Heterobifunctional compounds and methods of treating disease

Abstract

The present invention provides hetero-bifunctional compounds, crystalline forms of hetero-bifunctional compounds, pharmaceutical compositions and their use in the treatment of diseases such as cancer.

Description

TECHNICAL FIELD

[0001] The present invention provides heterobifunctional compounds, crystalline forms of heterobifunctional compounds, pharmaceutical compositions, and their use in treating diseases such as cancer. BACKGROUND

[0002] Despite the large amount of research effort and scientific progress reported in the literature regarding the treatment of cancer, this disease remains a major health problem. Solid tumors, including prostate cancer, breast cancer, and lung cancer, remain highly prevalent in the world population. The incidence of prostate cancer increases with age, and as human subjects live longer, the number of patients suffering from prostate cancer continues to increase accordingly. Breast cancer is one of the most common cancers in women and is the leading cause of death in women aged 50-55. Lung cancer is the leading cause of cancer patient deaths, with over 85% of lung cancers being non-small cell lung cancer (NSCLC). Many lung cancers are attributed to smoking. Current treatment options for these cancers are not effective for all patients and / or can have substantial adverse side effects.

[0003] New therapies are needed to address this unmet need in cancer therapy. In particular, new therapies are needed that achieve an anti-cancer effect via a different mechanism than commonly used therapies. Exemplary mechanisms of common anti-cancer therapies include (a) DNA alkylation that limits the ability of cells to proliferate, (b) topoisomerase inhibition, where a therapeutic agent inhibits the activity of topoisomerase, thereby limiting the separation of DNA strands, and (c) mitotic inhibition, where a therapeutic agent reduces the ability of cells to divide. New therapies that achieve an anti-cancer effect via a different mechanism offer the opportunity to more effectively treat cancer and / or treat cancers that have become resistant to currently available drugs.

[0004] The present invention addresses the foregoing needs and provides other related advantages. SUMMARY

[0005] The present invention provides heterobifunctional compounds, crystalline forms of heterobifunctional compounds, pharmaceutical compositions, and their use in treating diseases such as cancer. In particular, one aspect of the present invention provides a crystalline compound of Formula II-5:

[0006]

[0007] The compound can be part of a pharmaceutical composition comprising a pharmaceutically acceptable carrier.

[0008] Another aspect of the present invention provides a compound of Table 3 herein, or a pharmaceutically acceptable salt thereof. The compound can be part of a pharmaceutical composition comprising a pharmaceutically acceptable carrier.

[0009] Another aspect of the application provides a method of treating cancer. The method includes administering to a patient in need thereof a therapeutically effective amount of a compound described herein, such as a crystalline compound of Formula II-5 or a compound in Table 3, or a pharmaceutically acceptable salt thereof, to treat cancer.

[0010] Another aspect of the application provides a method of causing cancer cell death. The method includes contacting a cancer cell with an effective amount of a compound described herein, such as a crystalline compound of Formula II-5 or a compound in Table 3, or a pharmaceutically acceptable salt thereof, to cause cancer cell death. BRIEF DESCRIPTION OF DRAWINGS

[0011] Figure 1 An X-ray powder diffraction pattern of compound II-5 in crystalline Form A is depicted, as further described in Example 3.

[0012] Figure 2 A differential scanning calorimetry curve of compound II-5 in crystalline Form A is depicted, as further described in Example 3.

[0013] Figure 3 A thermogravimetric analysis curve of compound II-5 in crystalline Form A is depicted, as further described in Example 3.

[0014] Figure 4 An X-ray powder diffraction pattern of compound II-5 in crystalline Form B is depicted, as further described in Example 5.

[0015] Figure 5 A differential scanning calorimetry curve of compound II-5 in crystalline Form B is depicted, as further described in Example 5.

[0016] Figure 6 A thermogravimetric analysis curve of compound II-5 in crystalline Form B is depicted, as further described in Example 5. DETAILED DESCRIPTION

[0017] The present invention provides heterobifunctional compounds, crystalline forms of heterobifunctional compounds, pharmaceutical compositions, and uses thereof in the treatment of diseases such as cancer. Practice of the present invention employs, unless otherwise indicated, conventional techniques of organic chemistry, pharmacology, molecular biology (including recombinant techniques), cell biology, biochemistry, and immunology. Such techniques are explained in the literature, such as "Comprehensive Organic Synthesis" (B. M. Trost and I. Fleming, eds., 1991-1992); "Handbook of experimental immunology" (D. M. Weir and C. C. Blackwell, eds.); "Current protocols in molecular biology" (F. M. Ausubel et al., eds., 1987, and periodic updates); and "Current protocols in immunology" (J. E. Coligan et al., eds., 1991), each entirely incorporated herein by reference.

[0018] Aspects of the present invention are set forth in the following sections; however, aspects of the present invention described in one particular section are not limited to that section. Moreover, when a variable is not defined, the previous definition of the variable applies.

[0019] Definitions

[0020] The compounds of the present invention include those generally described herein and are further illustrated by the classes, subclasses, and species disclosed herein. As used herein, the following definitions shall apply unless otherwise indicated. Unless otherwise indicated, these definitions are applicable regardless of whether a term is used by itself or in combination with other terms. Thus, the definition of "alkyl" applies to "alkyl" as well as the "alkyl" portion of "-O-alkyl" and the like. For purposes of this invention, the chemical elements are identified in accordance with the Periodic Table of the Elements, CAS version, Handbook of Chemistry and Physics, 75thEd. Additionally, general principles of organic chemistry are described in "Organic Chemistry", Thomas Sorrell, University Science Books, Sausalito: 1999, and "March's Advanced Organic Chemistry", 5thEd., Smith, M.B. and March, J. eds., John Wiley & Sons, New York: 2001, the entire contents of which are incorporated herein by reference.

[0021] As used herein, the terms "aliphatic" or "aliphatic group" mean a straight-chain (i.e., unbranched) or branched, substituted or unsubstituted hydrocarbon chain that is completely saturated or that contains one or more units of unsaturation, or a monocyclic or bicyclic hydrocarbon that is completely saturated or that contains one or more units of unsaturation, but not aromatic, with the point of attachment being a single ring or double ring ring system. Unless otherwise specified, an aliphatic group contains from 1 to 6 aliphatic carbon atoms. In some embodiments, an aliphatic group contains from 1 to 5 aliphatic carbon atoms. In other embodiments, an aliphatic group contains from 1 to 4 aliphatic carbon atoms. In other embodiments, an aliphatic group contains from 1 to 3 aliphatic carbon atoms, and in other embodiments, an aliphatic group contains from 1 to 2 aliphatic carbon atoms. In some embodiments, "cycloaliphatic" refers to a monocyclic C3-C6 hydrocarbon that is completely saturated or that contains one or more units of unsaturation, but not aromatic, with the point of attachment being a single ring ring system. Suitable aliphatic groups include, but are not limited to, linear or branched, substituted or unsubstituted alkyl, alkenyl, alkynyl, and heterocombinations thereof, such as (cycloalkyl)alkyl, (cycloalkenyl)alkyl, or (cycloalkyl)alkenyl groups.

[0022] As used herein, the term "bicyclic" or "bicyclic ring system" refers to any bicyclic ring system, i.e., carbocyclic or heterocyclic, saturated or having one or more units of unsaturation, having one or more atoms that are common to two rings of the ring system. Thus, the term includes any permissible ring fusions, such as ortho-fusion or spiro-fusion. As used herein, the term "heterobicyclic" is a subset of "bicyclic" that requires the presence of one or more heteroatoms in one or both rings of the bicyclic ring system. Such heteroatoms can be present at the ring juncture and are optionally substituted, and can be selected from nitrogen (including N-oxides), oxygen, sulfur (including oxidized forms, such as sulfone and sulfonate), phosphorus (including oxidized forms, such as phosphonate), boron, and the like. In some embodiments, bicyclic groups have 7-12 ring members and 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. As used herein, the term "bridged bicyclic" refers to any bicyclic ring system, i.e., carbocyclic or heterocyclic, saturated or partially unsaturated, having at least one bridge. As defined by IUPAC, a "bridge" is an unbranched chain of atoms or valence bonds connecting two bridgeheads, where a "bridgehead" is any skeletal atom of the ring system that is bonded to three or more skeletal atoms not including hydrogen. In some embodiments, bridged bicyclic groups have 7-12 ring members and 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur. Such bridged bicyclic groups are well known in the art and include those set forth below, wherein each group is attached to the remainder of the molecule at any substitutable carbon or nitrogen atom. Unless otherwise specified, a bridged bicyclic group is optionally substituted with one or more substituents as set forth for aliphatic groups. Additionally or alternatively, any substitutable nitrogen of a bridged bicyclic group is optionally substituted. Exemplary bicyclic rings include:

[0023]

[0024] Exemplary bridged bicyclic rings include:

[0025]

[0026]

[0027] The term "lower alkyl" refers to a C 1-4 straight or branched chain alkyl group. Exemplary lower alkyl groups are methyl, ethyl, propyl, isopropyl, butyl, isobutyl, and t-butyl.

[0028] The term "lower haloalkyl" refers to a C 1-4 straight or branched chain alkyl group substituted with one or more halogen atoms.

[0029] The term "heteroatom" means one or more of oxygen, sulfur, nitrogen, phosphorus, or silicon (including any oxidized form of nitrogen, sulfur, phosphorus, or silicon; any quaternized form of an alkali nitrogen; or a substitutable nitrogen of a heterocyclic ring, e.g., N (as in 3,4-dihydro-2H-pyrrolyl), NH (as in pyrrolidinyl), or NR + (as in N-substituted pyrrolidinyl).

[0030] The term "unsaturated" as used herein means that the moiety has one or more units of unsaturation.

[0031] The term "divalent C 1-8 (or C 1-6 )saturated or unsaturated, straight or branched chain" refers to a straight or branched chain divalent alkylene, alkenylene, and alkynylene chain as defined herein.

[0032] The term "alkylene" refers to a divalent alkyl group. An "alkylene chain" is a polymethylene, i.e., -(CH2) n wherein n is a positive integer, preferably 1 to 6, 1 to 4, 1 to 3, 1 to 2, or 2 to 3. A substituted alkylene chain is a polymethylene in which one or more of the methylene hydrogen atoms are replaced by a substituent. Suitable substituents include those described below for substituted aliphatic groups.

[0033] The term "-(C0alkylene)-" refers to a bond. Thus, the term "-(C 0-3 alkylene)-" encompasses a bond (i.e., C0) and -(C 1-3 alkylene)- groups.

[0034] The term "alkenylene" refers to a divalent alkenyl group. A substituted alkenylene chain is a polymethylene containing at least one double bond and in which one or more of the hydrogen atoms are replaced by a substituent. Suitable substituents include those described below for substituted aliphatic groups.

[0035] The term "halogen" means F, Cl, Br, or I.

[0036] The term "aryl" used alone or as part of a larger moiety, e.g., in "aralkyl," "aralkoxy," or "aryloxyalkyl," refers to a monocyclic or bicyclic ring system having from five to fourteen ring members in which at least one ring is aromatic and in which each ring in the system contains from 3 to 7 ring members. The term "aryl" is used interchangeably with the term "aryl ring." In certain embodiments of the application, "aryl" refers to aromatic ring systems, including but not limited to phenyl, biphenyl, naphthyl, anthryl, and the like, which can bear one or more substituents. As used herein, also included within the scope of the term "aryl" are groups in which an aromatic ring is fused to one or more non-aromatic rings, such as indanyl, phthalimidyl, naphthalimidyl, phenanthridinyl, or tetrahydronaphthyl, and the like. The term "haloaryl" refers to aryl groups substituted with at least one halogen. Exemplary haloaryl groups include chlorophenyl (e.g., 3-chlorophenyl, 4-chlorophenyl), fluorophenyl, and the like. The term "phenylene" refers to divalent phenyl groups.

[0037] The terms "heteroaryl" and "heteroary-" used alone or as part of a larger portion (e.g., "heteroarylalkyl" or "heteroarylalkoxy") refer to a group having 5 to 10 ring atoms, preferably 5, 6, or 9 ring atoms; having 6, 10, or 14 π electrons shared in the ring array; and having one to five heteroatoms in addition to carbon atoms. The term "heteroatom" refers to nitrogen, oxygen, or sulfur, and includes any oxidized form of nitrogen or sulfur and any quaternized form of basic nitrogen. Heteroaryl groups include, but are not limited to, thienyl, furanyl, pyrroleyl, imidazolyl, pyrazolyl, triazolyl, tetrazolyl, oxazolyl, isoxazolyl, oxadiazolyl, thiazolyl, isothiazolyl, thiazolyl, pyridinyl, pyrazinyl, indazinyl, purinyl, naphridinyl, and pteridinyl. As used herein, the terms "heteroaryl" and "heteroary-" also include groups in which the heteroaryl ring is fused with one or more aryl, alicyclic, or heterocyclic rings, unless otherwise specified, wherein the linking group or linking point is on the heteroaryl ring or on a ring fused with the heteroaryl ring. Non-limiting examples include indolyl, isoindolyl, benzothiopheneyl, benzofuranyl, dibenzofuranyl, indazoleyl, benzimidazolyl, benzothiazolyl, quinolinyl, isoquinolinyl, terpineyl, phthalazinyl, quinazolinyl, quinoxalinyl, 4H-quinazinyl, carbazoleyl, acridineyl, phenazinyl, phenothiazinyl, phenotoxazinyl, tetrahydroquinolinyl, and tetrahydroisoquinolinyl. Heteroaryl groups can be monocyclic or bicyclic. The terms "heteroaryl" and "heteroaryl ring," "heteroaryl," or "heteroaryl" are used interchangeably, any of which includes optionally substituted rings. The term "heteroaryl" refers to an alkyl group substituted with a heteroaryl group, wherein the alkyl and heteroaryl portions are optionally substituted independently. The term "haloheteroaryl" refers to a heteroaryl group substituted with at least one halogen. Exemplary haloheteroaryls include chloropyridine, fluopyridine, chloropyrazole, fluopyrazole, etc. The term "heteroaryl" refers to a divalent heteroaryl group. Similarly, the terms "pyrazolyl," "imidazolyl," and "pyrrolyl" refer to divalent pyrazolyl, imidazolyl, and pyrrolyl, respectively. Similarly, the terms "pyridazinyl," "pyrimidinyl," "pyrazinyl," and "pyridinyl" refer to divalent pyridazinyl, pyrimidinyl, pyrazinyl, and pyridinyl, respectively.

[0038] As used herein, the terms “heterocycle,” “heterocyclic group,” “heterocyclic group,” and “heterocyclic ring” are used interchangeably and refer to a stable 5- to 7-membered monocyclic or 7- to 10-membered bicyclic heterocyclic moiety that is saturated or partially unsaturated and has one or more, preferably one to four, heteroatoms as defined above, in addition to a carbon atom. When used to refer to the ring atom of a heterocycle, the term “nitrogen” includes substituted nitrogen. For example, in a saturated or partially unsaturated ring having 0 to 3 heteroatoms selected from oxygen, sulfur, or nitrogen, nitrogen can be N (e.g., in 3,4-dihydro-2H-pyrrole), NH (e.g., in pyrrolealkyl), or... + NR (e.g., in N-substituted pyrroleyl groups).

[0039] Heterocycle can be attached at any heteroatom or carbon atom that results in a stable structure and any of the ring atoms can be optionally substituted. Examples of such saturated or partially unsaturated heterocyclyl groups include, but are not limited to, tetrahydrofuranyl, tetrahydrothiophenyl, pyrrolidinyl, piperidinyl, pyrrolinyl, tetrahydroquinolinyl, tetrahydroisoquinolinyl, decahydroquinolinyl, oxazolidinyl, piperazinyl, dioxanyl, dioxolanyl, diazepanyl, oxazepanyl, thiazepanyl, morpholinyl, 2-oxa-6-azaspiro[3.3]heptane, and quinuclidinyl. The terms “heterocycle,” “heterocyclyl,” “heterocyclyl ring,” “heterocyclic group,” “heterocyclic moiety,” and “heterocyclic radical” are used interchangeably herein, and also include groups in which the heterocyclyl ring is fused to one or more aryl, heteroaryl, or cycloaliphatic rings, such as indolinyl, 3H- indolyl, benzodioxolanyl, phenanthrollidinyl, or tetrahydroquinolinyl. The heterocyclyl group can be mono- or bicyclic. The term “heterocyclylalkyl” means an alkyl group substituted with a heterocyclyl group, wherein the alkyl and heterocyclyl moieties independently are optionally substituted. The term “heterocyclylalkylene” means a divalent heterocyclyl group. The terms “piperidinylene,” “piperazinylene,” and “azetidinylene” mean divalent piperidinyl, piperazinyl, and azetidinyl, respectively.

[0040] As used herein, the term “heterocycloalkyl” means a saturated heterocyclyl group. The term “heterocycloalkylene” means a divalent heterocycloalkyl group.

[0041] As used herein, the term “partially unsaturated” means a ring moiety that includes at least one double or triple bond. The term “partially unsaturated” is intended to encompass rings that have multiple sites of unsaturation, but is not intended to include aryl or heteroaryl moieties as defined herein.

