capsules made of gelatin and a plasticizer such as glycerol or sorbitol. The push-fit capsules can contain a compound in the form of granules, which can be mixed with fillers such as lactose, binders such as starches, and/or lubricants such as talc or magnesium stearate and, optionally, stabilizers, or in the form of extruded multiparticulates. In soft capsules, the active compounds are preferably dissolved or suspended in suitable liquids, such as fatty oils or liquid paraffin. In addition, stabilizers can be added
[0167] Possible pharmaceutica rectal administration include, for example, suppositories, which co ion of one or more active compounds with a suppository base. Sui bases include natural and synthetic triglycerides, and paraffin hydrocarbons, among others. It is also possible to use gelatin rectal capsules consisting of a combination of active compound with a base material such as, for example, a liquid triglyceride, polyethylene glycol, or paraffin hydrocarbon.
[0168] Suitable formulations for parenteral administration include aqueous solutions of the active compound in a water-soluble form such as, for example, a water-soluble salt, alkaline solution, or acidic solution. Alternatively, a suspension of the active compound can be prepared as an oily suspension. Suitable lipophilic solvents or vehicles for such as suspension may include fatty oils (for example, sesame oil), synthetic fatty acid esters (for example, ethyl oleate), triglycerides, or a polyethylene glycol such as polyethylene glycol-400 (PEG-400). An aqueous suspension may contain one or more substances to increase the viscosity of the suspension, including, for example, sodium carboxymethyl cellulose, sorbitol, and/or dextran. The suspension may optionally contain stabilizers.
[0169] In another embodiment, the present disclosure provides kits which comprise a Compound of the Disclosure (or a composition comprising a Compound of the Disclosure) packaged in a manner that facilitates their use to practice methods of the present disclosure. In one embodiment, the kit includes a Compound of the Disclosure (or a composition comprising a Compound of the Disclosure) packaged in a container, such as a sealed bottle or vessel, with a label affixed to the container or included in the kit that describes use of the compound or composition to practice the method of the disclosure. In one embodiment, the compound or composition is packaged in a unit dosage form. The kit further can include a device suitable for administering the composition according to the intended route of administration. General Synthesis of Compounds
[0170] Compounds of the Disclosure are prepared using methods known to those skilled in the art in view of this disclosure, or by the illustrative methods shown in the General Schemes below. In the General Schemes, R1, R2a, R3a, R4a, A, Y, and Z of Formulae A-D are as defined in connection with Formula I, unless otherwise indicated. In any of the General Schemes, suitable protecting can be employed in the synthesis, for example, when Z is (amino)alkyl or any other group that may group that may require protection. (See, Wuts, P. G. M.; Greene, T tective Groups in Organic Synthesis", 4th Ed., J. Wiley & Sons, NY, 2
1
Figure imgf000071_0001
[0171] Compound A is converted to compound B (i.e, a compound having Formula I, wherein R2b, R3b, R4b, and R5 are each hydrogen, and X is -S(=O)2-) by coupling with a suitable sulfonyl chloride (Z-SO2Cl) in the presence of a suitable base such as TEA or DIPEA in a suitable solvent such as dichloromethane, acetonitrile, or DMF. General Scheme 2
Figure imgf000072_0001
[0172] Compound A is converted to compound C (i.e, a compound having Formula I, wherein R2b, R3b, R4b, and R5 are each hydrogen, and X is -C(=O)-) by coupling with a suitable acide chloride (Z-COCl) in the presence of a suitable base such as TEA or DIPEA in a suitable solvent such as dichloromethane, acetonitrile, or DMF, or by coupling with a suitable carboxylic acid (Z-CO2H) in the presence of a suitable coupling reagent such as HATU and a suitable base such as TEA or DIPEA in a suitable solvent such as dichloromethane, acetonitrile, or DMF. General Scheme 3
Figure imgf000072_0002
[0173] Compound A is convert (i.e, a compound having Formula I, wherein R2b, R3b, R4b, and R5 are d X is -C(=O)C(R7)(H)-) by coupling with a suitable carboxylic acid (Z-C(H)R7-CO2H) in the presence of a suitable coupling reagent such as HATU and a suitable base such as TEA or DIPEA in a suitable solvent such as dichloromethane, acetonitrile, or DMF. EXAMPLES General Synthetic Methods [0174] General methods and experimental procedures for preparing and characterizing Compounds of the Disclosure are set forth in the general schemes above and the examples below. Wherever needed, reactions were heated using conventional hotplate apparatus or heating mantle or microwave irradiation equipment. Reactions were conducted with or without stirring, under atmospheric or elevated pressure in either open or closed vessels. Reaction progress was monitored using conventional techniques such as TLC, HPLC, UPLC, or LCMS using instrumentation and methods described below. Reactions were quenched and crude compounds isolated using conventional methods as described in the specific examples provided. Solvent removal was carried out with or without heating, under atmospheric or reduced pressure, using either a rotary or centrifugal evaporator.
[0175] Compound purification was carried out as needed using a variety of traditional methods including, but not limited to, preparative chromatography under acidic, neutral, or basic conditions using either normal phase or reverse phase HPLC or flash columns or Prep-TLC plates. Compound purity and mass confirmations were conducted using standard HPLC and / or UPLC and / or MS spectrometers and / or LCMS and / or GC equipment (i.e., including, but not limited to the following instrumentation: Waters Alliance 2695 with 2996 PDA detector connected with ZQ detector and ESI source; Shimadzu LDMS-2020; Waters Acquity H Class with PDA detector connected with SQ detector and ESI source; Agilent 1100 Series with PDA detector; Waters Alliance 2695 with 2998 PDA detector; AB SCIEX API 2000 with ESI source; Agilent 7890 GC). Exemplified compounds were dissolved in either MeOH or MeCN to a concentration of approximately 1 mg/mL and a n of 0.5-10 μL into an appropriate LCMS system.
[0176] Compound structure co carried out using standard 300 or 400 MHz NMR spectrometers with nOe s conducted whenever necessary.