[0042] As described herein, the compounds of the application can contain “optionally substituted” moieties. In general, the term “substituted,” whether preceded by the term “optionally” or not, means that the specified moiety can be substituted or unsubstituted. Unless otherwise indicated, “optionally substituted” groups can have suitable substituents at each substitutable position of the group, and when more than one position in any given structure can be substituted with more than one substituent, such moiety can be substituted independently with the same or different substituent(s). Combinations of substituents envisioned by this application are preferably those that result in the formation of stable or chemically feasible compounds. The term “stable,” as used herein, refers to compounds that are not substantially altered when subjected to conditions that can denature or destroy non-stable compounds.

[0043] Each optional substituent on a substitutable carbon is independently selected from the group consisting of monovalent substituents of: halogen; -(CH2)0-2SR; -(CH2)0-2OR; -(CH2)0-2Ph, which can be optionally substituted with R; -(CH2)0-2(CH2)0-2SRA; -(CH2)0-2O(CH2)0-2RA; -(CH2)0-2N(RA)2; -(C1-4straight or branched alkylene)O(NH2)C(O)O(CH2)0-1CH3;0-4 R°; -(CH2) 0-4 OR°; -O(CH2) 0-4 R°; -O-(CH2) 0-4 C(O)OR°; -(CH2) 0-4 CH(OR°)2; -(CH2) 0- 4SR°; -(CH2) 0-4 Ph, which can be substituted by R°; -(CH2) 0-4 O(CH2) 0-1 Ph, which can be substituted by R°; -CH=CHPh, which can be substituted by R°; -(CH2) 0-4 O(CH2) 0-1 -pyridyl, which can be substituted by R°; -NO2; -CN; -N3; -(CH2) 0-4 N(R°)2; -(CH2) 0-4 N(R°)C(O)R°; -N(R°)C(S)R°; -(CH2) 0-4 N(R°)C(O)NR°2; -N(R°)C(S)NR°2; -(CH2) 0-4 N(R°)C(O)OR°; -N(R°)N(R°)C(O)R°; -N(R°)N(R°)C(O)NR°2; -N(R°)N(R°)C(O)OR°; -(CH2) 0-4 C(O)R°; -C(S)R°; -(CH2) 0-4 C(O)OR°; -(CH2) 0-4 C(O)SR°; -(CH2) 0-4 C(O)OSiR°3; -(CH2) 0-4 OC(O)R°; -OC(O)(CH2) 0-4 SR-; SC(S)SR°; -(CH2) 0-4 SC(O)R°; -(CH2) 0-4 C(O)NR°2; -C(S)NR°2; -C(S)SR°; -SC(S)SR°; -(CH2) 0-4 OC(O)NR°2; -C(O)N(OR°)R°; -C(O)C(O)R°; -C(O)CH2C(O)R°; -C(NOR°)R°; -(CH2) 0-4 SSR°; -(CH2) 0-4 S(O)2R°; -(CH2) 0-4 S(O)2OR°; -(CH2) 0-4OS(O)2R°; -S(O)2NR°2; -S(O)(NR°)R°; -S(O)2N=C(NR°2)2; -(CH2) 0-4 S(O)R°; -N(R°)S(O)2NR°2; -N(R°)S(O)2R°; -N(OR°)R°; -C(NH)NR°2; -P(O)2R°; -P(O)R°2; -OP(O)R°2; -OP(O)(OR°)2; SiR°3; -(C 1-4 straight, branched, or cyclic alkylene)O-N(R°)2; or -(C 1-4 straight, branched, or cyclic alkylene)C(O)O-N(R°)2.

[0044] each R° is independently hydrogen, C 1-6 aliphatic, -CH2Ph, -O(CH2) 0-1 Ph, -CH2-(5-6 membered heteroaryl ring), or a 5-6 membered saturated, partially unsaturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or, notwithstanding the definition above, two independent occurrences of R°, together with their intervening atoms, form a 3-12 membered saturated, partially unsaturated, or aryl monocyclic or bicyclic ring, which can have 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, and which can be substituted at a saturated carbon atom of R° with a divalent substituent selected from =O and =S; or each R° is optionally substituted with a monovalent substituent independently selected from the group consisting of halogen, -(CH2) 0-2 R · , -(haloR · ), -(CH2) 0-2 OH, -(CH2) 0-2 OR · , -(CH2) 0-2 CH(OR · )2, -O(haloR · ), -CN, -N3, -(CH2) 0-2 C(O)R · , -(CH2) 0-2 C(O)OH, -(CH2) 0-2 C(O)OR · , -(CH2) 0-2 SR · , -(CH2) 0- 2SH, -(CH2) 0-2 NH2, -(CH2) 0-2 NHR · , -(CH2) 0-2 NR · 2, -NO2, -SiR · 3, -OSiR ·3. -C(O)SR · -(C 1-4 (straight-chain or branched-chain alkylene)C(O)OR · or -SSR · .

[0045] Each R · Selected independently from C 1-4 Aliphatic, -CH2Ph, -O(CH2) 0-1 Ph or a 5-6 member saturated, partially unsaturated, or aryl ring having 0-4 independent heteroatoms selected from nitrogen, oxygen, or sulfur, wherein each R · It is either unsubstituted or substituted with one or more halogens if a halogen group is preceding it; or the optional substituents on the saturated carbon are independently selected from =O, =S, =NNR. * 2、=NNHC(O)R * =NNHC(O)OR * =NNHS(O)2R * =NR * =NOR * -O(C(R) * 2)) 2-3 O- or -S(C(R) * 2)) 2-3 The divalent substituent of S-, or the divalent substituent bonded to the ortho-substituted carbon of the "optionally substituted" group, is -O(CR). * 2) 2-3 O-, where each independently occurring R * Selected from hydrogen, C 1-6 Aliphatic or unsubstituted 5-6 member saturated, partially unsaturated or aryl rings having 0-4 heteroatoms independently selected from nitrogen, oxygen or sulfur.

[0046] When R * It is C 1-6 When aliphatic, R * Optional halogen, -R · -(halogenated R) · -OH, -OR · -O(halogenated R) · -CN, -C(O)OH, -C(O)OR · -NH2, -NHR · -NR · 2 or -NO2 substitution, where each R · Selected independently from C 1-4 Aliphatic, -CH2Ph, -O(CH2) 0-1Ph or a 5-6 membered saturated, partially saturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, and wherein each R · is unsubstituted or substituted with one or more halogen, provided that when substituted, the R

[0047] The optional substituents on the nitrogen can be independently or wherein each is independently hydrogen, C 1-6 aliphatic, unsubstituted -OPh, or unsubstituted 5-6 membered saturated, partially saturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, or two independent occurrences of R together with their intervening atoms form an unsubstituted 3-12 membered saturated, partially saturated, or aryl monocyclic or bicyclic ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur; wherein when is C 1-6 aliphatic, is optionally substituted with halogen, -R · , -(haloR · ), -OH, -OR · , -O(haloR · ), -CN, -C(O)OH, -C(O)OR · , -NH2, -NHR · , -NR · 2, or -NO2, wherein each R · is independently selected from C 1-4 aliphatic, -CH2Ph, -O(CH2) 0-1 Ph, or a 5-6 membered saturated, partially saturated, or aryl ring having 0-4 heteroatoms independently selected from nitrogen, oxygen, or sulfur, and wherein each R · is unsubstituted or substituted with one or more halogen, provided that when substituted, the R

[0048] As used herein, the term "pharmaceutically acceptable salt" means those salts which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of humans and lower animals without undue toxicity, irritation, allergic response and the like, and are commensurate with a reasonable benefit / risk ratio. Pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge et al., describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 1977, 66, 1-19, incorporated herein by reference. Pharmaceutically acceptable salts of the compounds of this application include those derived from the following suitable inorganic and organic acids: ammonium, boric, bicarbonic, carbonic, hydrochloric, hydrobromic, phosphoric, sulfuric, and perchloric acid, and the like, or from bases such as ammonium, benzathines, calcium, choline, diethanolamine, diethylamine, diphendylamine, glutathione, lithium, magnesium, nuphage, potassium, sodium, triethanolamine, tromethamine, and the like. Examples of pharmaceutically acceptable, nontoxic acid addition salts are salts of an amino group with inorganic acids such as hydrochloric, hydrobromic, phosphoric, sulfuric, and perchloric acid, or with organic acids such as acetic, oxalic, maleic, tartaric, citric, succinic, or malonic acid, or by using other methods used in the art such as ion exchange. Other pharmaceutically acceptable salts include adipate, alginate, ascorbate, aspartate, besylate, benzoate, bicarbonate, borate, butyrate, camphorate, camphorsulfonate, citrate, cyclopentanepropionate, digluconate, dodecylsulfate, ethanesulfonate, formate, fumarate, glucoheptanoate, glycerophosphate, gluconate, glutamate, glycolate, hemicellulose, heptanoate, hexanoate, hydroiodide, 2-hydroxyethanesulfonate, lactobionate, lactate, laurate, lauryl sulfate, malate, maleate, malonate, methanesulfonate, 2-naphthalenesulfonate, nicotinate, nitrate, oleate, oxalate, palmitate, pamoate, pectinate, persulfate, 3-phenylpropionate, phosphate, pivalate, propionate, stearate, succinate, sulfate, tartrate, thiocyanate, p-toluenesulfonate, undecanoate, valerate salts, and the like.

[0049] Further, it is recognized that salts of basic pharmaceutical compounds can be prepared as pharmaceutically acceptable salts from pharmaceutically acceptable acids, as discussed for example in P. Stahl et al., Camille G. (ed) Handbook of Pharmaceutical Salts. Properties, Selection and Use. (2002) Zurich: Wiley-VCH; S. Berge et al, Journal of Pharmaceutical Sciences (1977) 66(1) 1-19; P. Gould, International J. of Pharmaceutics (1986) 33201-217; Anderson et al., The Practice of Medicinal Chemistry (1996), Academic Press, New York; and The Orange Book (Food & Drug Administration, Washington, D.C. on its website). These disclosures are incorporated herein by reference.

[0050] Salts derived from appropriate bases include alkali metal, alkaline earth metal, ammonium and N + (C 1-4 Representative alkali or alkaline earth salts include sodium, lithium, potassium, calcium, magnesium, and the like. Further pharmaceutically acceptable salts include, when appropriate, nontoxic ammonium, quaternary ammonium, and amine cations formed using counterions such as halide, hydroxide, carboxylate, sulfate, phosphate, nitrate, lower

[0051] Unless otherwise stated, structures depicted herein are also meant to include all isomeric (e.g., enantiomeric, diastereomeric, and geometric (or conformational)) forms of the structure; for example, the R and S configurations for each asymmetric center, Z and E double bond isomers, and Z and E conformational isomers. Therefore, single stereochemical isomers as well as enantiomeric, diastereomeric, and geometric (or conformational) mixtures of the present compounds are within the scope of the application. Unless otherwise stated, all tautomeric forms of the compounds of the application are within the scope of the application. The application includes compounds that differ only in the presence of one or more isotopically enriched atoms. For example, compounds having hydrogen 13 C or 14 C enriched carbon substitutions are within the scope of this application. Such compounds are useful, for example, as analytical tools, as probes in biological assays, or as therapeutics in accordance with the application.

[0052] Diastereomeric mixtures can be separated into their individual diastereomers on the basis of their physical chemical differences by methods known to those skilled in the art, such as by chromatography and / or fractional crystallization. Enantiomers can be separated by converting the enantiomeric mixtures into a mixture of diastereomers by reaction with an appropriately optically active compound (e.g., a chiral alcohol or Mosher's acid chloride), separating the diastereomers and converting (e.g., hydrolyzing) the individual diastereomers to the corresponding pure enantiomers. Alternatively, specific enantiomers of the compounds of the present application can be prepared by asymmetric synthesis. Further, where the molecule contains a basic functionality (such as an amino group) or an acidic functionality (such as a carboxylic acid), diastereomeric salts are formed with an appropriately optically active acid or base, followed by resolution of the resulting diastereomers by fractional crystallization or chromatographic means known in the art, and subsequent recovery of the pure enantiomers.

[0053] Individual stereoisomers of the compounds of the present application can, for example, be substantially free of other isomers, or can be mixed with other or selected stereoisomers. A chiral center in a compound of the present application can have the S or R configuration as defined by the IUPAC 1974 Recommendations. In addition, to the extent that a compound described herein can exist in atropisomers (e.g., substituted biaryls), all forms of such atropisomers are considered within the scope of the present application.

[0054] Chemical names, common names, and chemical structures can be used interchangeably to describe the same structure. If both a chemical structure and a chemical name are used to refer to a chemical compound, and there is an ambiguity between the structure and the name, the structure is to control. It is also noted that any carbon and heteroatom possessing an unsatisfied valence in the text, schemes, examples and tables herein is assumed to have sufficient number and kind of hydrogen present to satisfy the valences.

[0055] As used herein the term "a" and "an" means "one or more" and includes the plural unless the context dictates otherwise.

[0056] The term "alkyl" refers to saturated straight or branched chain hydrocarbon, such as a straight or branched chain group of 1-12, 1-10, or 1-6 carbon atoms, referred to herein as C1-C12, C1-C10, or C1-C6alkyl, respectively. 12 alkyl, C1-C 10Alkyl and C1-C6alkyl. Exemplary alkyl groups include, but are not limited to, methyl, ethyl, propyl, isopropyl, 2-methyl-1 -propyl, 2-methyl-2-propyl, 2-methyl-1 -butyl, 3-methyl-1 -butyl, 2-methyl-3-butyl, 2,2-dimethyl-1 -propyl, 2-methyl-1 -pentyl, 3-methyl-1 -pentyl, 4-methyl-1 -pentyl, 2-methyl-2-pentyl, 3-methyl-2-pentyl, 4-methyl-2-pentyl, 2,2-dimethyl-1 -butyl, 3,3-dimethyl-1 -butyl, 2-ethyl-1 -butyl, butyl, isobutyl, t-butyl, pentyl, isopentyl, neopentyl, hexyl, heptyl, octyl, and the like.

[0057] The term "cycloalkyl" refers to a monovalent saturated cyclic, bicyclic, or bridged cyclic (e.g., adamantyl) hydrocarbon radical of 3-12, 3-8, 4-8, or 4-6 carbons, referred to herein as, for example, "C3-C6 cycloalkyl" derived from a cycloalkane. Exemplary cycloalkyl groups include cyclohexyl, cyclopentyl, cyclobutyl, and cyclopropyl. The term "cycloalkylene" refers to a divalent cycloalkyl group.

[0058] The term "haloalkyl" refers to an alkyl group substituted with at least one halogen. Exemplary haloalkyl groups include -CH2F, -CHF2, -CF3, -CH2CF3, -CF2CF3, and the like. The term "chloroalkyl" refers to an alkyl group substituted with at least one chlorine. The term "bromoalkyl" refers to an alkyl group substituted with at least one bromine. The term "haloalkylene" refers to a divalent haloalkyl group.

[0059] The term "hydroxyalkyl" refers to an alkyl group substituted with at least one hydroxyl group. Exemplary hydroxyalkyl groups include -CH2CH2OH, -C(H)(OH)CH3, -CH2C(H)(OH)CH2CH2OH, and the like.

[0060] The term "heteroalkyl" refers to an alkyl group in which one or more carbon atoms are replaced with a heteroatom (e.g., N, O, or S). Exemplary heteroalkyl groups include -OCH3, -CH2OCH3, -CH2CH2N(CH3)2, and -CH2CH2OH. A heteroalkyl group can contain, for example, 2-4, 2-6, or 2-8 atoms selected from the group consisting of carbon and a heteroatom (e.g., N, O, or S). The phrase 3-8 membered heteroalkyl refers to a heteroalkyl group having 3 to 8 atoms selected from the group consisting of carbon and a heteroatom. The term "heteroalkylene" refers to a divalent heteroalkyl group.

[0061] The terms "alkenyl" and "alkynyl" are art-recognized and refer to unsaturated aliphatic groups analogous in length and possible substitution to the alkyl radicals described above, but that contain at least one double or triple bond respectively. The term "haloalkenyl" refers to an alkenyl group substituted with at least one halogen. The term "fluoroalkenyl" refers to an alkenyl group substituted with at least one fluorine. The term "nitroalkenyl" refers to an alkenyl group substituted with at least one nitro group.

[0062] The term "carbocyclyl" refers to a saturated or unsaturated, cyclic hydrocarbon group.

[0063] The term "alkoxyl" or "alkoxy" is art-recognized and refers to an alkyl group as previously defined having an oxy group attached thereto. Representative alkoxyl groups include methoxy, ethoxy, propyloxy, tert-butyloxy, and the like. The term "haloalkoxyl" refers to an alkoxyl group that is substituted with at least one halogen. Exemplary haloalkoxyl groups include -OCH2F, -OCHF2, -OCF3, -OCH2CF3, -OCF2CF3, and the like.

[0064] The term "oxo" is art-recognized and refers to a "=0" substituent. For example, cyclopentane substituted with an oxo group is cyclopentanone.

[0065] The term "amino" is art-recognized and refers to unsubstituted and substituted amines, for example, moieties that can be represented by the following general formula:

[0066]

[0067] wherein R 50 , R 51 , R 52 and R 53 each independently represent hydrogen, alkyl, alkenyl, -(CH2) m -R 61 , or R 50 and R 51 together with the N atom to which they are attached form a heterocyclic ring having 4 to 8 atoms in the ring structure; R 61 represents aryl, 3-7 membered cycloalkyl, 4-7 membered cycloalkenyl, 5-10 membered heteroaryl, or 3-10 membered heterocyclyl; and m is 0 or an integer in the range of 1 to 8.

[0068] The term "amido" is art-recognized and refers to unsubstituted and substituted amides, for example, moieties that can be represented by the following general formula:

[0069]

[0070] wherein R 50 and R 51 each independently represent hydrogen, alkyl, alkenyl, -(CH2) m -R 61 , or R 50 and R 51 together with the N atom to which they are attached form a heterocyclic ring having 4 to 8 atoms in the ring structure; R 61represents aryl, 3-7 membered cycloalkyl, 4-7 membered cycloalkenyl, 5-10 membered heteroaryl, or 3-10 membered heterocyclyl; and m is 0 or an integer in the range of 1 to 8; and R 52 is alkyl, alkenyl, or -(CH2) m -R 61 .

[0071] The symbol indicates a point of attachment.

[0072] Unless otherwise indicated, when any substituent or variable occurs multiple times in any constituent or compound of the present application, its definition in each occurrence is independent of any other occurrence.

[0073] One or more compounds of the present application can exist in unsolvated as well as solvated forms with pharmaceutically acceptable solvents such as water, ethanol, and the like, and it is intended that the application embrace both solvated and unsolvated forms. "Solvate" means a physical association between one or more solvent molecules and one or more of the compounds of the present application. This physical association can occur intramolecularly, intermolecularly, or intramolecularly and intermolecularly. In certain instances the association is such that the

[0074] As used herein, the terms "subject" and "patient" are used interchangeably and refer to an organism to be treated by the methods of the present application. Such organisms preferably include, but are not limited to, mammals (e.g., murines, simians, equines, bovines, porcines, canines, felines, and the like), and most preferably include humans.

[0075] The term "IC 50 " is art-recognized and refers to the concentration of a compound required for 50% inhibition of a target.

[0076] As used herein, the term "effective amount" means the amount of a compound sufficient to effect a beneficial or desired result (e.g., a therapeutic, ameliorative, inhibitory, or prophylactic result). An effective amount can be administered in one or more administrations, applications or dosages and is not intended to be limited to a particular formulation or administration route. As used herein, the term "treatment" includes any effect that alleviates, reduces, modulates, ameliorates, or eliminates a condition, disease, disorder, etc., or its symptoms.

[0077] As used herein, the term "pharmaceutical composition" refers to the combination of an active agent with a carrier, inert or active, making the composition especially suitable for diagnostic or therapeutic use in vivo or ex vivo.

[0078] As used herein, the term "pharmaceutically acceptable carrier" refers to any of the standard pharmaceutical carriers such as phosphate buffered saline solution, water, emulsions such as an oil / water or water / oil emulsion, and various types of wetting agents. The composition can also include stabilizers and preservatives. For examples of carriers, stabilizers, and adjuvants, see, e.g., Martin, Remington's Pharmaceutical Sciences, 15th Ed., Mack Publ. Co., Easton, PA

[1975] .

[0079] For therapeutic use, salts of the compounds of the application are contemplated as being pharmaceutically acceptable. However, salts of acids and bases that are non- pharmaceutically acceptable can also be used, for example, in the preparation or purification of a pharmaceutically acceptable compound.

[0080] Additionally, when the compounds of the application contain both a basic moiety, such as, but not limited to, a pyridine or imidazole, and an acidic moiety, such as, but not limited to, a carboxylic acid, zwitterions ("inner salts") can be formed. Such acidic salts and basic salts used within the scope of this application are pharmaceutically acceptable (i.e., non-toxic, physiologically acceptable) salts. Such salts of the compounds of the application can be formed, for example, by reaction of a compound of the application with an amount of acid or base, such as an equivalent amount, in a medium such as one in which the salt precipitates or in an aqueous medium, followed by lyophilization.

[0081] Throughout the description, where compositions are described as having, including, or comprising specific components, or where processes and methods are described as having, including, or comprising specific steps, it is contemplated that, additionally, there are compositions of the present application that consist essentially of, or consist of, the recited components, and that there are processes and methods according to the present application that consist essentially of, or consist of, the recited processing steps.

[0082] In general, percentages of compositions are specified by weight unless otherwise stated.

[0083] I. Heterobifunctional compounds

[0084] One aspect of the application provides heterobifunctional compounds. The compounds can be used in the pharmaceutical compositions and therapeutic methods described herein. Exemplary compounds, along with exemplary procedures for preparing the compounds, are described in the following sections.

[0085] Section A: Crystalline forms of compounds of Formula II-5

[0086] One aspect of the application provides a crystalline compound of Formula II-5:

[0087]

[0088] Crystalline forms of the compound of Formula II-5 can provide properties that facilitate the compound's suitability for manufacture. For example, crystalline Form A of the compound of Formula II-5 has been identified that has excellent properties. One benefit of crystalline Form A is that it exhibits superior stability relative to crystalline Form B.

[0089] Crystalline Form A

[0090] One aspect of the present application provides a crystalline compound of Formula II-5 having crystalline Form A. Crystalline Form A can be characterized by X-ray powder diffraction, differential scanning calorimetry, and / or purity.

[0091] In certain embodiments, the compound exhibits an X-ray powder diffraction pattern comprising peaks at the following diffraction angles (2Q): 13.1 ± 0.2, 17.6 ± 0.2, 18.3 ± 0.2, 18.7 ± 0.2, 19.5 ± 0.2, 23.8 ± 0.2, and 25.6 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 8.1 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 9.9 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 10.5 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 15.8 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 19.8 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 25.3 ± 0.2.

[0092] In certain embodiments, the compound exhibits an X-ray powder diffraction pattern comprising at least one peak from the following diffraction angles (2q): 13.1 ± 0.2, 17.6 ± 0.2, 18.3 ± 0.2, 18.7 ± 0.2, 19.5 ± 0.2, 23.8 ± 0.2, and 25.6 ± 0.2. In certain embodiments, the compound exhibits an X-ray powder diffraction pattern comprising at least two, three, or four peaks from the following diffraction angles (2q): 13.1 ± 0.2, 17.6 ± 0.2, 18.3 ± 0.2, 18.7 ± 0.2, 19.5 ± 0.2, 23.8 ± 0.2, and 25.6 ± 0.2. In certain embodiments, the compound exhibits an X-ray powder diffraction pattern comprising at least five peaks from the following diffraction angles (2q): 13.1 ± 0.2, 17.6 ± 0.2, 18.3 ± 0.2, 18.7 ± 0.2, 19.5 ± 0.2, 23.8 ± 0.2, and 25.6 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 8.1 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 9.9 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 10.5 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 15.8 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 19.8 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 25.3 ± 0.2.

[0093] In certain embodiments, the relative intensity of the peak at the diffraction angle (2q) is at least 20%. In certain embodiments, the relative intensity of the peak at the diffraction angle (2q) is at least 30%.

[0094] In certain embodiments, the compound is characterized by the following X-ray powder diffraction pattern expressed in terms of diffraction angles 2q, interplanar spacing d, and relative intensities (expressed as a percentage relative to the most intense peak):

[0095]

[0096]

[0097] In certain embodiments, the compound has an X-ray powder diffraction pattern substantially as shown in Figure 1

[0098] ​In certain embodiments, the compound has a melting point onset in the range of about 180 °C to about 200 °C as determined by differential scanning calorimetry. In certain embodiments, the compound has a melting point onset of about 192 °C as determined by differential scanning calorimetry. In certain embodiments, the compound has a melting point onset of 192 °C as determined by differential scanning calorimetry. In certain embodiments, the compound has substantially the same differential scanning calorimetry profile as shown in FIG. 2. Figure 2

[0099] In certain embodiments, the compound has a purity of greater than 98% by weight. In certain embodiments, the compound has a purity of greater than 99% by weight. In certain embodiments, the compound has a purity of greater than 99.5% by weight. In certain embodiments, the purity of the compound is determined by high performance liquid chromatography (HPLC).

[0100] Crystalline Form B

[0101] Another aspect of the application provides a crystalline compound of Formula II-5 having crystalline Form B. Crystalline Form B can be characterized by X-ray powder diffraction, differential scanning calorimetry, and / or purity.

[0102] In certain embodiments, the compound exhibits an X-ray powder diffraction pattern comprising peaks at the following diffraction angles (2Q): 9.4 ± 0.2, 12.8 ± 0.2, 15.4 ± 0.2, 18.2 ± 0.2, 18.7 ± 0.2, 20.2 ± 0.2, and 24.6 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises peaks at one or more of the following diffraction angles (2Q): 8.5 ± 0.2, 11.7 ± 0.2, 12.5 ± 0.2, 17.6 ± 0.2, 19.6 ± 0.2, 21.9 ± 0.2, and 23.5 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 8.5 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 11.7 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 12.5 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 17.6 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 19.6 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 21.9 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 23.5 ± 0.2.

[0103] ​In certain embodiments, the compound exhibits an X-ray powder diffraction pattern comprising at least one peak from the following diffraction angles (2q): 9.4 ± 0.2, 12.8 ± 0.2, 15.4 ± 0.2, 18.2 ± 0.2, 18.7 ± 0.2, 20.2 ± 0.2, and 24.6 ± 0.2. In certain embodiments, the compound exhibits an X-ray powder diffraction pattern comprising at least two, three, or four peaks from the following diffraction angles (2q): 9.4 ± 0.2, 12.8 ± 0.2, 15.4 ± 0.2, 18.2 ± 0.2, 18.7 ± 0.2, 20.2 ± 0.2, and 24.6 ± 0.2. In certain embodiments, the compound exhibits an X-ray powder diffraction pattern comprising at least five peaks from the following diffraction angles (2q): 9.4 ± 0.2, 12.8 ± 0.2, 15.4 ± 0.2, 18.2 ± 0.2, 18.7 ± 0.2, 20.2 ± 0.2, and 24.6 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at one or more of the following diffraction angles (2q): 8.5 ± 0.2, 11.7 ± 0.2, 12.5 ± 0.2, 17.6 ± 0.2, 19.6 ± 0.2, 21.9 ± 0.2, and 23.5 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 8.5 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 11.7 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 12.5 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 17.6 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 19.6 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 21.9 ± 0.2. In certain embodiments, the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2q): 23.5 ± 0.2.

[0104] In certain embodiments, the relative intensity of the peak at the diffraction angle (2q) is at least 20%. In certain embodiments, the relative intensity of the peak at the diffraction angle (2q) is at least 30%.

[0105] In certain embodiments, the compound is characterized by the following X-ray powder diffraction pattern expressed in terms of diffraction angles 2q, interplanar spacing d, and relative intensity (expressed as a percentage relative to the strongest peak):

[0106]

[0107]

[0109] In certain embodiments, the compound has an X-ray powder diffraction pattern substantially as shown in FIG. 2. Figure 4

[0110] In certain embodiments, the compound has a melting point onset in the range of about 180 °C to about 200 °C as determined by differential scanning calorimetry. In certain embodiments, the compound has a melting point onset of about 192 °C as determined by differential scanning calorimetry. In certain embodiments, the compound has a melting point onset of 192 °C as determined by differential scanning calorimetry. In certain embodiments, the compound has a differential scanning calorimetry curve substantially the same as shown in FIG. 3. Figure 5

[0111] In certain embodiments, the compound has a purity greater than 98% by weight. In certain embodiments, the compound has a purity greater than 99% by weight. In certain embodiments, the compound has a purity greater than 99.5% by weight. In certain embodiments, the purity of the compound is determined by high performance liquid chromatography (HPLC).

[0112] The above description describes various embodiments relating to crystalline forms of the compound of Formula II-5. The present patent application specifically encompasses all combinations of the described embodiments.

[0113] Part B: Additional Heterobifunctional Compounds

[0114] Another aspect of the present application provides a compound of Table 3, or a pharmaceutically acceptable salt thereof:

[0115] Table 3.

[0116]

[0117] In certain embodiments, the compound is a compound of Table 3.

[0118] II. Therapeutic Applications

[0119] ​​The heterobifunctional compounds described herein, such as the crystalline compound of Formula II-5 or the compounds in Table 3, or other compounds in Section I, or a pharmaceutically acceptable salt thereof, provide therapeutic benefit to patients suffering from cancer. Accordingly, one aspect of the present application provides a method of treating cancer. The method includes administering to a patient in need thereof a therapeutically effective amount of a compound described herein, such as the crystalline compound of Formula II-5 or the compounds in Table 3, or other compounds in Section I, or a pharmaceutically acceptable salt thereof, to treat cancer. In certain embodiments, the compound is the crystalline compound of Formula II-5. In certain embodiments, the particular crystalline compound of Formula II-5 is the compound defined by one of the embodiments described above. In certain embodiments, the compound is a compound in Table 3, or a pharmaceutically acceptable salt thereof. In certain embodiments, the compound is a compound in Table 3.

[0120] Cancer

[0121] In certain embodiments, the cancer is ovarian cancer, uterine cancer, endometrial cancer, cervical cancer, prostate cancer, testicular cancer, breast cancer, brain cancer, lung cancer, oral cancer, esophageal cancer, head and neck cancer, stomach cancer, colon cancer, rectal cancer, skin cancer, sebaceous gland cancer, bile duct and gall bladder cancer, liver cancer, pancreatic cancer, bladder cancer, urinary tract cancer, kidney cancer, eye cancer, thyroid cancer, lymphoma, or leukemia. In certain embodiments, the cancer is prostate cancer.

[0122] In certain embodiments, the cancer is squamous cell carcinoma, lung cancer (including small cell lung cancer, non-small cell lung cancer), vulvar cancer, thyroid cancer, lung adenocarcinoma and lung squamous carcinoma, peritoneal cancer, hepatocellular cancer, gastric cancer (including gastrointestinal cancer), pancreatic cancer, glioblastoma, cervical cancer, ovarian cancer, liver cancer, bladder cancer, hepatoma, breast cancer, colon cancer, rectal cancer, colorectal cancer, endometrial or uterine cancer, salivary gland cancer, kidney cancer, prostate cancer, liver cancer, anal cancer, penile cancer, and head and neck cancer. In certain embodiments, the cancer is at least one selected from the group consisting of ALL, T-lineage acute lymphoblastic leukemia (T-ALL), T-lineage lymphoblastic lymphoma (T-LL), peripheral T-cell lymphoma, adult T-cell leukemia, Pre-B ALL, Pre-B lymphoma, large B-cell lymphoma, Burkitts Lymphoma, B-cell ALL, Philadelphia chromosome positive ALL, Philadelphia chromosome positive CML, lymphoma, leukemia, multiple myeloma, myeloproliferative disease, large B-cell lymphoma, or B-cell lymphoma.

[0123] In certain embodiments, the cancer is a solid tumor or a leukemia. In certain other embodiments, the cancer is colon cancer, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, lung cancer, leukemia, bladder cancer, stomach cancer, cervical cancer, testicular cancer, skin cancer, rectal cancer, thyroid cancer, kidney cancer, uterine cancer, esophageal cancer, liver cancer, acoustic neuroma, oligodendroglioma, meningioma, neuroblastoma, or retinoblastoma. In certain other embodiments, the cancer is small cell lung cancer, non-small cell lung cancer, melanoma, cancer of the central nervous system tissue, brain cancer, Hodgkin's lymphoma, non-Hodgkin's lymphoma, cutaneous T-cell lymphoma, cutaneous B-cell lymphoma, or diffuse large B-cell lymphoma. In certain other embodiments, the cancer is breast cancer, colon cancer, small cell lung cancer, non-small cell lung cancer, prostate cancer, kidney cancer, ovarian cancer, leukemia, melanoma, or cancer of the central nervous system tissue. In certain other embodiments, the cancer is colon cancer, small cell lung cancer, non-small cell lung cancer, kidney cancer, ovarian cancer, kidney cancer, or melanoma.

[0124] In certain embodiments, the cancer is fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinoma, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilms' tumor, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, or hemangioblastoma.

[0125] In certain embodiments, the cancer is neuroblastoma, meningioma, hemangiopericytoma, multiple brain metastases, glioblastoma multiforme, glioblastoma, brain stem glioma, poor prognosis malignant brain tumors, malignant gliomas, anaplastic astrocytoma, anaplastic oligodendroglioma, neuroendocrine tumors, rectal adenocarcinoma, Dukes C and D colorectal cancer, inoperable colorectal cancer, metastatic hepatocellular carcinoma, Kaposi's sarcoma, karyotypically acute myeloblastoma, Hodgkin's lymphoma, non-Hodgkin's lymphoma, cutaneous T-cell lymphoma, cutaneous B-cell lymphoma, diffuse large B-cell lymphoma, low grade follicular lymphoma, metastatic melanoma, localized melanoma, malignant mesothelioma, malignant pleural effusion mesothelioma syndrome, peritoneal carcinoma, papillary serous carcinoma, gynecological sarcomas, soft tissue sarcomas, scleroderma, cutaneous vasculitis, Langerhans cell histiocytosis, leiomyosarcoma, fibrodysplasia ossificans progressiva, hormone refractory prostate cancer, high risk of resection of soft tissue sarcoma, inoperable hepatocellular carcinoma, Waidenstrom's macroglobulinemia, smoldering myeloma, indolent myeloma, fallopian tube cancer, androgen independent prostate cancer, androgen dependent stage IV non-metastatic prostate cancer, hormone non-sensitive prostate cancer, chemotherapy non-sensitive prostate cancer, castration resistant prostate cancer, castration resistant metastatic prostate cancer, papillary thyroid cancer, follicular thyroid cancer, medullary thyroid cancer, or leiomyoma.