[0177] The following abbreviations may be used herein:
Figure imgf000073_0001
Figure imgf000074_0001
EXAMPLE 1
Synthesis of (±)-trans-N-(1-cyclopropyl-4-methylpyrrolidin-3-yl)-2-oxoindoline-5- carboxamide (Cpd. No. 5)
and
(±)-cis- N-(1-cyclopropyl-4-methylpyrrolidin-3-yl)-2-oxoindoline-5-carboxamide
(Cpd. No. 6)
Figure imgf000075_0001
[0178] To a solution of 2-oxoindoline-5-carboxylic acid (200 mg, 1.12 mmol) in DMF (3 mL) was added HATU (644 mg, 1.69 mmol) and the reaction mixture was stirred for 30 min at room temperature. After being cooled to 0 °C 1-cyclopropyl-4-methylpyrrolidin- 3-amine (174 mg, 1.24 mmol) and DIPEA (0.29 mL, 1.69 mmol) were added successively to the reaction mixture. The reaction was further stirred at room temperature for 2 hrs. Completion of the reaction was confirmed by TLC. After completion, water (30 mL) was added to reaction mixture and product was extracted with ethyl acetate (3 x 25 mL). The combined organic layer was washed with brine, and dried over sodium sulfate and concentrated under vacuum to get crude product which was purified by column chromatography. The desire compound started eluting at 4% methanol in DCM. Evaporation of the fractions afforded 70 mg of pure racemic compound as a mixture of cis : trans isomer. The cis and trans isomers were separated out using reverse-phase prep. HPLC using 0.1% TFA in ACN and 0.1% TFA in water as mobile phase. Lyophilization of pure fractions afforded the pure title compounds. The yields of the isomers were (18 mg (5.33%) and 23 mg (6.81%) a and LCMS of the isomers were: 1H NMR (400 MHz, MeOD): į 7.7 97-6.95 (m, 1H), 4.15 (dd, J = 6 Hz, 1H), 3.60-3.58 (m, 1H), 3.51 (m , 1H), 3.18 (dd, J = 6 Hz, 1H), 2.82- 2.78 (m, 1H), 2.40-2.36 (t, J = 8.4 Hz, 1H), 2.25 2.21 (m, 1H), 1.21-1.20 (m, 3H), 0.53- 0.49 (m, 4H). LCMS (m/z): 300.25 [M+H]+. 1H NMR (400 MHz, MeOD): į 7.78-7.76 (m, 2H), 6.98-6.96 (m, 1H), 4.68 (dd, J=5.2 Hz, 1H), 3.61-3.59 (m, 1H), 3.24 (dd, J=5.2 Hz, 1H), 3.12 (dd, J=7.6 Hz, 1H), 2.72 (dd, J=6.4 Hz, 1H), 2.59-2.55 (m, 1H), 2.45 (t, J= 9.2 Hz, 1H), 1.86-1.84 (m, 1H), 0.97-0.95 (m, 3H), 0.54-0.48 (m, 4H). LCMS (m/z): 300.25 [M+H]+. EXAMPLE 2
SMYD3 Biochemical Assay General Materials
[0179] S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), Tris, Tween20, dimethylsulfoxide (DMSO), bovine skin gelatin (BSG), and Tris(2- carboxyethyl)phosphine hydrochloride solution (TCEP) were purchased from Sigma- Aldrich at the highest level of purity possible. 3H-SAM was purchase from American Radiolabeled Chemicals with a specific activity of 80 Ci/mmol. 384-well opaque white OptiPlates and SPA beads (Perkin Elmer, catalog # RPNQ0013) were purchased from PerkinElmer. Substrates
[0180] N-terminally GST-tagged MEKK2 (MAP3K2) protein corresponding to reference sequence AAF63496.3 was purchased from Life Technologies (catalog # PV4010). This protein was expressed in High Five insect cells and purified to >85 % purity. Protein identity was confirmed by MS/MS analysis after proteolytic digestion. The protein sequence used was:
MAPILGYWKIKGLVQPTRLLLEYLEEKYEEHLYERDEGDKWRNK KFELGLEFPNLPYYIDGDVKLTQSMAIIRYIADKHNMLGGCPKERA EISMLEGAVLDIRYGVSRIAYSKDFETLKVDFLSKLPEMLKMFEDR LCHKTYLNGDHVTHPDFMLYDALDVVLYMDPMCLDAFPKLVCF KKRIEAIPQIDKYLKSS ATFGGGDHPPKSDLVPRH NQTSLYKKAGTMDDQ VLHKASRPALSLQETRKA KSSSPKKQNDVRVKF RPVKLEDLRSKAKIAFGQS MDLHYTNNELVIPLTTQDDLDKALELLDRSIHMKSLKILLVINGST QATNLEPLPSLEDLDNTVFGAERKKRLSIIGPTSRDRSSPPPGYIPDE LHQVARNGSFTSINSEGEFIPESMEQMLDPLSLSSPENSGSGSCPSL DSPLDGESYPKSRMPRAQSYPDNHQEFSDYDNPIFEKFGKGGTYPR RYHVSYHHQEYNDGRKTFPRARRTQGNQLTSPVSFSPTDHSLSTSS GSSIFTPEYDDSRIRRRGSDIDNPTLTVMDISPPSRSPRAPTNWRLG
KLLGQGAFGRVYLCYDVDTGRELAVKQVQFDPDSPETSKEVNAL ECEIQLLKNLLHERIVQYYGCLRDPQEKTLSIFMEYMPGGSIKDQL KAYGALTENVTRKYTRQILEGVHYLHSNMIVHRDIKGANILRDST GNVKLGDFGASKRLQTICLSGTGMKSVTGTPYWMSPEVISGQGYG RKADIWSVACTVVEMLTEKPPWAEFEAMAAIFKIATQPTNPKLPP HVSDYTRDFLKRIFVEAKLRPSADELLRHMFVHYH.
(SEQ ID No. 1). Molecular Biology
[0181] Full-length human SMYD3 isoform 1 (BAB86333) was inserted into a modified pET21b plasmid containing a His6 tag and TEV and SUMO cleavage sites. Because two common variants of SMYD3 exist in the population, site directed mutagenesis was subsequently performed to change amino acid 13 from an asparagine to a lysine, resulting in plasmid pEPZ533. A lysine at position 13 conforms to the more commonly occurring sequence (NP_001161212). Protein Expression
[0182] E. coli (BL21 codonplus RIL strain, Stratagene) were transformed with plasmid pEPZ553 by mixing competent cells and plasmid DNA and incubating on ice for 30 minutes followed by heat shock at 42°C for 1 minute and cooling on ice for 2 minutes. Transformed cells were grown and selected on LB agar with 100 μg/mL ampicillin and 17 μg/mL chloramphenicol at 37°C overnight. A single clone was used to inoculate 200 mL of LB medium with 100 μg/mL ampicillin and 17 μg/mL chloramphenicol and incubated at 37°C on an orbital h k t 180 Once in log growth, the culture was diluted 1:100 into 2 L of LB m ntil OD600 was about 0.3 after which the culture was incubated at 15 Once OD600 reached about 0.4, IPTG was added to a final concentratio e cells were grown overnight at 15°C and 160 rpm. Cells were harvested by centrifugation at 8000 rpm, for 4 minutes at 4°C and stored at -80 oC for purification.
Protein Purification
[0183] Expressed full-length human His-tagged SMYD3 protein was purified from cell paste by Nickel affinity chromatography after equilibration of the resin with Buffer A (25 mM Tris, 200 mM NaCl, 5% glycerol, 5 mM E-mercaptoethanol, pH7.8). The column was washed with Buffer B (Buffer A plus 20 mM imidazole) and His-tagged SMYD3 was eluted with Buffer C (Buffer A plus 300 mM imidazole). The His tag, TEV and SUMO cleavage sites were removed generating native SMYD3 by addition of ULP1 protein at a ratio of 1:200 (ULP1:SMYD3). Imidazole was removed by dialysis overnight in Buffer A. The dialyzed solution was applied to a second Nickel column and the native SMYD3 protein was collected from the column flow-through. The flow- through was dialyzed in Buffer D (25 mM Tris, 5% glycerol, 5 mM E-mercaptoethanol, 50 mM NaCl, pH7.8) and ULP1 was removed using a Q sepharose fast flow column. SMYD3 was eluted in Buffer A and further purified using an S200 size-exclusion column equilibrated with Buffer A. SMYD3 was concentrated to 2 mg/mL with a final purity of 89%. Predicted Translation:
[0184] SMYD3 (Q9H7B4)
MEPLKVEKFATAKRGNGLRAVTPLRPGELLFRSDPLAYTVCKGSR GVVCDRCLLGKEKLMRCSQCRVAKYCSAKCQKKAWPDHKRECK CLKSCKPRYPPDSVRLLGRVVFKLMDGAPSESEKLYSFYDLESNIN KLTEDKKEGLRQLVMTFQHFMREEIQDASQLPPAFDLFEAFAKVIC NSFTICNAEMQEVGVGLYPSISLLNHSCDPNCSIVFNGPHLLLRAV RDIEVGEELTICYLDMLMTSEERRKQLRDQYCFECDCFRCQTQDK DADMLTGDEQVWKEVQESLKKIEELKAHWKWEQVLAMCQAIISS NSERLPDINIYQLKVLDCAMDACINLGLLEEALFYGTRTMEPYRIFF PGSHPVRGVQVMKVGKLQLHQGMFPQAMKNLRLAFDIMRVTHG REHSLIEDLILLLEECDANIRAS (SEQ ID No. 2). General Procedure for SMYD3 E MEKK2 protein substrate