[0126] In certain embodiments, the cancer is bone cancer, pancreatic cancer, skin cancer, cancer of the head or neck, cutaneous or intraocular melanoma, ovarian cancer, colon cancer, rectal cancer, cancer of the anal region, stomach cancer, gastrointestinal (gastric, colorectal, and duodenal), uterine cancer, carcinoma of the fallopian tubes, carcinoma of the endometrium, carcinoma of the cervix, carcinoma of the vagina, carcinoma of the vulva, Hodgkin's Disease, cancer of the esophagus, cancer of the small intestine, cancer of the endocrine system, cancer of the thyroid gland, cancer of the parathyroid gland, cancer of the adrenal gland, sarcoma of soft tissue, cancer of the urethra, cancer of the penis, prostate cancer, testicular cancer, chronic or acute leukemia, chronic myeloid leukemia, lymphocytic lymphoma, bladder cancer, cancer of the kidney or ureter, renal cell carcinoma, carcinoma of the renal pelvis, non-Hodgkin's lymphoma, neoplasms of the spinal axis, brain stem glioma, pituitary adenoma, adrenocortical cancer, gall bladder cancer, multiple myeloma, cholangiocarcinoma, fibrosarcoma, neuroblastoma, retinoblastoma, or a combination of one or more of the foregoing cancers.

[0127] In certain embodiments, the cancer is hepatocellular carcinoma, ovarian cancer, ovarian epithelial cancer, or fallopian tube cancer; papillary serous cystadenocarcinoma or uterine papillary serous carcinoma (UPSC); prostate cancer; testicular cancer; gallbladder cancer; cholangiocellular carcinoma of the liver; soft tissue and bone synovial sarcoma; rhabdomyosarcoma; osteosarcoma; chondrosarcoma; Ewing sarcoma; anaplastic thyroid cancer; adrenocortical adenoma; pancreatic cancer; pancreatic ductal carcinoma or pancreatic adenocarcinoma; gastrointestinal / stomach (GIST) cancer; lymphoma; squamous cell carcinoma of the head and neck (SCCHN); salivary gland cancer; glioma or brain cancer; neurofibromatosis-1 associated malignant peripheral nerve sheath tumor (MPNST); Waldenstrom macroglobulinemia; or medulloblastoma.

[0128] In certain embodiments, the cancer is hepatocellular carcinoma (HCC), hepatoblastoma, colon cancer, rectal cancer, ovarian cancer, ovarian epithelial cancer, fallopian tube cancer, papillary serous cystadenocarcinoma, uterine papillary serous carcinoma (UPSC), cholangiocellular carcinoma of the liver, soft tissue and bone synovial sarcoma, rhabdomyosarcoma, osteosarcoma, anaplastic thyroid cancer, adrenocortical adenoma, pancreatic cancer, pancreatic ductal carcinoma, pancreatic adenocarcinoma, glioma, neurofibromatosis-1 associated malignant peripheral nerve sheath tumor (MPNST), Waldenstrom macroglobulinemia, or medulloblastoma.

[0129] In certain embodiments, the cancer is a solid tumor, such as a sarcoma, a carcinoma, or a lymphoma. In certain embodiments, the cancer is renal cancer; hepatocellular carcinoma (HCC) or hepatoblastoma or liver cancer; melanoma; breast cancer; colorectal carcinoma or colorectal cancer; colon cancer; rectal cancer; anal cancer; lung cancer, such as non-small cell lung cancer (NSCLC) or small cell lung cancer (SCLC); ovarian cancer, ovarian epithelial cancer, ovarian carcinoma, or fallopian tube cancer; papillary serous cystadenocarcinoma or uterine papillary serous carcinoma (UPSC); prostate cancer; testicular cancer; gallbladder cancer; cholangiocellular carcinoma of the liver; soft tissue and bone synovial sarcoma; rhabdomyosarcoma; osteosarcoma; chondrosarcoma; Ewing sarcoma; anaplastic thyroid cancer; adrenocortical carcinoma; pancreatic cancer; pancreatic ductal carcinoma or pancreatic adenocarcinoma; gastrointestinal / stomach (GIST) cancer; lymphoma; squamous cell carcinoma of the head and neck (SCCHN); salivary gland cancer; glioma or brain cancer; neurofibromatosis-1 associated malignant peripheral nerve sheath tumor (MPNST); Waldenstrom macroglobulinemia; or medulloblastoma.

[0130] In certain embodiments, the cancer is a renal cell carcinoma, hepatocellular carcinoma (HCC), hepatoblastoma, colorectal carcinoma, colorectal cancer, colon cancer, rectal cancer, anal cancer, ovarian cancer, ovarian epithelial cancer, ovarian carcinoma, fallopian tube cancer, papillary serous cystadenocarcinoma, uterine papillary serous carcinoma (UPSC), cholangiocarcinoma, soft tissue and bone synovial sarcoma, rhabdomyosarcoma, osteosarcoma, chondrosarcoma, anaplastic thyroid cancer, adrenocortical carcinoma, pancreatic cancer, pancreatic ductal carcinoma, pancreatic adenocarcinoma, glioma, brain cancer, neurofibromatosis-1 associated malignant peripheral nerve sheath tumor (MPNST), Waldenstrom macroglobulinemia, or medulloblastoma.

[0131] In certain embodiments, the cancer is a hepatocellular carcinoma (HCC), hepatoblastoma, colon cancer, rectal cancer, ovarian cancer, ovarian epithelial cancer, ovarian carcinoma, fallopian tube cancer, papillary serous cystadenocarcinoma, uterine papillary serous carcinoma (UPSC), cholangiocarcinoma, soft tissue and bone synovial sarcoma, rhabdomyosarcoma, osteosarcoma, anaplastic thyroid cancer, adrenocortical carcinoma, pancreatic cancer, pancreatic ductal carcinoma, pancreatic adenocarcinoma, glioma, neurofibromatosis-1 associated malignant peripheral nerve sheath tumor (MPNST), Waldenstrom macroglobulinemia, or medulloblastoma.

[0132] In certain embodiments, the cancer is a hepatocellular carcinoma (HCC). In some embodiments, the cancer is a hepatoblastoma. In some embodiments, the cancer is colon cancer. In some embodiments, the cancer is rectal cancer. In some embodiments, the cancer is ovarian cancer or ovarian carcinoma. In some embodiments, the cancer is ovarian epithelial cancer. In some embodiments, the cancer is fallopian tube cancer. In some embodiments, the cancer is papillary serous cystadenocarcinoma. In some embodiments, the cancer is uterine papillary serous carcinoma (UPSC). In some embodiments, the cancer is cholangiocarcinoma. In some embodiments, the cancer is soft tissue and bone synovial sarcoma. In some embodiments, the cancer is rhabdomyosarcoma. In some embodiments, the cancer is osteosarcoma. In some embodiments, the cancer is anaplastic thyroid cancer. In some embodiments, the cancer is adrenocortical carcinoma. In some embodiments, the cancer is pancreatic cancer or pancreatic ductal carcinoma. In some embodiments, the cancer is pancreatic adenocarcinoma. In some embodiments, the cancer is glioma. In some embodiments, the cancer is malignant peripheral nerve sheath tumor (MPNST). In some embodiments, the cancer is neurofibromatosis-1 associated MPNST. In some embodiments, the cancer is Waldenstrom macroglobulinemia. In some embodiments, the cancer is medulloblastoma.

[0133] Causing death of cancer cells

[0134] Another aspect of the application provides a method of causing cancer cell death. The method comprises contacting a cancer cell with an effective amount of a compound described herein, such as a crystalline compound of Formula II-5 or a compound in Table 3, or a pharmaceutically acceptable salt thereof, or other compounds in Section I, to cause cancer cell death. In certain embodiments, the compound is a crystalline compound of Formula II-5. In certain embodiments, the particular crystalline compound of Formula II-5 is a compound defined by one of the embodiments described above. In certain embodiments, the compound is a compound in Table 3, or a pharmaceutically acceptable salt thereof. In certain embodiments, the compound is a compound in Table 3.

[0135] In certain embodiments, the cancer cell is selected from ovarian cancer, uterine cancer, endometrial cancer, cervical cancer, prostate cancer, testicular cancer, breast cancer, brain cancer, lung cancer, oral cancer, esophageal cancer, head and neck cancer, stomach cancer, colon cancer, rectal cancer, skin cancer, sebaceous gland cancer, bile duct and gall bladder cancer, liver cancer, pancreatic cancer, bladder cancer, urinary tract cancer, kidney cancer, eye cancer, thyroid cancer, lymphoma, or leukemia. In certain embodiments, the cancer cell is one or more of the cancers recited in the section above entitled "Cancers." In certain embodiments, the cancer cell is a prostate cancer cell.

[0136] Exemplary bioactivity assays

[0137] The biological activity of the compounds described herein can be evaluated using the assays described herein.

[0138] A. Assay for binding affinity to androgen receptor

[0139] Compounds can be tested for their ability to bind to the androgen receptor using the following procedure. The cytosolic fraction (106 cells / point) is incubated with 1 nM [3H]R1881 in the absence or presence of test compound in a buffer containing 25 mM Hepes-Tris (pH 7.4), 1 mM EDTA, 10 mM Na2Mo04, 2 mM DTT, 5 μM triamcinolone acetonide, and 10% glycerol at 4°C. After 2 hours, the incubation is terminated by the addition of 10 volumes of ice-cold buffer containing 50 mM Tris-HCl (pH 7.4), 1 mM EDTA, 0.1% BSA, and 10% glycerol. The cells are then filtered through Whatman GF / C filters, and the filters are washed with 10 volumes of ice-cold buffer. The filters are then dried and counted in a liquid scintillation counter. The IC50 value is calculated as the concentration of test compound that inhibits the binding of [3H]R1881 to the androgen receptor by 50%. 3H] Testosterone was incubated for 24 hours. Non-specific binding can be determined in the presence of 1 μM testosterone. After incubation, samples were rapidly filtered under vacuum through 0.3% PEI pre- soaked glass fiber filters (GF / B, Packard) and rinsed several times with ice-cold 50 mM Tris-HCl using a 96-sample cell harvester (Unifilter, Packard). Filters were dried and then counted for radioactivity in a scintillation counter (Topcount, Packard) using scintillation cocktail (Microscint 0, Packard). Results were expressed as percent inhibition of specific binding of the radioligand versus control. The standard reference compound was testosterone, which was tested at several concentrations in each experiment to obtain a competition curve from which the IC50value was calculated. 50

[0140] B. Determination of binding affinity to BRD4-BD1

[0141] ​The following experimental procedure can be used to test the ability of a compound to bind to BRD4-BD1. Compounds can be tested using the bromoKdELECT assay. A T7 phage strain displaying bromodomain is grown in parallel in E. coli hosts derived from BL21 strain in 24-well blocks. E. coli is grown to log phase and infected with T7 phage from frozen stocks (multiplicity of infection = 0.4) and incubated at 32°C with shaking until lysis (90-150 minutes). The lysate is centrifuged (5,000 x g) and filtered (0.2 μm) to remove cell debris. Streptavidin-coated magnetic beads are treated with biotinylated small molecule or acetylated peptide ligand for 30 minutes at room temperature to generate affinity resin for bromodomain assay. Ligandized beads are blocked with excess biotin and washed with blocking buffer (SeaBlock (Pierce), 1% BSA, 0.05% Tween 20, 1 mM DTT) to remove unbound ligand and reduce non-specific phage binding. Binding reactions are assembled by combining bromodomain binding domain, ligandized affinity beads, and test compound in 1 x binding buffer (17% SeaBlock, 0.33 x PBS, 0.04% Tween 20, 0.02% BSA, 0.004% sodium azide, 7.4 mM DTT). Test compounds are prepared as 1000 x stocks in 100% DMSO. An 11-point 3-fold dilution series of compound is used to determine Kd, with one DMSO control point. All compounds for Kd measurement are distributed in 100% DMSO by acoustic transfer (non-contact dispensing). Compounds are then diluted directly into the assay such that the final concentration of DMSO is 0.09%. All reactions can be performed in polypropylene 384-well plates. Each reaction has a final volume of 0.02 ml. Assay plates are incubated at room temperature with shaking for 1 hour and the affinity beads are washed with wash buffer (1 x PBS, 0.05% Tween 20). Beads are then resuspended in elution buffer (1 x PBS, 0.05% Tween 20, 2 μM non-biotinylated affinity ligand) and incubated at room temperature with shaking for 30 minutes. Bromodomain concentration in the eluate can be measured by qPCR.

[0142] C. Assay of binding affinity to BRD4-BD2

[0143] Compounds can be tested for their ability to bind to BRD4-BD2 using the following experimental procedure. Compounds can be tested using the bromoKdELECT assay. A T7 phage strain displaying bromodomain is grown in parallel in E. coli hosts derived from BL21 strain in 24-well blocks. E. coli is grown to log phase and infected with T7 phage from frozen stocks (multiplicity of infection = 0.4) and incubated at 32°C with shaking until lysis (90-150 minutes). The lysate is centrifuged (5,000 x g) and filtered (0.2 μιη) to remove cell debris. Streptavidin-coated magnetic beads are treated with biotinylated small molecule or acetylated peptide ligand for 30 minutes at room temperature to generate affinity resin for bromodomain assay. Ligandized beads are blocked with excess biotin and washed with blocking buffer (SeaBlock (Pierce), 1% BSA, 0.05% Tween 20, 1 mM DTT) to remove unbound ligand and reduce non-specific phage binding. Binding reactions are assembled by combining bromo-binding domain, ligandized affinity beads, and test compound in 1 x binding buffer (17% SeaBlock, 0.33 x PBS, 0.04% Tween 20, 0.02% BSA, 0.004% sodium azide, 7.4 mM DTT). Test compounds are prepared as 1000X stocks in 100% DMSO. An 11-point 3-fold compound dilution series is used to determine Kd, with one DMSO control point. All compounds for Kd measurement are distributed in 100% DMSO by acoustic transfer (non-contact dispensing). Compounds are then diluted directly into the assay such that the final concentration of DMSO is 0.09%. Reactions can be performed in polypropylene 384-well plates. Each reaction has a final volume of 0.02 ml. Assay plates are incubated at room temperature with shaking for 1 hour and the affinity beads are washed with wash buffer (1 x PBS, 0.05% Tween 20). Beads are then resuspended in elution buffer (1 x PBS, 0.05% Tween 20, 2 μΜ non-biotinylated affinity ligand) and incubated at room temperature with shaking for 30 minutes. Bromodomain concentration in the eluate can be measured by qPCR.

[0144] Combination therapy

[0145] Compounds useful in the methods of the present application can be used in combination with one or more additional therapeutic agents useful in the treatment of any of the diseases contemplated herein. These additional therapeutic agents can include compounds that are commercially available or that can be synthesized by one of skill in the art. These additional therapeutic agents are known to treat, prevent, or ameliorate the symptoms of the diseases or conditions contemplated herein.

[0146] Thus, in certain embodiments, the methods further comprise administering to the subject an additional therapeutic agent that treats a disease contemplated herein.

[0147] In certain embodiments, administration of a compound of the application to a subject allows for a lower dose of an additional therapeutic agent to be administered than would be required to achieve a similar result in treating a disease contemplated herein. For example, in certain embodiments, a compound of the application enhances the therapeutic activity of an additional therapeutic compound, thereby allowing a lower dose of the additional therapeutic compound to provide the same effect.

[0148] Synergy can be calculated, for example, using suitable methods such as Sigmoidal- E max equation (Holford and Scheiner, 1981, Clin. Pharmacokinet. 6:429-453), the Loewe additivity equation (Loewe and Muischnek, 1926, Arch. Exp. Pathol Pharmacol. 114:313-326), and the median-effect equation (Chou and Talalay, 1984, Adv. Enzyme Regul. 22:27-55). Each of the above-mentioned equations can be applied to experimental data to generate a corresponding graph to aid in the assessment of the effect of a drug combination. The corresponding graphs associated with the above equations are concentration-effect curves, isobologram curves, and combination index curves, respectively.

[0149] In certain embodiments, a compound of the application and a therapeutic agent are coadministered to a subject. In other embodiments, a compound of the application and a therapeutic agent are co-formulated and co-administered to a subject.

[0150] In certain embodiments, the compound is administered in combination with a second therapeutic agent having anti-cancer activity. In certain embodiments, the second therapeutic agent is mitomycin, tretinoin, ribomustin, gemcitabine, vincristine, etoposide, cladribine, mitobronitol, methotrexate, doxorubicin, carboquone, pentostatin, nitracrine, zinostatin, cetrorelix, letrozole, raltitrexed, daunorubicin, fadrozole, fotemustine, thymalfasin, sobuzoxane, nedaplatin, cytarabine, bicalutamide, vinorelbine, vesnarinone, aminoglutethimide, amsacrine, proglumide, elliptinium, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine, estramustine phosphate sodium, estramustine diphosphate sodium, estramustine sodium phosphate, estramustine diphosphate sodium, estramustine sodium phosphate,Acetate, Ketanserin, Doxifluridine, Etretinate, Isotretinoin, Streptozocin, Nimustine, Vindesine, Flutamide, Drogenil, Butocin, Carmofur, Razoxane, Sizofilan, Carboplatin, Mitolactalol, Tegafur, Ifosfamide, etc. Nimustine, picibanil, levamisole, teniposide, improsulfan, enocitabine, lisuride, oxymetholone, tamoxifen, progesterone, mepitiostane, epitiostanol, formestane, interferon-α, interferon-2α, interferon-β, interferon-γ, colony-stimulating factor-1, colony-stimulating factor-2, denileukin diftitox, interleukin-2, and luteinizing hormone-releasing factor.