[0185] The assays were all perfo onsisting of 25 mM Tris-Cl pH 8.0, 1 mM TCEP, 0.005% BSG, and 0.005% Tween 20, prepared on the day of use. Compounds in 100% DMSO (1ul) were spotted into a 384-well white opaque OptiPlate using a Bravo automated liquid handling platform outfitted with a 384-channel head (Agilent Technologies). DMSO (1ul) was added to Columns 11, 12, 23, 24, rows A-H for the maximum signal control and 1ul of SAH, a known product and inhibitor of SMYD3, was added to columns 11, 12, 23, 24, rows I-P for the minimum signal control. A cocktail (40ul) containing the SMYD3 enzyme was added by Multidrop Combi (Thermo-Fisher). The compounds were allowed to incubate with SMYD3 for 30 min at room temperature, then a cocktail (10ul) containing SAM and MEKK2 was added to initiate the reaction (final volume = 51ul). The final concentrations of the components were as follows: SMYD3 was 0.4 nM, 3H-SAM was 8 nM, MEKK2 was 12 nM, SAH in the minimum signal control wells was 1 mM, and the DMSO concentration was 2%. The assays were stopped by the addition of non-radiolabeled SAM (10ul) to a final concentration of 100 uM, which dilutes the 3H-SAM to a level where its incorporation into MEKK2 is no longer detectable. Radiolabeled MEKK2 was detected using a scintillation proximity assay (SPA). 10 uL of a 10 mg/mL solution of SPA beads in 0.5 M citric acid was added and the plates centrifuged at 600 rpm for 1 min to precipitate the radiolabeled MEKK2 onto the SPA beads. The plates were then read in a PerkinElmer TopCount plate reader to measure the quantity of 3H-labeled MEKK2 as disintegrations per minute (dpm) or alternatively, referred to as counts per minute (cpm). % inhibition calculation
Figure imgf000079_0001
[0186] Where dpm = disintegrations per minute, cmpd = signal in assay well, and min and max are the respective minimum and maximum signal controls. Four-parameter IC50 fit
Figure imgf000079_0002
[0187] Where top and bottom ar wed to float, but may be fixed at 100 or 0 respectively in a 3-paramet fficient normally allowed to float but may also be fixed at 1 in a 3-par % inhibition and X is the compound concentration. EXAMPLE 3
SMYD3 Cell Assay Trimethyl-MEKK2-In-Cell Western Assay
[0188] 293T/17 adherent cells were purchased from ATCC (American Type Culture Collection), Manassas, VA, USA. MEM/Glutamax medium, Optimem Reduced Serum medium, penicillin-streptomycin, 0.05% trypsin and 1x D-PBS were purchased from Life Technologies, Grand Island, NY, USA. PBS-10X was purchased from Ambion, Life Technologies, Grand Island, New York, USA. PBS with Tween 20 (PBST (10x)) was purchased from KPL, Gaithersburg, Maryland,USA. Tet System FBS- approved FBS US Source was purchased from Clontech, Mountain View, California, USA. Odyssey blocking buffer, 800CW goat anti-rabbit IgG (H+L) antibody, 680CW Goat anti-mouse IgG (H+L) and Licor Odyssey infrared scanner were purchased from Licor Biosciences, Lincoln, NE, USA. Tri-methyl-Lysine [A260]-MEKK2 antibody, MEKK2 and SMYD3 plasmids were made at Epizyme. Anti-flag monoclonal mouse antibody was purchased from Sigma, St. Louis, MO, USA. Methanol was purchased from VWR, Franklin, MA, USA. 10% Tween 20 was purchased from KPL, Inc., Gaithersburg, Maryland, USA. Fugene was purchased from Promega, Madison, WI, USA. The Biotek ELx405 was purchased from BioTek, Winooski, Vermont, USA. The multidrop combi was purchased from Thermo Scientific, Waltham, Massachusetts, USA.
[0189] 293T/17 adherent cells were maintained in growth medium (MEM/Glutamax medium supplemented with 10% v/v Tet System FBS and cultured at 37 °C under 5% CO2. Cell treatment, In Cell Western (ICW) for detection of trimethyl-lysine-MEKK2 and MEKK2.
[0190] 293T/17 cells were seede at a concentration of 33,333 cells per cm2 in 30 mL medium per T150 d at 37 °C under 5% CO2. Plasmids were prepared for delivery to cells by first mixing 1350 μL Opti-MEM with Fugene (81 μL) in a sterile Eppendorf and incubated for five minutes at room temperature (RT). MEKK2-flag (13.6 ug/T150) MEKK2 p3XFlag-CMV-14 with C-3XFlag and SMYD3 (0.151 ug/T150) SMYD3 p3XFlag-CMV-14 without C-3XFlag plasmids were aliquotted to a 1.7 mL sterile microfuge tube. The gene ID for MEKK2 and SMYD3 is NM_006609.3 and Q9H7B4, respectively. Entire volume of Opti-MEM/Fugene mixture was then added to a microfuge tube containing DNA plasmid, mixed and then incubated x 15 minutes at RT. The medium on the 293T/17 cells was refreshed, and the DNA/Fugene complex is added aseptically to each flask, rocked gently, and incubated at 37 C for 5 hours. Medium was then removed, and cells were washed once with PBS in the flask. Trypsin 0.05% (3mL) was added and cells incubated for three minutes. Room temperature MEM+10% Tet system FBS was added and cells were mixed gently, and counted using the Vi-cell. Cells were seeded at 100,000 cells/mL in 50 μL MEM/10%Tet FBS/Pen/Strep to a 384 well black/clear poly-D-lysine coated plate containing test agent diluted in DMSO. The final top concentration of test compound was 40 μM. The total concentration of DMSO did not exceed 0.2% (v/v). Plates were incubated x 30 minutes at RT in low-airflow area, followed by incubation at 37 °C under 5% CO2 for 24 hours. Medium was aspirated from all wells of assay plates prior to fixation and permeabilization with ice cold (-20 ºC) methanol (90 μL/well) for ten minutes. Plates were rinsed with PBS three times on BioTek ELx405. PBS was removed with a final aspiration, and Odyssey blocking buffer (50 μL/well) was added to each well and incubated for one hour at RT. Primary antibody solution was prepared (anti-trimethyl-MEKK2 at 1:600 dilution plus mouse anti-flag antibody at 1:10,000 dilution in diluent (Odyssey Blocking buffer + 0.1% Tween 20)) and 20 μL per well was dispensed using the Multidrop Combi. Assay plates were then sealed with foil, and incubated overnight at 4º C. Plates were washed five times with PBS-Tween (1X) on Biotek ELx405 and blotted on paper towel to remove excess reagent. Detection antibody solution (IRDye 800 CW goat anti-rabbit IgG diluted 1:400 in diluent (Odyssey Blocking buffer + 0.1% Tween 20), plus IRDye 680CW goat anti-mouse IgG at 1:500 in diluent (Odyssey Blocking buffer + 0.1% Tween 20) was added (20 μL/well) and incubated in dark for one hour at RT. Plat ed four times with PBS-T (1X) on ELx405. A final rinse with wa (115 μL/well x three washes on the ELx405). Plates were then ce own, on paper towel, at 200 x g to remove excess reagent. Plates were left to dry in dark for one hour. The Odyssey Imager was used to measure the integrated intensity of 700 and 800 wavelengths at resolution of 84 μm, medium quality, focus offset 4.0, 700 channel intensity = 3.5 to measure the MEKK2-flag signal, 800 channel intensity = 5 to measure the Trimethyl-MEKK2 signal of each well. Calculations:
[0191] First, the ratio for each well was determined by:
Figure imgf000082_0002
[0192] Each plate included fourteen control wells of DMSO only treatment (Minimum Inhibition) as well as fourteen control wells for maximum inhibition (Background). The average of the ratio values for each control type was calculated and used to determine the percent inhibition for each test well in the plate. Reference compound was serially diluted two-fold in DMSO for a total of nine test concentrations, beginning at 40 μM. Percent inhibition was calculated below . P
Figure imgf000082_0001
[0193] Non-linear regression curves were generated to calculate the IC50 and dose-response relationship using triplicate wells per concentration of compound. EXAMPLE 4