[0151] In some embodiments, the second therapeutic agent is an mTOR inhibitor that inhibits cell proliferation, angiogenesis, and glucose uptake. Approved mTOR inhibitors for use in this invention include everolimus (…). Novartis; temsirolimus ( Pfizer); and sirolimus (Pfizer); Pfizer).

[0152] In some embodiments, the second therapeutic agent is a poly-ADP-ribose polymerase (PARP) inhibitor. PARP inhibitors approved for use in this invention include olaparib. AstraZeneca); Rucaparib (AstraZeneca); Clovis Oncology); and niraparib (Zejula®) Tesaro). Other PARP inhibitors under investigation that can be used in the present application include talazoparib (MDV3800 / BMN 673 / LT00673, Medivation / Pfizer / Biomarin); veliparib (ABT-888, AbbVie); and BGB-290 (BeiGene, Inc.).

[0153] In certain embodiments, the second therapeutic agent is a phosphatidylinositol 3 kinase (PI3K) inhibitor. Approved PI3K inhibitors that can be used in the present application include idelalisib (Zydelig®) Gilead). Other PI3K inhibitors under investigation that can be used in the present application include alpelisib (BYL719, Novartis); taselisib (GDC-0032, Genentech / Roche); pictilisib (GDC-0941, Genentech / Roche); copanlisib (BAY806946, Bayer); duvelisib (formerly IPI-145, Infinity Pharmaceuticals); PQR309 (Piqur Therapeutics, Switzerland); and TGR1202 (formerly RP5230, TG Therapeutics).

[0154] In certain embodiments, the second therapeutic agent is a proteasome inhibitor. Approved proteasome inhibitors that can be used in the present application include bortezomib (Velcade®) Takeda); carfilzomib (Kyprolis® Amgen); and ixazomib (Ninlaro® Takeda).

[0155] In certain embodiments, the second therapeutic agent is a histone deacetylase (HDAC) inhibitor. Approved HDAC inhibitors that can be used in the present application include vorinostat (Zolinza® Merck); romidepsin (Istodax® Celgene); panobinostat (Farydak® Novartis); and belinostat (Novartis); Spectrum Pharmaceuticals). Other HDAC inhibitors under investigation that may be used in this invention include entinostat (SNDX-275, Syndax Pharmaceuticals) (NCT00866333); and chidamide ( HBI-8000, Chipscreen Biosciences, China).

[0156] In some embodiments, the second therapeutic agent is a CDK inhibitor, such as a CDK 4 / 6 inhibitor. CDK 4 / 6 inhibitors approved for use in this invention include palbociclib (…). Pfizer); and Ribociclib (Pfizer); Novartis). Other CDK 4 / 6 inhibitors under investigation that may be used in this invention include abemaciclib (Ly2835219, Eli Lilly); and trilaciclib (G1T28, G1 Therapeutics).

[0157] In some embodiments, the second therapeutic agent is an indoleamine (2,3)-dioxygenase (IDO) inhibitor. Investigative IDO inhibitors that may be used in this invention include epacadostat (INCB024360, Incyte); indoximod (NLG-8189, NewLink Genetics Corporation); capmanitib (INC280, Novartis); GDC-0919 (Genentech / Roche); PF-06840003 (Pfizer); BMS:F001287 (Bristol-Myers Squibb); Phy906 / KD108 (Phytoceutica); and an enzyme that breaks down kynurenine (Kyn Therapeutics).

[0158] In some embodiments, the second therapeutic agent is a growth factor antagonist, such as an antagonist of platelet-derived growth factor (PDGF) or epidermal growth factor (EGF) or its receptor (EGFR). PDGF antagonists approved for use in this invention include olaratumab (…). Eli Lilly). EGFR antagonists approved for use in this invention include cetuximab (…). Eli Lilly); necitumumab (Eli Lilly); Eli Lilly); panitumumab (Pantumumab) Amgen); and osimertinib (targeting activated EGFR, AstraZeneca).

[0159] In some embodiments, the second therapeutic agent is an aromatase inhibitor. Approved aromatase inhibitors for use in this invention include exemestane (…). Pfizer); Anastazole AstraZeneca; and letrozole ( Novartis).

[0160] In some embodiments, the second therapeutic agent is a hedgehog protein pathway antagonist. Hedgehog protein pathway inhibitors approved for use in this invention include sodegib (…). SunPharmaceuticals); and vismodegib ( Genentech, both are used to treat basal cell carcinoma.

[0161] In some embodiments, the second therapeutic agent is a folic acid inhibitor. Folic acid inhibitors approved for use in this invention include pemetrexed. Eli Lilly).

[0162] In some embodiments, the second therapeutic agent is a CC chemokine receptor 4 (CCR4) inhibitor. Investigative CCR4 inhibitors that may be used in this invention include mogamulizumab (…). Kyowa Hakko Kirin, Japan).

[0163] In certain embodiments, the second therapeutic agent is an isocitrate dehydrogenase (IDH) inhibitor. Investigational IDH inhibitors that can be used in the present application include AG120 (Celgene; NCT02677922); AG221 (Celgene, NCT02677922; NCT02577406); BAY1436032 (Bayer, NCT02746081); IDH305 (Novartis, NCT02987010).

[0164] In certain embodiments, the second therapeutic agent is an arginase inhibitor. Investigational arginase inhibitors that can be used in the present application include AEB1102 (pegylated recombinant arginase, Aeglea Biotherapeutics), which is being investigated in Phase 1 clinical trials for acute myeloid leukemia and myelodysplastic syndromes (NCT02732184) and solid tumors (NCT02561234); and CB-1158 (Calithera Biosciences).

[0165] In certain embodiments, the second therapeutic agent is a glutaminase inhibitor. Investigational glutaminase inhibitors that can be used in the present application include CB-839 (Calithera Biosciences).

[0166] In certain embodiments, the second therapeutic agent is an antibody that binds to a tumor antigen, i.e., a protein expressed on the cell surface of a tumor cell. Approved antibodies that bind to a tumor antigen that can be used in the present application include rituximab (RITUXAN®) Genentech / Biogen Idec); ofatumumab (ARZERRA®) (anti-CD20, GlaxoSmithKline); obinutuzumab (GAZYVA®) (anti-CD20, Genentech); ibritumomab (ZEVALIN®) (anti-CD20 and yttrium-90, Spectrum Pharmaceuticals); daratumumab (DARZALEX®) (anti-CD38, Janssen Biotech); dinutuximab (UNITUXIN®) (anti-glycolipid GD2, United Therapeutics); trastuzumab (HERCEPTIN®) (anti-HER2, Genentech); ADO-trastuzumab (anti-HER2, fused with emtansine). Genentech); and pertuzumab (anti-HER2, Genentech); and brentuximab vedotin (an anti-CD30 drug conjugate). Seattle Genetics.

[0167] In some embodiments, the second therapeutic agent is a topoisomerase inhibitor. Approved topoisomerase inhibitors for use in this invention include irinotecan (Irinotecan). Merrimack Pharmaceuticals); Topotecan GlaxoSmithKline). Topoisomerase inhibitors under investigation that can be used in this invention include pixantrone (…). CTIBiopharma).

[0168] In some implementations, the second therapeutic agent is a nucleoside inhibitor or another therapeutic agent that interferes with normal DNA synthesis, protein synthesis, cell replication, or otherwise inhibits rapidly proliferating cells. Such nucleoside inhibitors or other therapeutic agents include trabectedin (a guanidine alkylating agent). Janssen Oncology); nitrogen mustard (mechlorethamine) (alkylating agent, Aktelion Pharmaceuticals); Vincristine ( Eli Lilly; Teva Pharmaceuticals; Talon Therapeutics); Temozolomide (alkylating agent 5-(3-methyltriazine-1-yl)-imidazol-4-carboxamide (MTIC)) The prodrug, Merck); cytarabine injection (ara-C, an antimetabolite cytidine analog, Pfizer); lomustine (an alkylating agent, ... Bristol-Myers Squibb; NextSource Biotechnology); azacitidine (a pyrimidine nucleoside analog of cytidine, Celgene); omacetaxine mepesuccinate (cephalotaxine ester) (inhibitor of protein synthesis, Teva Pharmaceuticals); asparaginase Erwinia chrysanthemi (asparagine-depleting enzyme, Lundbeck; EUSAPharma); eribulin mesylate (microtubule inhibitor, tubulin-based antimitotic agent, Eisai); cabazitaxel (microtubule inhibitor, tubulin-based antimitotic agent, Sanofi-Aventis); capacetrine (thymidylate synthase inhibitor, Genentech); bendamustine (bifunctional nitrogen mustard derivative, believed to form interstrand DNA crosslinks, Cephalon / Teva); ixabepilone (semisynthetic analog of epothilone B, microtubule inhibitor, tubulin-based antimitotic agent, Bristol-Myers Squibb); nelarabine (prodrug of deoxyguanosine analogs, nucleoside metabolism inhibitor, Novartis); clorafabine (prodrug of ribonucleotide reductase inhibitor, competitive inhibitor of deoxycytidine, Sanofi-Aventis); and trifluridine with tipiracil (thymidine-based nucleoside analog with thymidine phosphorylase inhibitor, Taiho Oncology).

[0169] In certain embodiments, the second therapeutic agent is a platinum-based therapeutic agent, also known as a platinum drug. Platinum drugs cause DNA crosslinks, thereby inhibiting DNA repair and / or DNA synthesis, primarily in rapidly proliferating cells, such as cancer cells. Approved platinum-based therapeutic agents that can be used in the present application include cisplatin (Platinol®) Bristol-Myers Squibb); carboplatin (Paraplatin® Bristol-Myers Squibb; also Teva; Pfizer); oxaliplatin ( Sanofi-Aventis); and nedaplatin (Nelvanax® Shionogi). Other platinum-based therapeutic agents that have undergone clinical testing and can be used in the present application include picoplatin (Poniard Pharmaceuticals); and satraplatin (JM-216, Agennix).

[0170] In certain embodiments, the second therapeutic agent is a taxane compound that causes disruption of microtubules essential for cell division. Taxane compounds approved for use in the present application include paclitaxel (Taxol® Bristol-Myers Squibb), docetaxel (Taxotere® Sanofi-Aventis; Sun Pharmaceutical), albumin-bound paclitaxel (Abraxane® Abraxis / Celgene), and cabazitaxel (Jevtana® Sanofi-Aventis). Other taxane compounds that have undergone clinical testing and can be used in the present application include SID530 (SK Chemicals, Co.) (NCT00931008).

[0171] In certain embodiments, the second therapeutic agent is an inhibitor of anti-apoptotic proteins such as BCL-2. Anti-apoptotic agents approved for use in the present application include venetoclax (Venclexta® AbbVie / Genentech); and blinatumomab (Blincyto® Amgen). Other therapeutic agents that target apoptotic proteins that have undergone clinical testing and can be used in the present application include navitoclax (ABT-263, Abbott), BCL-2 inhibitor (NCT02079740).

[0172] In certain embodiments, the second therapeutic agent is a selective estrogen receptor modulator (SERM) that interferes with the synthesis or activity of estrogen. SERMs approved for use in the present application include raloxifene (Evista® Eli Lilly).

[0173] In certain embodiments, the second therapeutic agent is an inhibitor of the interaction between the two major p53 inhibitory proteins, MDMX and MDM2. Inhibitors of the p53 inhibitory proteins under investigation that can be used in the present application include ALRN-6924 (Aileron), which is a stapled peptide that binds and disrupts the interaction of MDMX and MDM2 with p53 equivalently. ALRN-6924 is currently being evaluated in clinical trials for the treatment of AML, advanced myelodysplastic syndrome (MDS), and peripheral T-cell lymphoma (PTCL) (NCT02909972; NCT02264613).

[0174] In certain embodiments, the second therapeutic agent is an inhibitor of the transforming growth factor-beta (TGF-beta or TGFp). Inhibitors of the TGF-beta proteins under investigation that can be used in the present application include NIS793 (Novartis), which is an anti-TGF-beta antibody being tested in the clinic for the treatment of various cancers, including breast cancer, lung cancer, hepatocellular carcinoma, colorectal cancer, pancreatic cancer, prostate cancer, and renal cancer (NCT02947165). In some embodiments, the TGF-beta protein inhibitor is fresolimumab (GC1008; Sanofi-Genzyme), which is being investigated for melanoma (NCT00923169), renal cell carcinoma (NCT00356460), and non-small cell lung cancer (NCT02581787). Additionally, in some embodiments, the additional therapeutic agent is a TGF-beta trap, such as described in Connolly et al. (2012) Int'l J. Biological Sciences 8:964-978. One therapeutic compound currently in clinical trials for the treatment of solid tumors is M7824 (Merck KgaA - formerly MSB0011459X), which is a bispecific anti-PD-L1 / TGFp trap compound (NCT02699515; and (NCT02517398). M7824 consists of a fully human IgGl antibody against PD-L1 fused to the extracellular domain of human TGF-beta receptor II, which can act as a TGFp "trap."

[0175] In certain embodiments, the second therapeutic agent is a cancer vaccine. In some embodiments, the cancer vaccine is selected from sipuleucel-T (Provenge® Dendreon / Valeant Pharmaceuticals), which has been approved for the treatment of asymptomatic or mildly symptomatic metastatic hormone -refractory prostate cancer; and talimogene laherparepvec (Imlygic® BioVex / Amgen (formerly known as T-VEC) is a genetically modified oncolytic virus therapy approved for the treatment of unresectable cutaneous, subcutaneous, and nodular lesions of melanoma. In some embodiments, additional therapeutic agents are selected from oncolytic virus therapies, such as pexastimogene devacirepvec (PexaVec / JX-594, SillaJen / formerly Jennerex Biotherapeutics), an engineered thymidine kinase (TK)-deficient vaccinia virus expressing GM-CSF targeting hepatocellular carcinoma (NCT02562755) and melanoma (NCT00429312); pelareorep (… Oncolytics Biotech describes a respiratory enterovirus (ROV) variant that fails to replicate in non-RAS-activated cells in various cancers, including colorectal cancer (NCT01622543), prostate cancer (NCT01619813), head and neck squamous cell carcinoma (NCT01166542), pancreatic adenocarcinoma (NCT00998322), and non-small cell lung cancer (NSCLC) (NCT01622543). 00861627); enadenotucirev (NG-348, PsiOxus, formerly known as ColoAd1), an engineered adenovirus to express full-length CD80 and T-cell receptor CD3 protein-specific antibody fragments in ovarian cancer (NCT02028117), metastatic or advanced epithelial tumors (such as colorectal cancer, bladder cancer, head and neck squamous cell carcinoma, and salivary gland cancer) (NCT02636036); ONCOS-102 (Targovax / formerly Oncos), an engineered adenovirus to express GM-CSF in melanoma (NCT03003676) and peritoneal diseases, colorectal cancer, or ovarian cancer (NCT02963831); GL-ONC1 (GLV-1h68 / GLV-1h153, Genelux) GmbH, which is a vaccinia virus engineered to express β-galactosidase (β-gal) / β-glucuronidase or β-gal / human sodium iodide cotransporter (hNIS) in peritoneal metastatic cancer (NCT01443260), fallopian tube cancer, and ovarian cancer (NCT 02759588); or CG0070 (Cold Genesys), which is an adenovirus engineered to express GM-CSF in bladder cancer (NCT02365818).

[0176] In certain embodiments, the second therapeutic agent is an immune checkpoint inhibitor selected from a PD-1 antagonist, a PD-L1 antagonist, or a CTLA-4 antagonist. In some embodiments, a compound disclosed herein, or a pharmaceutically acceptable salt thereof, is administered in combination with nivolumab (anti-PD-1 antibody, Bristol-Myers Squibb); pembrolizumab (anti-PD-1 antibody, Merck); ipilimumab (anti-CTLA-4 antibody, Bristol-Myers Squibb); durvalumab (anti-PD-L1 antibody, AstraZeneca); or atezolizumab (anti-PD-L1 antibody, Genentech). Other immune checkpoint inhibitors suitable for use in the present application include REGN2810 (Regeneron), which is an anti-PD-1 antibody being tested in patients with basal cell carcinoma (NCT03132636), NSCLC (NCT03088540), squamous cell carcinoma of the skin (NCT02760498), lymphoma (NCT02651662), and melanoma (NCT03002376); pidilizumab (CureTech), also known as CT-011, which is an antibody that binds to PD-1, is in clinical trials for diffuse large B-cell lymphoma and multiple myeloma; avelumab (PF-06801591) Pfizer / Merck KGaA, also known as MSB0010718C), which is a fully human IgGl anti-PD-Ll antibody, is in clinical trials for non-small cell lung cancer, Merkel cell carcinoma, mesothelioma, solid tumors, renal cancer, ovarian cancer, bladder cancer, head and neck cancer, and gastric cancer; and PDR001 (Novartis), which is an inhibitory antibody that binds to PD-1, is in clinical trials for non-small cell lung cancer, melanoma, triple-negative breast cancer, and advanced or metastatic solid tumors. Tremelimumab (CP-675,206; Astrazeneca) is a fully human monoclonal antibody against CTLA-4 that has been studied in clinical trials for a variety of indications, including: mesothelioma, colorectal cancer, renal cancer, breast cancer, lung cancer and non-small cell lung cancer, pancreatic ductal adenocarcinoma, pancreatic cancer, germ cell cancer, head and neck squamous cell carcinoma, hepatocellular carcinoma, prostate cancer, endometrial cancer, liver metastases, hepatocarcinoma, large B-cell lymphoma, ovarian cancer, cervical cancer, metastatic anaplastic thyroid cancer, urothelial cancer, fallopian tube cancer, multiple myeloma, bladder cancer, soft tissue sarcoma, and melanoma. AGEN-1884 (Agenus) is an anti-CTLA4 antibody being studied in a Phase 1 clinical trial for advanced solid tumors (NCT02694822).

[0177] Another aspect of the application provides the use of a compound described herein, such as a crystalline compound of Formula II-5 or a compound in Table 3, or a pharmaceutically acceptable salt thereof, or other compounds in Section I, to manufacture a medicament. In certain embodiments, the medicament is used to treat a disease described herein, such as a cancer.

[0178] Another aspect of the application provides the use of a compound described herein, such as a crystalline compound of Formula II-5 or a compound in Table 3, or a pharmaceutically acceptable salt thereof, or other compounds in Section I, to manufacture a medicament. In certain embodiments, the medicament is used to treat a disease described herein, such as a cancer.

[0179] Another aspect of the application provides the use of a compound described herein, such as a crystalline compound of Formula II-5 or a compound in Table 3, or a pharmaceutically acceptable salt thereof, or other compounds in Section I, to manufacture a medicament. In certain embodiments, the medicament is used to treat a disease described herein, such as a cancer.

[0180] Evaluation of cell growth inhibition of HEK293 cells and HeLa cells

[0181] The ability of compounds to inhibit the proliferation of HEK293 cells or HeLa cells can be evaluated according to the following procedure. HEK293 and HeLa cells are cultured in DMEM medium supplemented with 10% fetal bovine serum and 1% Penn / Strep. Cells are seeded at 500 cells / well in white 384-well plates in 25 pL of complete medium. After seeding, the plates are spun at 300 x g for three minutes and incubated at 37 °C and 5% CO2 in a humidified tissue culture incubator. Twenty-four hours later, compounds are titrated in 100% DMSO and diluted in complete cell culture medium. Twenty-five pL aliquots of the compound / medium mixture are added to the cells, bringing the total volume in the wells to 50 pL. DMSO alone is used as a negative control. The plates are then spun at 300 x g for three minutes and stored at 37 °C and 5% CO2 for three days. Cell viability is quantified on day 0 and day 3 of compound treatment using CellTiter-Glo 2.0 reagent (Promega). After equilibrating the microplate at room temperature for 30 minutes, 25 pL of CellTiter-Glo 2.0 reagent is dispensed into each well, bringing the total volume to 75 pL. The plate is mixed on a shaker at 500 rpm for 2 minutes and then incubated at room temperature for 10 minutes. After a quick spin, luminescence readings are measured using an EnVision plate reader. Data are normalized to day 0 and day 3 readings of DMSO treatment. A four-parameter nonlinear regression curve fit is applied to the dose response data in GraphPad Prism data analysis software to determine the half-maximal growth inhibition concentration (GI50) for each compound. 50 ).

[0182] III. Pharmaceutical compositions and dosing considerations

[0183] As indicated above, the present invention provides pharmaceutical compositions comprising a therapeutically effective amount of one or more of the compounds described above, formulated together with one or more pharmaceutically acceptable carriers (additives) and / or diluents. The pharmaceutical compositions may be specifically formulated for administration in solid or liquid form, including those suitable for: (1) oral administration, e.g., enemas (aqueous or non-aqueous solutions or suspensions), tablets (e.g., those intended for buccal, sublingual, and systemic absorption), pills, powders, granules, pastes for tongue administration; (2) parenteral administration, e.g., by subcutaneous, intramuscular, intravenous, or epidural injection, as, for example, a sterile solution or suspension or a sustained-release formulation; (3) topical administration, e.g., as creams, ointments, or controlled-release patches or sprays for skin application; (4) intravaginal or rectal administration, e.g., as pessaries, creams, or foams; (5) sublingual; (6) ocular; (7) transdermal; or (8) nasal. In some embodiments, the present invention provides a pharmaceutical composition comprising a compound described herein (e.g., a crystalline compound of formula II-5 or a compound in Table 3 or a pharmaceutically acceptable salt thereof) and a pharmaceutically acceptable carrier.

[0184] As used herein, the phrase "therapeutic effective amount" means the amount of a compound, material, or composition comprising the compounds of the present invention that effectively produces a desired therapeutic effect in at least a subpopulation of animals, given a reasonable benefit / risk ratio applicable to any medical treatment.

[0185] The phrase “pharmaceutically acceptable” is used in this document to refer to compounds, materials, compositions, and / or dosage forms that, to the extent of reasonable medical judgment, are suitable for use in human and animal tissues without excessive toxicity, irritation, allergic reactions, or other problems or complications, and that match a reasonable benefit / risk ratio.

[0186] Wetting agents, emulsifiers and lubricants (such as sodium lauryl sulfate and magnesium stearate), as well as colorants, releasing agents, coating agents, sweeteners, flavoring agents and aroma agents, preservatives and antioxidants may also be present in the composition.

[0187] Examples of pharmaceutically acceptable antioxidants include: (1) water-soluble antioxidants, such as ascorbic acid, cysteine ​​hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite, etc.; (2) oil-soluble antioxidants, such as ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, α-tocopherol, etc.; and (3) metal chelating agents, such as citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitol, tartaric acid, phosphoric acid, etc.

[0188] Formulations of the present application include those suitable for oral, nasal, topical (including buccal and sublingual), rectal, vaginal and / or parenteral administration. The formulations can conveniently be presented in unit dosage form and can be prepared by any methods well known in the art of pharmacy. The amount of active ingredient which can be combined with a carrier material to produce a single dosage form will vary depending upon the host being treated, the particular mode of administration. The amount of active ingredient which can be combined with a carrier material to produce a single dosage form will generally be that amount of a compound which produces a therapeutic effect. Generally, this amount will range from about 0.1% to about 99% of the active ingredient, preferably from about 5% to about 70%, most preferably from about 10% to about 30% by weight of the total composition. Solid dosage forms for oral administration can include capsules, tablets, pills, powders, granules or suppositories.

[0189] In certain embodiments, formulations of the present application comprise an excipient selected from the group consisting of cyclodextrins, celluloses, liposomes, micelle-forming agents (e.g., bile acids) and polymeric carriers (e.g., polyesters and polyanhydrides); and a compound of the present application. In certain embodiments, the foregoing formulations facilitate oral bioavailability of a compound of the present application.

[0190] Methods of preparing these formulations or compositions include the step of bringing into association a compound of the present application with the carrier and, optionally, one or more accessory ingredients. In general, the formulations are prepared by uniformly and intimately associating a compound of the present application with a liquid carrier or a finely divided solid carrier or both, and then, if necessary, shaping the product.

[0191] Formulations of the present application suitable for oral administration can be in the form of capsules, cachets, tablets, pills, lozenges (using a flavored basis, usually sucrose and acacia or tragacanth), powders, granules, or as a solution or a suspension in an aqueous or non-aqueous liquid, or as an oil-in-water or water-in-oil emulsion, or as an elixir or syrup, or as pastilles (using an inert base, such as gelatin and glycerin, or sucrose and acacia), and / or as mouth washes and the like, each containing a predetermined amount of a compound of the present application as an active ingredient. A compound of the present application can also be administered as a bolus, electuary or paste.

[0192] In solid dosage forms of the application for oral administration (for example, capsules, tablets, pills, dragees, powders, granules, throat lozenges, etc.), the active ingredient is mixed with one or more pharmaceutically-acceptable carriers, such as sodium citrate or dicalcium phosphate, and / or any of the following: (1) fillers or extenders, such as starches, lactose, sucrose, glucose, mannitol, and / or silicic acids; (2) binders, such as carboxymethylcellulose, alginates, gelatin, polyvinyl pyrrolidone, sucrose, and / or acacia; (3) humectants, such as glycerol; (4) disintegrating agents, such as agar-agar, calcium carbonate, potato or tapioca starch, alginic acid, certain silicates, and sodium carbonate; (5) solution retarders, such as paraffin; (6) absorption accelerators, such as quaternary ammonium compounds, and surfactants, such as poloxamer and sodium lauryl sulfate; (7) wetting agents, such as cetyl alcohol, glycerol monostearate, and non-ionic surfactants; (8) absorbents, such as kaolin and bentonite clay; (9) lubricants, such as talc, calcium stearate, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, zinc stearate, sodium stearate, stearic acid, and mixtures thereof; (10) coloring agents; and (11) release-affecting agents, such as crospovidone or ethyl cellulose. The pharmaceutical compositions also can contain a buffering agent. Solid compositions of a similar type can also be employed as fillers in soft and hard-filled gelatin capsules using such excipients as lactose or milk sugar, as well as high molecular weight polyethylene glycols.

[0193] Tablets can be prepared by compression or molding, optionally with one or more accessory ingredients. Binding agents (e.g., gelatin or hydroxypropyl methylcellulose), lubricants, inert diluents, preservatives, disintegrating agents (e.g., sodium starch glycolate or croscarmellose sodium), surface-active or dispersing agents can be used. Molded tablets can be made by molding in a suitable machine a mixture of the powdered compound moistened with a liquid diluent.

[0194] The tablets and other solid dosage forms of the pharmaceutical compositions of the present application, such as dragees, capsules, pills and granules, can optionally be scored or prepared with coatings and shells, such as enteric coatings and other coatings well known in the pharmaceutical formulating art. They can also be formulated so as to provide a slow or controlled release of the active ingredient therein, including time release or sustained release formulations, as for example, by different ratios of hydroxypropylmethyl cellulose and other polymer matrices, liposomes and / or microspheres. They can be formulated for rapid release, e.g., by freeze- drying. They can be sterilized by, for example, filtration through a bacteria-retaining filter, or by incorporating sterilizing agents in the form of sterile solid compositions which can be dissolved in sterile water, or some other sterile injectable medium, just prior to use. To these compositions also can be added adjuvants customarily used in the art, and depending on the dosage form desired. Embodiments of embedding compositions which can be used include polymeric substances and waxes. The active ingredient can also be in micro-encapsulated form, if appropriate, with one or more of the above-described excipients.

[0195] Liquid dosage forms for oral administration of the compounds of the application include pharmaceutically acceptable emulsions, microemulsions, solutions, suspensions, syrups and elixirs. In addition to the active ingredient, the liquid dosage forms can contain inert diluents commonly used in the art, such as water or other solvents, solubilizing agents and emulsifiers, such as ethyl alcohol, isopropyl alcohol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1,3- butylene glycol, oils (in particular, cottonseed, groundnut, corn, germ, olive, castor and sesame oils), glycerol, tetrahydrofuryl alcohol, polyethylene glycols and fatty acid esters of sorbitan, and mixtures thereof.

[0196] In addition to inert diluents, the oral compositions can include adjuvants such as wetting agents, emulsifying and suspending agents, sweetening, flavoring, coloring, perfuming and preservative agents.

[0197] Suspensions, in addition to the active ingredient, can contain suspending agents, as for example, ethoxylated isostearyl alcohols, polyoxyethylene sorbitol and sorbitan esters, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar-agar and tragacanth, and mixtures thereof.

[0198] Formulations of the pharmaceutical compositions of the application for rectal or vaginal administration can be presented as a suppository, which can be prepared from a mixture of one or more compounds of the application with one or more suitable nonirritating excipients or carriers comprising, for example, cocoa butter, polyethylene glycol, a suppository wax or a salicylate, and which is solid at room temperature, but liquid at body temperature and, therefore, will melt in the rectum or vaginal cavity to release the active component.

[0199] Formulations of the present application suitable for vaginal administration also include pessaries, tampons, creams, gels, pastes, foams or spray formulations containing such carriers as are known in the art to be appropriate.

[0200] Dosage forms for topical or transdermal administration of a compound of this application include powders, sprays, ointments, pastes, creams, lotions, gels, solutions, patches and inhalants. The active compound can be mixed under sterile conditions with a pharmaceutically acceptable carrier and any needed preservatives, buffers or propellants which can be required.

[0201] Ointments, pastes, creams and gels can contain, in addition to an active compound of this application, excipients such as animal and vegetable fats, oils, waxes, paraffins, starch, tragacanth, cellulose derivatives, polyethylene glycols, silicones, bentonites, silicic acid, talc and zinc oxide, or mixtures thereof.

[0202] Powders and sprays can contain, in addition to a compound of this application, excipients such as lactose, talc, silicic acid, aluminum hydroxide, calcium silicates and polyamide powder or mixtures of these substances. Sprays can additionally contain customary propellants such as chlorofluorohydrocarbons and volatile unsubstituted hydrocarbons, such as butane and propane.

[0203] Transdermal patches have the added advantage of providing controlled delivery of a compound of the present application to the body. Such dosage forms can be made by dissolving or dispensing the compound in the proper medium. Absorption enhancers can also be used to increase the flux of the compound across the skin. The rate of such flux can be controlled by either providing a rate controlling membrane or dispersing the compound in a polymer matrix or gel.

[0204] Ophthalmic formulations, eye ointments, powders, solutions, and the like, are also contemplated as being within the scope of this application.

[0205] Pharmaceutical compositions of this application suitable for parenteral administration include one or more compounds of the application in combination with one or more pharmaceutically acceptable sterile isotonic aqueous or nonaqueous solutions, dispersions, suspensions or emulsions, or sterile powders which can be reconstituted into sterile injectable solutions or dispersions just prior to use, which can contain sugars, alcohols, antioxidants, buffers, bacteriostats, solutes which render the formulation isotonic with the blood of the intended recipient or suspending agents.

[0206] Examples of suitable aqueous and nonaqueous carriers which can be employed in the pharmaceutical compositions of the application include water, ethanol, polyols (such as glycerol, propylene glycol, polyethylene glycol, and the like), and suitable mixtures thereof, vegetable oils, such as olive oil, and injectable organic esters, such as ethyl oleate. Proper fluidity can be maintained, for example, by the use of coating materials, such as lecithin, by the maintenance of the required particle size in the case of dispersion and by the use of surfactants.

[0207] These compositions can also contain adjuvants such as preservatives, wetting agents, emulsifying agents, and dispersing agents. Prevention of the action of microorganisms can be ensured by the inclusion of various antibacterial and antifungal agents, for example, parabens, chlorobutanol, phenol, sorbic acid, and the like. It can also be desirable to include isotonic agents, for example, sugars, sodium chloride, and the like, into the compositions. In addition, prolonged absorption of the injectable pharmaceutical form can be brought about by the inclusion of agents which delay absorption, for example, aluminum monostearate and gelatin.

[0208] In some cases, in order to prolong the effect of a drug, it is desirable to slow its absorption from subcutaneous or intramuscular injection. This can be accomplished by the use of a liquid suspension of crystalline or amorphous material having poor water solubility. The rate of absorption of the drug then depends upon its rate of dissolution that, in turn, can depend upon crystal size and crystalline form. Alternatively, delayed absorption of a parenterally administered drug form is accomplished by dissolving or suspending the drug in an oil vehicle.

[0209] Injectable depot forms are made by forming microencapsule matrices of the subject compound in biodegradable polymers such as polylactide-polyglycolide. Depending on the ratio of drug to polymer, and the particular polymer / drug composition utilized, the rate of drug release can be controlled. Examples of other biodegradable polymers include poly(orthoesters) and poly(anhydrides). Depot injectable formulations are also prepared by entrapping the drug in liposomes or microemulsions that are compatible with body tissues.

[0210] When the compounds of the present application are administered as pharmaceuticals, to humans and animals, they can be given per se or as a pharmaceutical composition containing, for example, 0.1 to 99% (more preferably, 10 to 30%) of active ingredient in combination with a pharmaceutically acceptable carrier.

[0211] The formulations of the present application can be orally, parenterally, topically, or rectally administered. It will be noted that the formulations are administered in forms conventionally used for such purposes: for example, in tablets or capsules for oral administration; in suppositories for rectal administration; in injectable, inhalable, eye drops, ointments, or creams for parenteral administration; in injectable, infusable, or inhalable for parenteral administration; in lotions or ointments for topical administration; and in suppositories for rectal administration. Oral administration is preferred.

[0212] The phrases "parenteral administration" and "administered parenterally" as used herein, mean modes of administration other than oral and topical administration, usually by injection, and include, but are not limited to, intravenous, intramuscular, intraarterial, intrathecal, intracapsular, intraorbital, intracardiac, intradermal, intraperitoneal, transtracheal, subcutaneous, subcuticular, intraarticular, subcapsular, subarachnoid, intraspinally, and intrasternal injection and infusion.