SMYD2 ASSAY General Materials
[0194] S-adenosylmethionine (SAM), S-adenosylhomocysteine (SAH), bicine, Tween20, dimethylsulfoxide (DMSO), bovine skin gelatin (BSG), and Tris(2- carboxyethyl)phosphine hydrochloride (TCEP) were purchased from Sigma-Aldrich at the highest level of purity possible. 3H-SAM was purchase from American Radiolabeled Chemicals with a specific activity of 80 Ci/mmol. 384-well streptavidin Flashplates were purchased from PerkinElmer. Substrates
[0195] Peptide was synthesized with a N-terminal linker-affinity tag motif and a C- terminal amide cap by 21st Century Biochemicals. The peptide was high high-perfomance liquid chromatography (HPLC) purified to greater than 95% purity and confirmed by liquid chromatography mass spectrometry (LC-MS). The sequence was ARTKQTARKSTGGKAPRKQLATKAARKSA(K-Biot)-amide. (SEQ ID NO: 3) Production of Recombinant SMYD2 Enzymes for Biochemical Enzyme Activity Assays
[0196] Full length SMYD2 (NP_064582.2) was cloned into a pFastbac-Htb-lic vector with an N-terminal His6 tag and FLAG tag, preceded by a TEV protease cleavage site. The protein was expressed in Sf9 insect cells. Cells were resuspended in lysis buffer (25 mM HEPES-NaOH, pH 7.5, 200 mM NaCl, 5% glycerol, and 5 mM ȕ-ME) and lysed by sonication. The protein was purified by Ni-NTA (Qiagen), followed by TEV cleavage to remove the His6 tag, subtractive Ni-NTA (Qiagen), and gel filtration chromatography using an S200 column (GE Healthcare). Purified protein was stored in 20 mM Tris-HCl, pH 8.0, 100 mM NaCl, and 1 mM TCEP. General Procedure for SMYD2 Enzyme Assays on Peptide Substrates
[0197] The assays were all performed in a buffer consisting of 20mM Bicine (pH=7.6), 1mM TCEP, 0.005% Bovine Skin Gelatin, and 0.002% Tween20, prepared on the day of use. Compounds in 100% DMSO (1ul) were spotted into a polypropylene 384-well V- bottom plates (Greiner) using a Platemate Plus outfitted with a 384-channel head (Thermo Scientific). DMSO (1ul) was added to Columns 11, 12, 23, 24, rows A-H for the maximum signal control and 1ul of SAH, a known product and inhibitor of SMYD2, was added to columns 11, 12, 23, 24, rows I-P for the minimum signal control. A cocktail (40ul) containing the SMYD2 enzyme was added by Multidrop Combi (Thermo-Fisher). The compounds were allowed to incubate with SMYD2 for 30 min at room temperature, then a cocktail (10ul) containing 3H-SAM and peptide was added to initiate the reaction (final volume = 51ul). The final concentrations of the components were as follows: SMYD2 was 1.5nM, 3H-SAM was 10nM, and peptide was 60nM, SAH in the minimum signal control wells was 1000uM and the DMSO concentration was 2%. The assays were stopped by the addition of no (10ul) to a final concentration of 600uM, which dilutes the 3H-S ere its incorporation into the peptide substrate is no longer detectable n in the 384-well polypropylene plate was then transferred to a 384-well Flashplate and the biotinylated peptides were allowed to bind to the streptavidin surface for at least 1 hour before being washed three times with 0.1%Tween20 in a Biotek ELx405 plate washer. The plates were then read in a PerkinElmer TopCount plate reader to measure the quantity of 3H-labeled peptide bound to the Flashplate surface, measured as disintegrations per minute (dpm) or alternatively, referred to as counts per minute (cpm). % inhibition calculation
Figure imgf000084_0001
[0198] Where dpm = disintegrations per minute, cmpd = signal in assay well, and min andmax are the respective minimum and maximum signal controls. Four-parameter IC50 fit
Figure imgf000084_0002
[0199] Where top and bottom are the normally allowed to float, but may be fixed at 100 or 0 respectively in a 3-parameter fit. The Hill Coefficient normally allowed to float but may also be fixed at 1 in a 3-parameter fit. I is the compound concentration.
[0200] SMYD2 biochemical assay data for representative Compounds of the Disclosure are presented in Tables 1, 3, and 4 in the column titled "SMYD2 Bicohem IC50 (μM)."
[0201] Having now fully described this invention, it will be understood by those of ordinary skill in the art that the same can be performed within a wide and equivalent range of conditions, formulations, and other parameters without affecting the scope of the invention or any embodiment thereof.
[0202] Other embodiments of the invention will be apparent to those skilled in the art from consideration of the specification and practice of the invention disclosed herein. It is intended that the specification considered as exemplary only, with a true scope and spirit of the inven d by the following claims.
[0203] All patents and publica are fully incorporated by reference herein in their entirety.

Claims

What is Claimed Is: 1. A compound having Formula I:
Figure imgf000085_0001
or a pharmaceutically acceptable salt or hydrate thereof, wherein: A is selected from the group consisting of 1,2,3-triazolyl, 1,2,4-triazolyl, 1-imidazolyl, 1-isoquinolinyl, 1-pyrazolyl, 2-(1,2,3,4-tetrahydroquinolinyl), 2-benzo[d]imidazolyl, 2-benzo[d]thiazolyl, 2-chromenyl-4-one, 2-furanyl, 2-imidazo[1,2-b]pyridazinyl, 2-imidazolyl, 2-indolyl, 2-naphthalenyl, 2-pyrazinyl, 2-pyridyl, 2-pyrimidinyl, 2-pyrrolidinyl, 2-pyrrolyl, 2-quinolinyl, 2-quinoxalinyl, 2-thiazolo[5,4-c]pyridinyl, 2-thiazolyl, 2-thiophenyl, 3-(1,2,3,4-tetrahydroisoquinoline), 3-(1,2,4-oxadiazolyl), 3-imidazo[1,2-a]pyrimidinyl, 3-indazolyl, 3-indolyl, 3-isothiazolyl, 3-pyrazolyl, 3-pyridazinyl, 3-pyridinyl-2-one, 3-pyridyl, 3-pyrrolo[3,2-b]pyridinyl, 3-quinolinyl, 4-(2,2-difluorobenzo[d][1,3]dioxolyl), 4-cyclohexanyl-1-amine, 4-imidazolyl, 4-indolinyl-2-one, 4-indolyl, 4-isothiazolyl, 4-oxazolyl, 4-piperidinyl, 4-pyrazolyl, 4-pyridyl, 4-quinolinyl, 5-(1,3-dihydro-2H-benzo[d]imidazolyl-2-one), 5-(1,3-dihydro- 2H-pyrrolo[2,3-b]pyridinyl-2-one), 5-(1,3-dihydro-2H-pyrrolo[2,3-c]pyridinyl-2-one), 5-(2,2-difluorobenzo[d][1,3]dioxolyl), 5-(2,4-dihydro-3H-1,2,4-triazolyl-3-one), 5-4H- furo[3,2-b]pyrrolyl, 5-ben zolyl, 5-benzo[d][1,3]dioxolyl, 5-benzo[d]oxazolyl-2(3H)-one, eptyl-2-ene, 5-indolinyl-2,3-dione, 5-indolinyl-2-one, 5-indolyl, ne, 5-isoxazolyl, 5-pyrazolo[3,4- c]pyridinyl, 5-pyrazolyl, 5-pyrimidinyl, 5-thiazolyl, 6-(1,2,3,4-tetrahydronaphthalenyl), 6-(3,4-dihydroquinolinyl-2(1H)-one), 6-(3,4-dihydroquinoxalinyl-2(1H)-one), 6-(4,5- dihydropyridazinyl-3(2H)-one), 6-benzo[b][1,4]oxazinyl-3-one, 6-benzo[d]imidazolyl, 6-benzo[d]oxazolyl-2(3H)-one, 6-benzo[d]thiazolyl, 6-chromenyl-2-one, 6-imidazo[2,1- b]thiazole, 6-indazolyl, 6-indolinyl-2-one, 6-indolyl, 6-isoquinolinyl, 6-quinolinyl, 6-quinoxalinyl, 6-quinoxalinyl-2(1H)-one, 7-(3,4-dihydroquinolinyl-2(1H)-one), 7-(3,4- dihydroquinoxalin-2(1H)-one), 7-benzo[b][1,4]oxazinyl-3-one, 7-indolinyl-2-one, 7-quinolinyl, 8-benzo[b][1,4]oxazinyl-3-one, cyclopropanyl, phenyl, 4-(prop-1-en-1-yl)- imidazole, 1-butanyl-imidazole, sec-butylcyclopropane, 2-(ethylsulfonyl)propanyl, 1-isobutylpyrrolidine, 4-pyridyl 1-oxide, and 5-benzo[c][1,2,5]oxadiazolyl 1-oxide, each of which is optionally substituted with one, two, or three substituents independently selected from the group consisting of halo, hydroxy, alkoxy, amino, alkylamino, dialkylamino, (amino)alkyl,(alkylamino)alkyl, (dialkylamino)alkyl, C1-6 alkyl, haloalkyl, hydroxyalkyl, (carboxamido)alkyl, (cycloalkyl)alkyl, optionally substituted C3-12 cycloalkyl, optionally substituted C6-14 aryl, optionally substituted 5- to 14-membered heteroaryl, optionally substituted 4- to 14-membered heterocyclo, and aralkyl; Y is -C(R1a)(R1b)-; or Y is absent; B is:
Figure imgf000086_0001
; X is selected from the group consisting of -S(=O)2-, -S(=O)2N(R6)-, -S(=O)2C(R7)(H)-, -C(=O)-, -C(=O)N(R6)-, -C(=O)O-, and -C(=O)C(R7)(H)-; or X is absent; Z is selected from the group consisting of hydrogen, optionally substituted C1-6 alkyl, hydroxyalkyl, (amino)alk ino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (hetero o)(hydroxy)alkyl, (amino)(aryl)alkyl, (hydroxy)(aryl)alkyl, (aralkylam kyl, optionally substituted C6-14 aryl, optionally substituted 4- to 14-membered heterocyclo, optionally substituted 5- to 14-membered heteroaryl, optionally substituted C3-12 cycloalkyl, aralkyl, and heteroaralkyl; R1s and R1b are independently selected from the group consisting of hydrogen and C1-4 alkyl; R2a, R2b, R3a, R3b, R4a, and R4b are each independently selected from the group consisting of hydrogen, amino, alkylamino, dialkylamino, cycloalkylamino, halo, hydroxy, C1-6 alkyl, alkoxy, haloalkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally substituted C3-12 cycloalkyl, optionally substituted C6-14 aryl, optionally substituted 4- to 14-membered heterocyclo, optionally substituted 5- to 14-membered heteroaryl, alkoxyalkyl, aralkyl, alkoxycarbonyl, sulfonamido, carboxamido, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or R2a and R2b taken together with the carbon atom to which they are attached form a carbonyl; and R3a, R3b, R4a, and R4b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, aralkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or R3a and R3b taken together with the carbon atom to which they are attached form a carbonyl; and R2a, R2b, R4a, and R4b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or R4a and R4b taken together with the carbon atom to which they are attached form a carbonyl; and R2a, R2b, R3a, an dependently selected from the group consisting of hydrogen, hydrox oxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl )C(=O)R8a; and -CH2N(H)C(=O)R8b; or R2a and R2b taken together with the carbon atom to which they are attached form a C3-6 cycloalkyl or C3-6 heterocyclo; and R3a, R3b, R4a, and R4b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, aralkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or R3a and R3b taken together with the carbon atom to which they are attached form a C3-6 cycloalkyl or C3-6 heterocyclo; and R2a, R2b, R4a, and R4b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; or R4a and R4b taken together with the carbon atom to which they are attached form a C3-6 cycloalkyl or C3-6 heterocyclo; and R2a, R2b, R3a, and R3b are each independently selected from the group consisting of hydrogen, hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, -N(H)C(=O)R8a; and -CH2N(H)C(=O)R8b; R5 is selected from the group consisting of hydrogen and C1-4 alkyl; R6 is selected from the group consisting of hydrogen and C1-4 alkyl; R7 is selected from the group consisting of hydrogen, C1-4 alkyl, amino, alkylamino, dialkylamino, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, and hydroxyalkyl; R8a is selected from the group consisting of C1-6 alkyl, haloalkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally substituted C aryl, optionally substituted 4- to 14-membered heterocyclo, opti 5- to 14-membered heteroaryl, optionally substituted C3-12 cyclo heteroaralkyl; and R8b is selected from the group consisting of C1-6 alkyl, haloalkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally substituted C6-14 aryl, optionally substituted 4- to 14-membered heterocyclo, optionally substituted 5- to 14-membered heteroaryl, optionally substituted C3-12 cycloalkyl, aralkyl, and heteroaralkyl.
2. The compound of claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein B is selected from the rou consistin of:
Figure imgf000089_0001
.
3. The compound of claim 1, or a pharmaceutically acceptable salt or hydrate thereof, wherein B is selected from the group consisting of:
Figure imgf000089_0002
R4a is selected from the group consisting of hydroxy, C1-6 alkyl, alkoxy, hydroxyalkyl, C3-12 cycloalkyl, optionally substituted C6-14 aryl, alkoxyalkyl, and aralkyl.
4. The compound of claim 3, or a pharmaceutically acceptable salt or hydrate thereof, wherein R4a is C1-4 alkyl.
5. The compound of claim 4, or a pharmaceutically acceptable salt or hydrate thereof, wherein R4a is methyl.
6. The compound of any on a pharmaceutically acceptable salt or solvate thereof, wherein X is sel p consisting of -S(=O)2- and -C(=O)-; or X is absent.
7. The compound of claim 6, or a pharmaceutically acceptable salt or solvate thereof, wherein X is -S(=O)2-.
8. The compound of claim 6, or a pharmaceutically acceptable salt or solvate thereof, wherein X is -C(=O)-.
9. The compound of any one of claims 1-8, or a pharmaceutically acceptable salt or solvate thereof, wherein X is absent.
10. The compound of any one of claims 1-9, or a pharmaceutically acceptable salt or solvate thereof, wherein Z is selected from the group consisting of optionally substituted C1-6 alkyl, hydroxyalkyl, (amino)alkyl, (alkylamino)alkyl, (dialkylamino)alkyl, (cycloalkylamino)alkyl, (heterocyclo)alkyl, optionally substituted C6-14 aryl, optionally substituted 4- to 14-membered heterocyclo, optionally substituted 5- to 14-membered heteroaryl, optionally substituted C3-12 cycloalkyl, aralkyl, and heteroaralkyl.
11. The compound of claim 10, or a pharmaceutically acceptable salt or solvate thereof, wherein Z is selected from the group consisting of C1-6 alkyl and optionally substituted C3-12 cycloalkyl.
12. The compound of any one of claims 1-11, or a pharmaceutically acceptable salt or solvate thereof, wherein Y is -CH2-.
13. The compound of any one of claims 1-11, or a pharmaceutically acceptable salt or solvate thereof, wherein Y is absent.
14. The compound of any one of claims 1-13, or a pharmaceutically acceptable salt or solvate thereof, wherein A is selected from the group consisting of 5-indolinyl-2-one, 6- benzo[b][1,4]oxazinyl-3-one, 5-pyrazolyl, 3-isothiazolyl, 6-benzo[d]oxazolyl-2(3H)-one, 1,2,3-triazolyl, phenyl, 5-(1,3 dihydro 2H benzo[d]imidazolyl-2-one), 3-pyridazinyl, 1,2,3-triazolyl, 3-isothiazolyl, 2-pyridyl, 1,3,4-thiadiazole, and 1,3,4-oxadiazole.
15. The compound of claim 14, or a pharmaceutically acceptable salt thereof, wherein A is 5-indolinyl-2-one.
16. The compound of claim 1, or a pharmaceutically acceptable salt thereof, selected from any one or more of the compounds provided in Tables 1, 3, and 4 of the specification.
17. A pharmaceutical composition comprising the compound of any one of claims 1-16, or a pharmaceutically acceptable salt or solvate thereof, and a pharmaceutically acceptable carrier.
18. A method of treating a patient comprising administering to the patient a therapeutically effective amount of the compound of any one of claims 1-16, or a pharmaceutically acceptable salt or hydrate thereof, wherein the patient has cancer.
19. The method of claim 18, wherein the cancer is selected from the group consisting of adrenal cancer, acinic cell carcinoma, acoustic neuroma, acral lentigious melanoma, acrospiroma, acute eosinophilic leukemia, acute erythroid leukemia, acute lymphoblastic leukemia, acute megakaryoblastic leukemia, acute monocytic leukemia, acute promyelocytic leukemia, adenocarcinoma, adenoid cystic carcinoma, adenoma, adenomatoid odontogenic tumor, adenosquamous carcinoma, adipose tissue neoplasm, adrenocortical carcinoma, adult T-cell leukemia/lymphoma, aggressive NK-cell leukemia, AIDS-related lymphoma, alveolar rhabdomyosarcoma, alveolar soft part sarcoma, ameloblastic fibroma, anaplastic large cell lymphoma, anaplastic thyroid cancer, angioimmunoblastic T-cell lymphoma, angiomyolipoma, angiosarcoma, astrocytoma, atypical teratoid rhabdoid tumor, B-cell chronic lymphocytic leukemia, B- cell prolymphocytic leukemia, B-cell lymphoma, basal cell carcinoma, biliary tract cancer, bladder cancer, blastoma bone cancer Brenner tumor, Brown tumor, Burkitt's lymphoma, breast cancer, brain , carcinoma in situ, carcinosarcoma, cartilage tumor, cementoma, my ndroma, chordoma, choriocarcinoma, choroid plexus papilloma, cl of the kidney, craniopharyngioma, cutaneous T-cell lymphoma, cervical cancer, colorectal cancer, Degos disease, desmoplastic small round cell tumor, diffuse large B-cell lymphoma, dysembryoplastic neuroepithelial tumor, dysgerminoma, embryonal carcinoma, endocrine gland neoplasm, endodermal sinus tumor, enteropathy-associated T-cell lymphoma, esophageal cancer, fetus in fetu, fibroma, fibrosarcoma, follicular lymphoma, follicular thyroid cancer, ganglioneuroma, gastrointestinal cancer, germ cell tumor, gestational choriocarcinoma, giant cell fibroblastoma, giant cell tumor of the bone, glial tumor, glioblastoma multiforme, glioma, gliomatosis cerebri, glucagonoma, gonadoblastoma, granulosa cell tumor, gynandroblastoma, gallbladder cancer, gastric cancer, hairy cell leukemia, hemangioblastoma, head and neck cancer, hemangiopericytoma, hematological malignancy, hepatoblastoma, hepatosplenic T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, invasive lobular carcinoma, intestinal cancer, kidney cancer, laryngeal cancer, lentigo maligna, lethal midline carcinoma, leukemia, leydig cell tumor, liposarcoma, lung cancer, lymphangioma, lymphangiosarcoma, lymphoepithelioma, lymphoma, acute lymphocytic leukemia, acute myelogeous leukemia, chronic lymphocytic leukemia, liver cancer, small cell lung cancer, non-small cell lung cancer, MALT lymphoma, malignant fibrous histiocytoma, malignant peripheral nerve sheath tumor, malignant triton tumor, mantle cell lymphoma, marginal zone B-cell lymphoma, mast cell leukemia, mediastinal germ cell tumor, medullary carcinoma of the breast, medullary thyroid cancer, medulloblastoma, melanoma, meningioma, merkel cell cancer, mesothelioma, metastatic urothelial carcinoma, mixed Mullerian tumor, mucinous tumor, multiple myeloma, muscle tissue neoplasm, mycosis fungoides, myxoid liposarcoma, myxoma, myxosarcoma, nasopharyngeal carcinoma, neurinoma, neuroblastoma, neurofibroma, neuroma, nodular melanoma, ocular cancer, oligoastrocytoma, oligodendroglioma, oncocytoma, optic nerve sheath meningioma, optic nerve tumor, oral cancer, osteosarcoma, ovarian cancer, Pancoast tumor, papillary thyroid cancer, paraganglioma, pinealoblastoma, pineocytoma, pituicytoma, pituitary adenoma, pituitary tumor, plasmacytoma, polyembryoma, precursor T-lymphoblastic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, preimary peritoneal cancer, prostate cancer, pancreatic cancer pharyngeal cancer, pseudomyxoma periotonei, renal cell carcinoma, renal ma, retinoblastoma, rhabdomyoma, rhabdomyosarcoma, Richter's tr l cancer, sarcoma, Schwannomatosis, seminoma, Sertoli cell tumor, se mal tumor, signet ring cell carcinoma, skin cancer, small blue round cell tumors, small cell carcinoma, soft tissue sarcoma, somatostatinoma, soot wart, spinal tumor, splenic marginal zone lymphoma, squamous cell carcinoma, synovial sarcoma, Sezary's disease, small intestine cancer, squamous carcinoma, stomach cancer, T-cell lymphoma, testicular cancer, thecoma, thyroid cancer, transitional cell carcinoma, throat cancer, urachal cancer, urogenital cancer, urothelial carcinoma, uveal melanoma, uterine cancer, verrucous carcinoma, visual pathway glioma, vulvar cancer, vaginal cancer, Waldenstrom's macroglobulinemia, Warthin's tumor, and Wilms' tumor.
20. The pharmaceutical composition of claim 17 for use in treating cancer.
21. The pharmaceutical composition of claim 20, wherein the cancer is selected from the group consisting of adrenal cancer, acinic cell carcinoma, acoustic neuroma, acral lentigious melanoma, acrospiroma, acute eosinophilic leukemia, acute erythroid leukemia, acute lymphoblastic leukemia, acute megakaryoblastic leukemia, acute monocytic leukemia, acute promyelocytic leukemia, adenocarcinoma, adenoid cystic carcinoma, adenoma, adenomatoid odontogenic tumor, adenosquamous carcinoma, adipose tissue neoplasm, adrenocortical carcinoma, adult T-cell leukemia/lymphoma, aggressive NK-cell leukemia, AIDS-related lymphoma, alveolar rhabdomyosarcoma, alveolar soft part sarcoma, ameloblastic fibroma, anaplastic large cell lymphoma, anaplastic thyroid cancer, angioimmunoblastic T-cell lymphoma, angiomyolipoma, angiosarcoma, astrocytoma, atypical teratoid rhabdoid tumor, B-cell chronic lymphocytic leukemia, B-cell prolymphocytic leukemia, B-cell lymphoma, basal cell carcinoma, biliary tract cancer, bladder cancer, blastoma, bone cancer, Brenner tumor, Brown tumor, Burkitt's lymphoma, breast cancer, brain cancer, carcinoma, carcinoma in situ, carcinosarcoma, cartilage tumor, cementoma, myeloid sarcoma, chondroma, chordoma, choriocarcinoma, choroid plexus papilloma, clear-cell sarcoma of the kidney, craniopharyngioma, cutaneous T-cell lymphoma, cervical cancer, colorectal cancer, Degos disease, desmoplastic small round cell tumor, diffuse large B-cell lymphoma, dysembryoplastic neuroepithelial tumor dysgerminoma, embryonal carcinoma, endocrine gland neoplasm, en umor, enteropathy-associated T-cell lymphoma, esophageal cancer, f a, fibrosarcoma, follicular lymphoma, follicular thyroid cancer, gang intestinal cancer, germ cell tumor, gestational choriocarcinoma, giant cell fibroblastoma, giant cell tumor of the bone, glial tumor, glioblastoma multiforme, glioma, gliomatosis cerebri, glucagonoma, gonadoblastoma, granulosa cell tumor, gynandroblastoma, gallbladder cancer, gastric cancer, hairy cell leukemia, hemangioblastoma, head and neck cancer, hemangiopericytoma, hematological malignancy, hepatoblastoma, hepatosplenic T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, invasive lobular carcinoma, intestinal cancer, kidney cancer, laryngeal cancer, lentigo maligna, lethal midline carcinoma, leukemia, leydig cell tumor, liposarcoma, lung cancer, lymphangioma, lymphangiosarcoma, lymphoepithelioma, lymphoma, acute lymphocytic leukemia, acute myelogeous leukemia, chronic lymphocytic leukemia, liver cancer, small cell lung cancer, non-small cell lung cancer, MALT lymphoma, malignant fibrous histiocytoma, malignant peripheral nerve sheath tumor, malignant triton tumor, mantle cell lymphoma, marginal zone B-cell lymphoma, mast cell leukemia, mediastinal germ cell tumor, medullary carcinoma of the breast, medullary thyroid cancer, medulloblastoma, melanoma, meningioma, merkel cell cancer, mesothelioma, metastatic urothelial carcinoma, mixed Mullerian tumor, mucinous tumor, multiple myeloma, muscle tissue neoplasm, mycosis fungoides, myxoid liposarcoma, myxoma, myxosarcoma, nasopharyngeal carcinoma, neurinoma, neuroblastoma, neurofibroma, neuroma, nodular melanoma, ocular cancer, oligoastrocytoma, oligodendroglioma, oncocytoma, optic nerve sheath meningioma, optic nerve tumor, oral cancer, osteosarcoma, ovarian cancer, Pancoast tumor, papillary thyroid cancer, paraganglioma, pinealoblastoma, pineocytoma, pituicytoma, pituitary adenoma, pituitary tumor, plasmacytoma, polyembryoma, precursor T-lymphoblastic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, preimary peritoneal cancer, prostate cancer, pancreatic cancer, pharyngeal cancer, pseudomyxoma periotonei, renal cell carcinoma, renal medullary carcinoma, retinoblastoma, rhabdomyoma, rhabdomyosarcoma, Richter's transformation, rectal cancer, sarcoma, Schwannomatosis, seminoma, Sertoli cell tumor, sex cord-gonadal stromal tumor, signet ring cell carcinoma, skin cancer, small blue round cell tumors, small cell carcinoma, soft tissue sarcoma, somatostatinoma, soot wart, spinal tumor, splenic marginal zone lymphoma squamous cell carcinoma, synovial sarcoma, Sezary's disease, small intestin s carcinoma, stomach cancer, T-cell lymphoma, testicular cancer, the er, transitional cell carcinoma, throat cancer, urachal cancer, urogenit carcinoma, uveal melanoma, uterine cancer, verrucous carcinoma, visual pathway glioma, vulvar cancer, vaginal cancer, Waldenstrom's macroglobulinemia, Warthin's tumor, and Wilms' tumor.