[0213] As used herein, the phrases “systemic administration,” “administered systemically,” “peripheral administration,” and “administered peripherally” refer to the administration of a compound, drug, or other material in a manner other than direct administration to the central nervous system, so that it enters the patient’s system and undergoes metabolism and other similar processes, such as subcutaneous administration.

[0214] These compounds can be administered to humans and other animals for therapeutic purposes via any suitable route of administration, including oral, nasal (e.g., via spray), rectal, vaginal, parenteral, intracerebral, and topical (e.g., via powder, ointment, or drops), including buccal and sublingual administration.

[0215] Regardless of the chosen route of administration, the compounds and / or pharmaceutical compositions of the present invention, which can be used in a suitable hydrated form, are formulated into pharmaceutically acceptable dosage forms using conventional methods known to those skilled in the art.

[0216] The actual dose level of the active ingredient in the pharmaceutical composition of the present invention may vary to obtain an amount of active ingredient that is effective in achieving the desired therapeutic response for a particular patient, composition and administration mode, while being non-toxic to the patient.

[0217] The selected dosage level will depend on a variety of factors, including the activity of the specific compound of the present invention or its ester, salt or amide, the route of administration, the time of administration, the rate of excretion or metabolism of the specific compound, the rate and extent of absorption, the duration of treatment, other drugs, compounds and / or materials used in combination with the specific compound, the age, sex, weight, disease, general health and medical history of the patient being treated, and similar factors well known in the medical field.

[0218] A physician or veterinarian of ordinary skill in the art can readily determine and prescribe an effective amount of the desired pharmaceutical composition. For example, a physician or veterinarian may begin with a dose of the compound of the invention used in the pharmaceutical composition at a level below that required to achieve the desired therapeutic effect and gradually increase the dose until the desired effect is achieved.

[0219] In general, a suitable daily dose of a compound of the application will be that amount of the compound that is the lowest dose effective to produce a therapeutic effect. Generally, such an effective dose will depend on the factors described above. Compounds are preferably administered in the range of about 0.01 mg / kg to about 200 mg / kg, more preferably about 0.1 mg / kg to about 100 mg / kg, even more preferably about 0.5 mg / kg to about 50 mg / kg. When a compound described herein is co-administered with another agent (e.g., as a sensitizing agent), the effective amount can be lower than the amount used alone.

[0220] If desired, the effective daily dose of the active compound can be administered as two, three, four, five, six or more sub-doses administered separately at appropriate intervals throughout the day, optionally, in unit dosage forms. Preferred dosing is once per day.

[0221] The present application further provides a unit dosage form (such as a tablet or capsule) comprising a therapeutically effective amount of a compound described herein for use in treating a medical condition described herein.

[0222] IV. Medical Kits

[0223] Another aspect of the present application is a kit comprising (i) a compound described herein, such as a crystalline compound of Formula II-5 or a compound in Table 3 or other compounds in Section I, or a pharmaceutically acceptable salt thereof, and (ii) instructions for use, such as for treating cancer.

[0224] Examples

[0225] The application now being generally described, will be more readily understood by reference to the following examples, which are included merely for purposes of illustration of certain aspects and embodiments of the present application, and are not intended to limit the application.

[0226] General methods

[0227] All reactions were carried out under a dry nitrogen or argon atmosphere. Glassware was oven dried prior to use. Common reagents or materials were obtained from commercial sources and used without further purification unless otherwise noted. Anhydrous N,N-diisopropylethylamine (DIPEA) was obtained by distillation from potassium hydroxide. Solvents were dried by passing through columns of activated 4A molecular sieves. The following abbreviations are used: aq = aqueous; Bn = benzyl; Boc = tert-butyloxycarbonyl; DCM = dichloromethane; DIPEA = N,N-diisopropylethylamine; DMF = dimethylformamide; DMSO = dimethylsulfoxide; EDC = 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide; EtOAc = ethyl acetate; EtOH = ethanol; HOBt = 1-hydroxybenzotriazole; HPLC = high performance liquid chromatography; LCMS = liquid chromatography-mass spectrometry; MeOH = methanol; NMR = nuclear magnetic resonance; RT = room temperature; sat. = saturated; TFA = trifluoroacetic acid; THF = tetrahydrofuran; TLC = thin layer chromatography. TMSolvents were dried over solvent drying systems. PTLC refers to preparative thin layer chromatography separation. Abbreviations: HFIP (hexafluoroisopropanol), HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid). Flash column chromatography was performed using silica gel 60 (230-400 mesh). Analytical thin layer chromatography (TLC) was performed on Merck silica gel plates with QF-254 indicator and visualized by UV or KMn04.

[0228] at room temperature on an Agilent DD2 500 (500 MHz 1 H; 125 MHz 13 C) or Agilent DD2 600 (600 MHz 1 H; 150 MHz 13 C) or Agilent DD2 400 (400 MHz 1 H; 100 MHz 13 C) spectrometer 1 H and 13 C NMR spectra. Chemical shifts are reported in ppm relative to residual CDCl3(δ 7.26 ppm 1 H; δ 77.0 ppm 13 C), CD3OD (δ 3.31 ppm 1 H; δ 49.00 ppm 13 C) or d6-DMSO (δ 2.50 ppm 1 H; δ 39.52 ppm 13 C). NMR chemical shifts are reported in ppm relative to the internal solvent peak and coupling constants are measured in Hz (bs = broad signal). In most cases, only the peaks of the major rotamers are reported.

[0229] Mass spectra were obtained using an Agilent 1100 series LC / MSD spectrometer. Analytical HPLC analysis was performed on a 250 x 4.6 mm C-18 column using gradient conditions (10-100% B, flow rate = 1.0 mL / min, 20 min) or as described in the LC-MS method table.

[0230] Preparative HPLC was performed on a 250 x 21.2 mm C-18 column using gradient conditions (10-100% B, flow rate = 10.0 mL / min, 20 min) unless otherwise indicated. The eluents used were: solvent A (H20 with 0.1% TFA) and solvent B (CH3CN with 0.1% TFA). Final products were typically purified via reverse phase HPLC, PTLC or flash column chromatography.

[0231]

[0232]

[0233]

[0234]

[0235]

[0236] Example 1 -Synthesis of N-[(1r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [13-C-2(6),4,7,10,12-pentene-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5-carboxamide(II-5)

[0237]

[0238]

[0239] Step 1: Preparation of 2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [Decadecyl-2(6),4,7,10,12-pentaen-9-yl]acetic acid. To 2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [1.0 g, 2.1 mmol, 1.0 equivalent] tert-butyl acetate (1.0 g, 2.1 mmol, 1.0 equivalent) was added to a solution of TFA (7.7 g, 67 mmol, 5.0 mL, 30 equivalent) in DCM (10 mL). The mixture was stirred at 25 °C for 0.5 h. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was 2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [Decadecano-2(6),4,7,10,12-pentene-9-yl]acetic acid (877 mg, 99% yield) was used directly in the next step without purification.

[0240] Step 2: Preparation of 2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamide. To a solution of 2-[(9S)-7-(4- chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]acetic acid (877 mg, 2.19 mmol, 1.0 equiv) and NH4Cl (351 mg, 6.56 mmol, 3.0 equiv) in DMF (8 mL) were added HATU (915 mg, 2.41 mmol, 1.1 equiv) and DIEA (848 mg, 6.56 mmol, 3.0 equiv). The mixture was stirred at 25 °C for 1 h. The mixture was filtered and concentrated. The residue was purified by prep-HPLC (column: Waters Xbridge CI 8 150 mm x 50 mm x 10 µm; mobile phase: [water(NH4HCO3)-ACN]; B%: 24%-54%, 10 min) to give 2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia-1,8,11,12- tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]acetamide (740 mg, 84% yield).

[0241] Step 3: Preparation of 2-[[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole. 1,3-Dioxol-2-one (103 mg, 1.20 mmol, 1.2 equiv) and 2-[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6A solution of 2-[[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (300 mg, 707 μmol, 1.0 equiv) and tert-butyl 2-hydroxy-7-azaspiro[3.5]nonane-7-carboxylate (853 mg, 3.54 mmol, 5.0 equiv) in dioxane (5 mL) was added tBuXPhos Pd G3 (112 mg, 141 μmol, 0.2 equiv) and Cs2CO3 (1.1 g, 3.5 mmol, 5.0 equiv). The mixture was degassed and purged with N2 3 times, then the mixture was stirred at 90 °C under N2 atmosphere for 2 h. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (SiO2, eluent 0-3% methanol / dichloromethane gradient, 18 mL / min) to give 2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0

[0242] Step 4: Preparation of tert-butyl 2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2- ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]nonane-7-carboxylate. To a solution of 2-[[(9S)-7-(4-chlorophenyl)-4,5,13-trimethyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (300 mg, 707 μmol, 1.0 equiv) and tert-butyl 2-hydroxy-7-azaspiro[3.5]nonane-7-carboxylate (853 mg, 3.54 mmol, 5.0 equiv) in dioxane (5 mL) was added tBuXPhos Pd G3 (112 mg, 141 μmol, 0.2 equiv) and Cs2CO3 (1.1 g, 3.5 mmol, 5.0 equiv). The mixture was degassed and purged with N2 3 times, then the mixture was stirred at 90 °C under N2 atmosphere for 2 h. The reaction mixture was filtered and concentrated under reduced pressure to give a residue. The residue was purified by flash silica gel chromatography (SiO2, eluent 0-3% methanol / dichloromethane gradient, 18 mL / min) to give 2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2g Silica flash column, eluent 0-3% methanol / dichloromethane gradient, 18 mL / min) to give 2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6tert-butyl 2-[[(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-4,5,13-trimethyl-3- thia- 1,8, 11, 12-tetraazatricyclo[8.3.0.0

[0243] Step 5: Preparation of 2-[[(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-4,5,13- trimethyl-3-thia- 1,8, 11, 12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole. To a mixture of tert-butyl 2-[[(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-4,5,13-trimethyl-3-thia- 1,8, 11, 12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (214 mg, 332 pmol, 99% yield, TFA) was used directly in the next step without purification. 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (214 mg, 332 pmol, 99% yield, TFA) was used directly in the next step without purification.

[0244] Step 6: Preparation of N-[(lr,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8, 11, 12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (214 mg, 332 pmol, 99% yield, TFA) was used directly in the next step without purification. 2,6To a solution of [2-(6),4,7,10,12-pentacosa-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (80 mg, 151 pmol, 1.0 equiv) and 2-chloro-N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]pyrimidine-5-carboxamide (94 mg, 227 pmol, 1.5 equiv) in NMP (0.5 mL) was added DIEA (60 mg, 460 pmol, 0.1 mL, 3.0 equiv). The mixture was stirred at 25 °C for 8 h. The reaction mixture was filtered and concentrated to give a residue. The residue was purified by prep-HPLC (column: Waters Xbridge C18 150 mm x 50 mm x 10 pm; mobile phase: [water (NH4HCO3) - ACN]; gradient: 54% - 84% B, 10 min) to give N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]tricyclo[8.3.0.0 1 H NMR (400 MHz, CD3OD) d 8.74 (s, 2H), 7.91 (d, J = 0.8 Hz, 1H), 7.54 (d, J = 8.6 Hz, 1H), 7.34 (d, J = 8.8 Hz, 2H), 7.14 (d, J = 0.8 Hz, 1H), 6.85 (d, J = 8.9 Hz, 2H), 6.64 (d, J = 2.1 Hz, 1H), 6.57 (dd, J = 2.1, 8.6 Hz, 1H), 4.79 - 4.71 (m, 2H), 4.27 - 4.25 (m, 1H), 4.16 - 4.12 (m, 1H), 3.95 (s, 7H), 3.90 - 3.83 (m, 2H), 2.72 (s, 3H), 2.61 - 2.50 (m, 2H), 2.48 (s, 3H), 2.03 - 1.92 (m, 2H), 1.72 (s, 7H), 1.30 (s, 6H), 1.24 (s, 6H). LC-MS: MS (ES + ): RT = 2.698 min, m / z = 907.4 [M+H] + .

[0245] Example 2-Synthesis of N-[(1r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [Decadecyl-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5-carboxamide crystal form A (II-5 of crystal form A)

[0246]

[0247] Part 1 - Preparation of 2-[[(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [Deca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole

[0248]

[0249] To 2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclic[8.3.0.0] 2,6 [Tetrate-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]nonane-7-carboxylic acid tert-butyl ester (22.5 g, 35.78 mmol, 1.0 equivalent) was added to a solution of DCM (50 mL) with TFA (31 mL). The mixture was stirred at 25 °C for 1 hour. The reaction mixture was quenched at 0 °C by adding NaHCO3 (300 mL). The aqueous layer was washed with (DCM / MeOH = 10:1, 300 mL × 3), and the combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure to give 2-[[(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] as a yellow solid. 2,6 [Decadecano-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (18.5 g, 35.0 mmol, 98% yield). LC-MS: MS(ES+): RT = 0.686 min, m / z = 529.3 [M+H] + ].

[0250] Part 2 - Preparation of tert-butyl N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)- 2,2,4,4-tetramethyl-cyclobutyl]carbamate

[0251]

[0252] To a solution of tert-butyl N-((1 r,3r)-3-hydroxy-2,2,4,4-tetramethyl- cyclobutyl)carbamate (65.0 g, 267 mmol, 1.0 equiv) in DMF (1 L) was added NaH (10.7 g, 267 mmol, 60% purity, 1.0 equiv) at 0 °C. The reaction mixture was stirred at 0 °C for 0.5 h. Then 4-fluoro-2-methoxy-benzonitrile (40.4 g, 267 mmol, 1.0 equiv) was added. The reaction mixture was stirred at 25 °C for 12 h. The reaction mixture was quenched with saturated aqueous NH4Cl (20 mL) at 0 °C and diluted with water (5 L). The mixture was extracted with ethyl acetate (1 L x 3). The combined organic layers were washed with brine (1 L), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. The resulting residue was recrystallized from EtOH (1 L), and the resulting solid was filtered. The filter cake was dried to give tert-butyl N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]carbamate (56.5 g, 151 mmol, 56% yield) as a white solid. LC-MS: MS (ES+): RT = 0.966 min, m / z = 375.3 [M+H + ] at 0 °C. The reaction mixture was stirred at 0 °C for 0.5 h. Then 4-fluoro-2-methoxy-benzonitrile (40.4 g, 267 mmol, 1.0 equiv) was added. The reaction mixture was stirred at 25 °C for 12 h. The reaction mixture was quenched with saturated aqueous NH4Cl (20 mL) at 0 °C and diluted with water (5 L). The mixture was extracted with ethyl acetate (1 L x 3). The combined organic layers were washed with brine (1 L), dried over anhydrous Na2SO4, filtered, and concentrated in vacuo. The resulting residue was recrystallized from EtOH (1 L), and the resulting solid was filtered. The filter cake was dried to give tert-butyl N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]carbamate (56.5 g, 151 mmol, 56% yield) as a white solid. LC-MS: MS (ES+): RT = 0.966 min, m / z = 375.3 [M+H

[0253] Part 3 - Preparation of 4-((1 r,3r)-3-amino-2,2,4,4-tetramethyl-cyclobutoxy)-2- methoxy-benzonitrile

[0254]

[0255] To a solution of tert-butyl N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]carbamate (50 g, 133.52 mmol, 1 eq) in dioxane (125 mL) was added HC1 / dioxane (4 M, 125 mL, 3.74 eq). The mixture was stirred at 50 °C for 12 h. The reaction mixture was concentrated under reduced pressure to give the crude product 4-((1 r,3r)-3-amino-2,2,4,4-tetramethyl-cyclobutoxy)-2-methoxy- benzonitrile (41.5 g, 134 mmol, 100% yield, HC1 salt) as a white solid and used in the next step without further purification. LC-MS: MS (ES+): RT = 0.492 min, m / z = 275.2 [M+H + ].

[0256] Part 4 - Preparation of 2-chloropyrimidine-5-carbonyl chloride

[0257]

[0258] To a solution of 2-chloropyrimidine-5-carboxylic acid (32.0 g, 202 mmol, 1.0 eq) in DCM (240 mL) was added oxalyl dichloride (76.9 g, 606 mmol, 53.0 mL, 3.0 eq) and DMF (738 mg, 10.1 mmol, 776 μί, 0.1 eq). The mixture was stirred at 40 °C for 1 h. The reaction mixture was concentrated under reduced pressure to give 2-chloropyrimidine-5-carbonyl chloride (35.7 g, 202 mmol, 99.94% yield) as a yellow oil which was used in the next step without further purification.

[0259] Part 5 - Preparation of 2-chloro-N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]pyrimidine-5-carboxamide

[0260]

[0261] To a solution of 4-((1 r,3 r)-3-amino-2,2,4,4-tetramethyl-cyclobutyloxy)-2- methoxy-benzonitrile (41.5 g, 134 mmol, 1.0 eq, HC1 salt) in THF (215 mL) was added DIEA (86.3 g, 668 mmol, 116 mL, 5.0 eq) at 0 °C, followed by 2-chloropyrimidine-5- carbonyl chloride (35.5 g, 200 mmol, 1.5 eq) in THF (215 mL). The mixture was stirred at 0 °C for 0.5 h. The mixture was poured into ice water (2 L), then extracted with EA (2 L x 3). The combined organic layers were washed with brine (2 L), dried, filtered, and concentrated under reduced pressure to give a residue. The crude product was wet milled with EA (500 mL) at 25 °C for 12 min to give 2-chloro-N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]pyrimidine-5-carboxamide (45.0 g, 108 mmol, 81% yield) as a yellow solid.