22. A compound of any one of claims 1-16, or a pharmaceutically acceptable salt or hydrate thereof, for use in treatment of cancer.
23. The compound of claim 22, wherein the cancer is selected from the group consisting of adrenal cancer, acinic cell carcinoma, acoustic neuroma, acral lentigious melanoma, acrospiroma, acute eosinophilic leukemia, acute erythroid leukemia, acute lymphoblastic leukemia, acute megakaryoblastic leukemia, acute monocytic leukemia, acute promyelocytic leukemia, adenocarcinoma, adenoid cystic carcinoma, adenoma, adenomatoid odontogenic tumor, adenosquamous carcinoma, adipose tissue neoplasm, adrenocortical carcinoma, adult T-cell leukemia/lymphoma, aggressive NK-cell leukemia, AIDS-related lymphoma, alveolar rhabdomyosarcoma, alveolar soft part sarcoma, ameloblastic fibroma, anaplastic large cell lymphoma, anaplastic thyroid cancer, angioimmunoblastic T-cell lymphoma, angiomyolipoma, angiosarcoma, astrocytoma, atypical teratoid rhabdoid tumor, B-cell chronic lymphocytic leukemia, B- cell prolymphocytic leukemia, B-cell lymphoma, basal cell carcinoma, biliary tract cancer, bladder cancer, blastoma, bone cancer, Brenner tumor, Brown tumor, Burkitt's lymphoma, breast cancer, brain cancer, carcinoma, carcinoma in situ, carcinosarcoma, cartilage tumor, cementoma, myeloid sarcoma, chondroma, chordoma, choriocarcinoma, choroid plexus papilloma, clear-cell sarcoma of the kidney, craniopharyngioma, cutaneous T-cell lymphoma, cervical cancer, colorectal cancer, Degos disease, desmoplastic small round cell tumor, diffuse large B-cell lymphoma, dysembryoplastic neuroepithelial tumor, dysgerminoma, embryonal carcinoma, endocrine gland neoplasm, endodermal sinus tumor, enteropathy-associated T-cell lymphoma, esophageal cancer, fetus in fetu, fibroma, fibrosarcoma, follicular lymphoma, follicular thyroid cancer, ganglioneuroma, gastrointestinal cancer, germ cell tumor, gestational choriocarcinoma, giant cell fibroblastoma, giant cell tumor of the bone, glial tumor, glioblastoma multiforme, glioma, gliomatosis cerebri glucagonoma, gonadoblastoma, granulosa cell tumor, gynandroblastoma, gall astric cancer, hairy cell leukemia, hemangioblastoma, head and hemangiopericytoma, hematological malignancy, hepatoblastoma, h l lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, invasive lobular carcinoma, intestinal cancer, kidney cancer, laryngeal cancer, lentigo maligna, lethal midline carcinoma, leukemia, leydig cell tumor, liposarcoma, lung cancer, lymphangioma, lymphangiosarcoma, lymphoepithelioma, lymphoma, acute lymphocytic leukemia, acute myelogeous leukemia, chronic lymphocytic leukemia, liver cancer, small cell lung cancer, non-small cell lung cancer, MALT lymphoma, malignant fibrous histiocytoma, malignant peripheral nerve sheath tumor, malignant triton tumor, mantle cell lymphoma, marginal zone B-cell lymphoma, mast cell leukemia, mediastinal germ cell tumor, medullary carcinoma of the breast, medullary thyroid cancer, medulloblastoma, melanoma, meningioma, merkel cell cancer, mesothelioma, metastatic urothelial carcinoma, mixed Mullerian tumor, mucinous tumor, multiple myeloma, muscle tissue neoplasm, mycosis fungoides, myxoid liposarcoma, myxoma, myxosarcoma, nasopharyngeal carcinoma, neurinoma, neuroblastoma, neurofibroma, neuroma, nodular melanoma, ocular cancer, oligoastrocytoma, oligodendroglioma, oncocytoma, optic nerve sheath meningioma, optic nerve tumor, oral cancer, osteosarcoma, ovarian cancer, Pancoast tumor, papillary thyroid cancer, paraganglioma, pinealoblastoma, pineocytoma, pituicytoma, pituitary adenoma, pituitary tumor, plasmacytoma, polyembryoma, precursor T-lymphoblastic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, preimary peritoneal cancer, prostate cancer, pancreatic cancer, pharyngeal cancer, pseudomyxoma periotonei, renal cell carcinoma, renal medullary carcinoma, retinoblastoma, rhabdomyoma, rhabdomyosarcoma, Richter's transformation, rectal cancer, sarcoma, Schwannomatosis, seminoma, Sertoli cell tumor, sex cord-gonadal stromal tumor, signet ring cell carcinoma, skin cancer, small blue round cell tumors, small cell carcinoma, soft tissue sarcoma, somatostatinoma, soot wart, spinal tumor, splenic marginal zone lymphoma, squamous cell carcinoma, synovial sarcoma, Sezary's disease, small intestine cancer, squamous carcinoma, stomach cancer, T-cell lymphoma, testicular cancer, thecoma, thyroid cancer, transitional cell carcinoma, throat cancer, urachal cancer, urogenital cancer, urothelial carcinoma, uveal melanoma, uterine cancer, verrucous carcinoma, visual pathway glioma, vulvar cancer, vaginal cancer, Waldenstrom's macroglobulinemia, Warthin's tumor, and Wilms' tumor.
24. Use of a compound of a 16, or a pharmaceutically acceptable salt or hydrate thereof, for the m icament for treatment of cancer.
25. The use of claim 24, wherein the cancer is selected from the group consisting of adrenal cancer, acinic cell carcinoma, acoustic neuroma, acral lentigious melanoma, acrospiroma, acute eosinophilic leukemia, acute erythroid leukemia, acute lymphoblastic leukemia, acute megakaryoblastic leukemia, acute monocytic leukemia, acute promyelocytic leukemia, adenocarcinoma, adenoid cystic carcinoma, adenoma, adenomatoid odontogenic tumor, adenosquamous carcinoma, adipose tissue neoplasm, adrenocortical carcinoma, adult T-cell leukemia/lymphoma, aggressive NK-cell leukemia, AIDS-related lymphoma, alveolar rhabdomyosarcoma, alveolar soft part sarcoma, ameloblastic fibroma, anaplastic large cell lymphoma, anaplastic thyroid cancer, angioimmunoblastic T-cell lymphoma, angiomyolipoma, angiosarcoma, astrocytoma, atypical teratoid rhabdoid tumor, B-cell chronic lymphocytic leukemia, B- cell prolymphocytic leukemia, B-cell lymphoma, basal cell carcinoma, biliary tract cancer, bladder cancer, blastoma, bone cancer, Brenner tumor, Brown tumor, Burkitt's lymphoma, breast cancer, brain cancer, carcinoma, carcinoma in situ, carcinosarcoma, cartilage tumor, cementoma, myeloid sarcoma, chondroma, chordoma, choriocarcinoma, choroid plexus papilloma, clear-cell sarcoma of the kidney, craniopharyngioma, cutaneous T-cell lymphoma, cervical cancer, colorectal cancer, Degos disease, desmoplastic small round cell tumor, diffuse large B-cell lymphoma, dysembryoplastic neuroepithelial tumor, dysgerminoma, embryonal carcinoma, endocrine gland neoplasm, endodermal sinus tumor, enteropathy-associated T-cell lymphoma, esophageal cancer, fetus in fetu, fibroma, fibrosarcoma, follicular lymphoma, follicular thyroid cancer, ganglioneuroma, gastrointestinal cancer, germ cell tumor, gestational choriocarcinoma, giant cell fibroblastoma, giant cell tumor of the bone, glial tumor, glioblastoma multiforme, glioma, gliomatosis cerebri, glucagonoma, gonadoblastoma, granulosa cell tumor, gynandroblastoma, gallbladder cancer, gastric cancer, hairy cell leukemia, hemangioblastoma, head and neck cancer, hemangiopericytoma, hematological malignancy, hepatoblastoma, hepatosplenic T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, invasive lobular carcinoma, intestinal cancer, kidney cancer, laryngeal cancer, lentigo maligna lethal midline carcinoma, leukemia, leydig cell tumor, liposarcoma, lung cancer, lym hangiosarcoma, lymphoepithelioma, lymphoma, acute lymphocyti e myelogeous leukemia, chronic lymphocytic leukemia, liver can g cancer, non-small cell lung cancer, MALT lymphoma, malignant fibrous histiocytoma, malignant peripheral nerve sheath tumor, malignant triton tumor, mantle cell lymphoma, marginal zone B-cell lymphoma, mast cell leukemia, mediastinal germ cell tumor, medullary carcinoma of the breast, medullary thyroid cancer, medulloblastoma, melanoma, meningioma, merkel cell cancer, mesothelioma, metastatic urothelial carcinoma, mixed Mullerian tumor, mucinous tumor, multiple myeloma, muscle tissue neoplasm, mycosis fungoides, myxoid liposarcoma, myxoma, myxosarcoma, nasopharyngeal carcinoma, neurinoma, neuroblastoma, neurofibroma, neuroma, nodular melanoma, ocular cancer, oligoastrocytoma, oligodendroglioma, oncocytoma, optic nerve sheath meningioma, optic nerve tumor, oral cancer, osteosarcoma, ovarian cancer, Pancoast tumor, papillary thyroid cancer, paraganglioma, pinealoblastoma, pineocytoma, pituicytoma, pituitary adenoma, pituitary tumor, plasmacytoma, polyembryoma, precursor T-lymphoblastic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, preimary peritoneal cancer, prostate cancer, pancreatic cancer, pharyngeal cancer, pseudomyxoma periotonei, renal cell carcinoma, renal medullary carcinoma, retinoblastoma, rhabdomyoma, rhabdomyosarcoma, Richter's transformation, rectal cancer, sarcoma, Schwannomatosis, seminoma, Sertoli cell tumor, sex cord-gonadal stromal tumor, signet ring cell carcinoma, skin cancer, small blue round cell tumors, small cell carcinoma, soft tissue sarcoma, somatostatinoma, soot wart, spinal tumor, splenic marginal zone lymphoma, squamous cell carcinoma, synovial sarcoma, Sezary's disease, small intestine cancer, squamous carcinoma, stomach cancer, T-cell lymphoma, testicular cancer, thecoma, thyroid cancer, transitional cell carcinoma, throat cancer, urachal cancer, urogenital cancer, urothelial carcinoma, uveal melanoma, uterine cancer, verrucous carcinoma, visual pathway glioma, vulvar cancer, vaginal cancer, Waldenstrom's macroglobulinemia, Warthin's tumor, and Wilms' tumor.
26. A kit comprising the compound of any one of claims 1-16, or a pharmaceutically acceptable salt or hydrate thereof, and instructions for administering the compound, or a pharmaceutically acceptable salt or hydrate thereof, to a patient having cancer.
27. The kit of claim 26, whe selected from the group consisting of adrenal cancer, acinic cell car euroma, acral lentigious melanoma, acrospiroma, acute eosinophilic throid leukemia, acute lymphoblastic leukemia, acute megakaryoblastic leukemia, acute monocytic leukemia, acute promyelocytic leukemia, adenocarcinoma, adenoid cystic carcinoma, adenoma, adenomatoid odontogenic tumor, adenosquamous carcinoma, adipose tissue neoplasm, adrenocortical carcinoma, adult T-cell leukemia/lymphoma, aggressive NK-cell leukemia, AIDS-related lymphoma, alveolar rhabdomyosarcoma, alveolar soft part sarcoma, ameloblastic fibroma, anaplastic large cell lymphoma, anaplastic thyroid cancer, angioimmunoblastic T-cell lymphoma, angiomyolipoma, angiosarcoma, astrocytoma, atypical teratoid rhabdoid tumor, B-cell chronic lymphocytic leukemia, B- cell prolymphocytic leukemia, B-cell lymphoma, basal cell carcinoma, biliary tract cancer, bladder cancer, blastoma, bone cancer, Brenner tumor, Brown tumor, Burkitt's lymphoma, breast cancer, brain cancer, carcinoma, carcinoma in situ, carcinosarcoma, cartilage tumor, cementoma, myeloid sarcoma, chondroma, chordoma, choriocarcinoma, choroid plexus papilloma, clear-cell sarcoma of the kidney, craniopharyngioma, cutaneous T-cell lymphoma, cervical cancer, colorectal cancer, Degos disease, desmoplastic small round cell tumor, diffuse large B-cell lymphoma, dysembryoplastic neuroepithelial tumor, dysgerminoma, embryonal carcinoma, endocrine gland neoplasm, endodermal sinus tumor, enteropathy-associated T-cell lymphoma, esophageal cancer, fetus in fetu, fibroma, fibrosarcoma, follicular lymphoma, follicular thyroid cancer, ganglioneuroma, gastrointestinal cancer, germ cell tumor, gestational choriocarcinoma, giant cell fibroblastoma, giant cell tumor of the bone, glial tumor, glioblastoma multiforme, glioma, gliomatosis cerebri, glucagonoma, gonadoblastoma, granulosa cell tumor, gynandroblastoma, gallbladder cancer, gastric cancer, hairy cell leukemia, hemangioblastoma, head and neck cancer, hemangiopericytoma, hematological malignancy, hepatoblastoma, hepatosplenic T-cell lymphoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, invasive lobular carcinoma, intestinal cancer, kidney cancer, laryngeal cancer, lentigo maligna, lethal midline carcinoma, leukemia, leydig cell tumor, liposarcoma, lung cancer, lymphangioma, lymphangiosarcoma, lymphoepithelioma, lymphoma, acute lymphocytic leukemia, acute myelogeous leukemia, chronic lymphocytic leukemia, liver cancer, small cell lung cancer, non-small cell lung cancer, MALT lymphoma, malignant fibrous histiocytoma, malignant peripheral nerve sheath tumor, malignant triton tumor, m ma, marginal zone B-cell lymphoma, mast cell leukemia, mediastina medullary carcinoma of the breast, medullary thyroid cancer, medu ma, meningioma, merkel cell cancer, mesothelioma, metastatic urothelial carcinoma, mixed Mullerian tumor, mucinous tumor, multiple myeloma, muscle tissue neoplasm, mycosis fungoides, myxoid liposarcoma, myxoma, myxosarcoma, nasopharyngeal carcinoma, neurinoma, neuroblastoma, neurofibroma, neuroma, nodular melanoma, ocular cancer, oligoastrocytoma, oligodendroglioma, oncocytoma, optic nerve sheath meningioma, optic nerve tumor, oral cancer, osteosarcoma, ovarian cancer, Pancoast tumor, papillary thyroid cancer, paraganglioma, pinealoblastoma, pineocytoma, pituicytoma, pituitary adenoma, pituitary tumor, plasmacytoma, polyembryoma, precursor T-lymphoblastic lymphoma, primary central nervous system lymphoma, primary effusion lymphoma, preimary peritoneal cancer, prostate cancer, pancreatic cancer, pharyngeal cancer, pseudomyxoma periotonei, renal cell carcinoma, renal medullary carcinoma, retinoblastoma, rhabdomyoma, rhabdomyosarcoma, Richter's transformation, rectal cancer, sarcoma, Schwannomatosis, seminoma, Sertoli cell tumor, sex cord-gonadal stromal tumor, signet ring cell carcinoma, skin cancer, small blue round cell tumors, small cell carcinoma, soft tissue sarcoma, somatostatinoma, soot wart, spinal tumor, splenic marginal zone lymphoma, squamous cell carcinoma, synovial sarcoma, Sezary's disease, small intestine cancer, squamous carcinoma, stomach cancer, T-cell lymphoma, testicular cancer, thecoma, thyroid cancer, transitional cell carcinoma, throat cancer, urachal cancer, urogenital cancer, urothelial carcinoma, uveal melanoma, uterine cancer, verrucous carcinoma, visual pathway glioma, vulvar cancer, vaginal cancer, Waldenstrom's macroglobulinemia, Warthin's tumor, and Wilms' tumor.
28. A method of treating a SMYD protein mediated disorder comprising administering to a subject in need thereof a compound of any one of claims 1-16, or a pharmaceutically acceptable salt or hydrate thereof in an effective amount to treat the SMYD protein mediated disorder.
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