[0262] Part 6 - Preparation of N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12- tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide (II-5)

[0263]

[0264] To a solution of 4-((1 r,3 r)-3-amino-2,2,4,4-tetramethyl-cyclobutyloxy)-2- methoxy-benzonitrile (41.5 g, 134 mmol, 1.0 eq, HC1 salt) in THF (215 mL) was added DIEA (86.3 g, 668 mmol, 116 mL, 5.0 eq) at 0 °C, followed by 2-chloropyrimidine-5- carbonyl chloride (35.5 g, 200 mmol, 1.5 eq) in THF (215 mL). The mixture was stirred at 0 °C for 0.5 h. The mixture was poured into ice water (2 L), then extracted with EA (2 L x 3). The combined organic layers were washed with brine (2 L), dried, filtered, and concentrated under reduced pressure to give a residue. The crude product was wet milled with EA (500 mL) at 25 °C for 12 min to give 2-chloro-N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]pyrimidine-5-carboxamide (45.0 g, 108 mmol, 81% yield) as a yellow solid. 2,6To a solution of [2-(6),4,7,10,12-pentacosa-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (13.0 g, 24.6 mmol, 1.0 equiv) and 2-chloro-N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]pyrimidine-5-carboxamide (10.7 g, 25.8 mmol, 1.1 equiv) in NMP (30 mL) was added DIEA (9.53 g, 73.8 mmol, 12.9 mL, 3.0 equiv). The mixture was stirred at 25 °C for 8 h. The reaction mixture was diluted with DCM / MeOH (10:1, 300 mL) and washed with H2O (100 mL x 3). The organic phase was dried and concentrated. The crude product was purified by column chromatography (SiO2, R f = 0.4, EA / MeOH = 10 / 1) to give N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]thirteen-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5-carboxamide (19.5 g, 21.2 mmol, 87% yield, 98.5% purity). 1 H NMR (400 MHz, CD3OD) δ 8.74 (s, 2H), 7.91 (d, J = 0.8 Hz, 1H), 7.54 (d, J = 8.6 Hz, 1H), 7.34 (d, J = 8.8 Hz, 2H), 7.14 (d, J = 0.8 Hz, 1H), 6.85 (d, J = 8.9 Hz, 2H), 6.64 (d, J = 2.1 Hz, 1H), 6.57 (dd, J = 2.1, 8.6 Hz, 1H), 4.79 - 4.71 (m, 2H), 4.27 - 4.25 (m, 1H), 4.16 - 4.12 (m, 1H), 3.95 (s, 7H), 3.90 - 3.83 (m, 2H), 2.72 (s, 3H), 2.61 - 2.50 (m, 2H), 2.48 (s, 3H), 2.03 - 1.92 (m, 2H), 1.72 (s, 7H), 1.30 (s, 6H), 1.24 (s, 6H). LC-MS: MS (ES + ): RT = 2.698 min, m / z = 907.4 [M+H + ].

[0265] Part 7 - Preparation of N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3- thia- 1,8, 11, 12-tetraazatricyclo [8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine- 5-carboxamide crystalline Form A (II-5 in crystalline Form A)

[0266]

[0267] To N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4- [(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2 ,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine- 5-carboxamide (90.0 g, 99.2 mmol) was added CH3CN (450 mL) and then allowed to reflux for 1 hour until the reaction mixture became clear. The mixture was slowly cooled to 25 °C and a white precipitate formed. After filtration, the filter cake was washed with CH3CN (50 mL x 2) and dried to give N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4- [(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine- 5-carboxamide crystalline Form A (77.0 g, 84.9 mmol, 86% yield, 99.7% purity). The filtrate was concentrated to give a light brown foam (13.0 g, 13.5 mmol, 14% yield, 94% purity). LC-MS: MS (ES + ): RT = 0.667 min, m / z = 907.5 [M+H + ]. 1H NMR (400 MHz, CD3OD) δ 8.75-8.70 (m, 2H), 7.92-7.89 (m, 1H), 7.53 (d, J = 8.6 Hz, 1H), 7.32 (d, J = 8.7 Hz, 2H), 7.13 (s, 1H), 6.84 (d, J = 8.8 Hz, 2H), 6.63 (d, J = 2.2 Hz, 1H), 6.56 (dd, J = 2.1, 8.6 Hz, 1H), 4.84-4.81 (m, 1H), 4.77-4.71 (m, 1H), 4.25 (s, 1H), 4.13 (s, 1H), 3.99-3.89 (m, 7H), 3.88-3.82 (m, 2H), 2.71 (s, 3H), 2.58-2.50 (m, 2H), 2.46 (s, 3H), 2.02-1.93 (m, 2H), 1.74-1.66 (m, 7H), 1.28 (s, 6H), 1.23 (s, 6H).

[0268] Example 3 -N-[(1r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 Physical Characterization of Crystalline Form A of [7-(2,2-dimethyl-1-oxo-1,3,5-triazinan-7-yl)-5-oxo-4,5-dihydro-1H-pyrrol-2-yl]-(2-oxo-3-piperidinyl)methanone (II-1) as Crystalline Form A

[0269] X-ray powder diffraction was performed using a LYNXEYE_XE_T detector operating in reflection mode. The samples were prepared on a single crystal silicon, flat surface sample holder. The parameters for the XRPD were:

[0270]

[0271] Parameters Values Residence time (seconds / step) 0.4 Rotation Yes (15 rpm)

[0272] Figure 1 The X-ray powder diffraction pattern of the title compound is provided in FIG. 1. The table of the X-ray powder diffraction pattern is provided in Table 1 below, which lists the diffraction angles 2-theta, interplanar distances d, and relative intensities (expressed as a percentage relative to the strongest peak): Figure 1

[0273] X-ray powder diffraction pattern data

[0274]

[0275] ​

[0276] Differential scanning calorimetry analysis of the title compound was performed using a TA Discovery 2500 instrument at a nitrogen flow rate of 50 mL / min. The sample (1-2 mg) was weighed into a Tzero sample pan with a Tzero hermetic lid with a pinhole (0.7 mm diameter) and analyzed according to the following parameters: a ramp method, a heating rate of 10 °C / min, and a temperature range of 30 to 250 °C or until decomposition. Figure 2 The differential scanning calorimetry curve of the title compound obtained according to this procedure is provided in FIG. 1. A melting peak was observed at an onset temperature of 192.0 °C with a heat of fusion of 38 J / g.

[0277] Thermogravimetric analysis of the title compound was performed using a TA Discovery 5500 instrument. The nitrogen flow rate was 10 mL / min on the balance and 25 mL / min in the sample chamber. The sample (2-10 mg) was weighed directly into an open aluminum pan and analyzed according to the following parameters: a ramp method, a heating rate of 10.0 °C / min, and a temperature range of ambient temperature (below 35 °C) to 300 °C (or until the weight was <80%). Figure 3 The thermogravimetric analysis curve of the title compound is provided in FIG. 2. A weight loss of 0.2% w / w was observed at 180 °C.

[0278] The title compound was determined to be 99.7% pure by high pressure liquid chromatography (HPLC).

[0279] Storage stability

[0280] To investigate the effect of relative humidity on the physical stability of Compound II-5 in crystalline Form A, a sample of Compound II-5 in crystalline Form A was maintained at a temperature of 23-25 °C in an environment having a relative humidity level (RH) in the range of 23% RH to 53% RH. No change in the crystalline form was observed for Compound II-5 in crystalline Form A after a period of one week of storage under the foregoing conditions.

[0281] Example 4 - Synthesis of N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12- tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide in crystalline Form B (II-5 in crystalline Form B)

[0282] About 50 mg of amorphous N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine- 5-carboxamide was equilibrated in 0.15-0.35 mL EtOAc / MTBE solvent (1 / 4, v / v) on a magnetic stir plate with a stir bar at a rate of 600 rpm for 24 days. On the sixth day, about 1 mg of a nucleating agent was added to the mixture to help crystallization. On the twentieth day, about 1 mg of N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine- 5-carboxamide crystalline Form A was added to the mixture. On the twenty-fourth day, the resulting suspension was filtered through a 0.45 μιη membrane filter by centrifugation. The isolated solid was collected and determined to be Compound II-5 in crystalline Form B.

[0283] Example 5 -N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 Physical Characterization of N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0

[0284] The title compound was characterized using the instrumental methods described in Example 3.

[0285] Figure 4 The X-ray powder diffraction pattern obtained for the title compound is provided in Table 2 below. Figure 4 The table of X-ray powder diffraction pattern features is provided in Table 2 below, which lists the diffraction angle 2-theta, interplanar spacing d, and relative intensity (expressed as a percentage relative to the strongest peak):

[0286] X-ray powder diffraction pattern data

[0287]

[0288]

[0289] The title compound was subjected to differential scanning calorimetry analysis using the method described in Example 3. Figure 5 The differential scanning calorimetry curve of the title compound obtained according to this procedure is provided in Figure 6. A melting peak was observed at an onset temperature of 191.7 °C with a shoulder on the peak.

[0290] The title compound was subjected to thermogravimetric analysis using the method described in Example 3. Figure 6 The thermogravimetric analysis curve of the title compound is provided in Figure 7. A weight loss of 1.4% w / w was observed at 180 °C.

[0291] The title compound was determined to be 99.6% pure by high pressure liquid chromatography (HPLC).

[0292] Example 6 N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4- [(9S)-4,5, 13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8, 11, 12-tetraazatricyclo [8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide crystal Form A and N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4- [(9S)-4,5, 13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8, 11, 12-tetraazatricyclo [8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide crystal Form B

[0293] Approximately 3 mg of N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4- [(9S)-4,5, 13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8, 11, 12-tetraazatricyclo [8.3.0.0 2,6N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4- [(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 Compound II-5 in crystalline Form B was added to a saturated solution of 0.18 mL of the selected solvent. The resulting suspension was stirred at 25 °C and 50 °C, respectively, for 1 week. The solid fraction (wet cake) was isolated by centrifugal filtration at the respective temperature and investigated by XRPD.

[0294] The results are provided in the table below. The results show that Compound II-5 in crystalline Form B was converted to Compound II-5 in crystalline Form A in each of the solvent systems listed in the table below under stability study conditions. This demonstrates the superior stability of Compound II-5 in crystalline Form A over Compound II-5 in crystalline Form B.

[0295]

[0296] Example 7 - Synthesis of N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]-2-[2-[4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12- tetraazatricyclo[8.3.0.0 2,6 N-[(1 r,3 r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4- [(9S)-4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0

[0297]

[0298]

[0299] Step 1: Preparation of (3S)-4-[[3-(4-chlorobenzoyl)-4-ethyl-2- thienyl]amino]-3-(9H-fluoren-9-ylmethoxycarbonylamino)-4-oxo-butyric acid tert-butyl ester. To a solution of (2-amino-4-ethyl-3-thienyl)-(4-chlorophenyl)methanone (4.72 g, 17.3 mmol, 1.0 eq) in THF (40 mL) was added (2S)-4-tert-butoxy-2-(9H-fluoren-9-ylmethoxycarbonylamino)-4-oxo-butyric acid (7.28 g, 17.6 mmol, 1.0 eq) and pyridine (4.22 g, 53.1 mmol, 4.28 mL, 3.0 eq) at -15 °C, followed by POCl3(2.71 g, 17.6 mmol, 1.65 mL, 1.0 eq). The mixture was stirred at 0 °C for 3 h. The mixture was concentrated, the residue was diluted with HCl (1 M, 150 mL), and the mixture was extracted with ethyl acetate (200 mL x 3). The combined organic layers were washed with saturated sodium bicarbonate solution (200 mL), dried over sodium sulfate, filtered, and concentrated under reduced pressure to give (3S)-4-[[3-(4-chlorobenzoyl)-4-ethyl-2-thienyl]amino]-3-(9H-fluoren-9- ylmethoxycarbonylamino)-4-oxo-butyric acid tert-butyl ester (11.6 g, 17.6 mmol, 100% yield) as a yellow solid.

[0300] Step 2: Preparation of (3S)-3-amino-4-[[3-(4-chlorobenzoyl)-4-ethyl-2- thienyl]amino]-4-oxo-butyric acid tert-butyl ester. To a solution of (3S)-4-[[3-(4-chlorobenzoyl)-4-ethyl-2-thienyl]amino]-3-(9H-fluoren-9- ylmethoxycarbonylamino)-4-oxo-butyric acid tert-butyl ester (11.6 g, 17.6 mmol, 1.0 eq) in DCM (120 mL) was added piperidine (3.01 g, 35.8 mmol, 3.49 mL, 2.0 eq). The mixture was stirred at 25 °C for 0.5 h. The reaction mixture was concentrated under reduced pressure to give (3S)-3-amino-4-[[3-(4-chlorobenzoyl)-4-ethyl-2-thienyl]amino]-4-oxo-butyric acid tert-butyl ester (7.73 g, 17.6 mmol, 100% yield) as a colorless liquid.

[0301] Step 3: Preparation of tert-butyl 2-[(3S)-5-(4-chlorophenyl)-6-ethyl-2-oxo- 1,3-dihydrothieno[2,3-e][1,4]diazepine-3-yl]acetate. To a solution of tert-butyl (3S)-3-amino-4-[[3-(4-chlorobenzoyl)-4-ethyl-2- thienyl]amino]-4-oxo-butanoate (7.73 g, 17.7 mmol, 1.0 equiv) in EtOH (50 mL) was added HOAc (5.25 g, 87.4 mmol, 5.55 mL, 4.9 equiv). The mixture was stirred at 80 °C for 2 h. The reaction mixture was concentrated under reduced pressure to remove the solvent. The residue was made basic with aqueous sodium bicarbonate solution (150 mL) and the mixture was extracted with ethyl acetate (100 mL x 2). The combined organic layers were filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 22 / 1 to 3 / 1) to give tert-butyl 2-[(3S)-5-(4-chlorophenyl)-6-ethyl-2-oxo-1,3-dihydrothieno[2,3- e][1,4]diazepine-3-yl]acetate as a yellow solid (6.54 g, 15.5 mmol, 88% yield).

[0302] Step 4: Preparation of tert-butyl 2-[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.02,6]tetadeca-2(6),4,7,10,12-pentaen-9-yl]acetate. t-BuOK (1.0M, 18mL, 1.1 equivalent) was added to a solution of tert-butyl 2-[(3S)-5-(4-chlorophenyl)-6-ethyl-2-oxo-1,3-dihydrothieno[2,3-e][1,4]diazacycloheptatrien-3-yl]acetate (7.21 g, 16.7 mmol, 1.0 equivalent) in THF (70 mL) at -78 °C. The reaction mixture was slowly heated to 25 °C and stirred for 30 min. After cooling the mixture to -78°C, [chloro(phenoxy)phosphoryl]oxybenzene (5.39 g, 20.5 mmol, 4.16 mL, 1.2 equivalents) was added to the reaction mixture, and the mixture was slowly heated to 25°C over 30 minutes. Acetylhydrazine (1.97 g, 26.7 mmol, 1.6 equivalents) and EtOH (35 mL) were added to the reaction mixture, and it was stirred at 90°C for 1 hour. The reaction mixture was quenched by adding saturated NH4Cl solution (100 mL) at 0°C, and the mixture was diluted with H2O (200 mL) and extracted with ethyl acetate (250 mL × 2). The combined organic layers were dried over Na2SO4, filtered, and concentrated under reduced pressure to give the residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 1 to 0 / 1) to give 2-[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] as a white solid. 2,6 [Tetrate-2(6),4,7,10,12-pentene-9-yl]tert-butyl acetate (6.82 g, 14.9 mmol, 89% yield).

[0303] Step 5: Preparation of 2-[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [Decadecyl-2(6),4,7,10,12-pentaen-9-yl]acetic acid. To 2-[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6To a solution of tert-butyl [trideca-2(6),4,7,10,12-pentaen-9-yl]acetate (6.82 g, 14.8 mmol, 1.0 equiv) in DCM (40 mL) was added TFA (20 mL). The mixture was stirred at 25 °C for 0.5 h. The reaction mixture was concentrated under reduced pressure to give 2-[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]acetic acid (7.66 g, 14.8 mmol, crude, TFA salt).

[0304] Step 6: Preparation of 2-[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]-N-(2,2-dimethoxyethyl)acetamide. To a solution of 2-[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]acetic acid (7.66 g, 14.8 mmol, 1.0 equiv, TFA salt), 2,2-dimethoxyethylamine (1.88 g, 17.8 mmol, 1.94 mL, 1.2 equiv) in DCM (40 mL) was added TEA (9.03 g, 89.6 mmol, 12.2 mL, 6.0 equiv) and T4P (16.1 g, 22.3 mmol, 50% purity, 1.5 equiv) at 0 °C. The mixture was stirred at 25 °C for 12 h. The mixture was concentrated, and the residue was diluted with H2O (100 mL) and extracted with DCM (100 mL x 2). The combined organic layers were filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, Petroleum ether / Ethyl acetate = 1 / 1 to 0 / 1) to give 2-[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]-N-(2,2-dimethoxyethyl)acetamide (6.12 g, 12.5 mmol, 84% yield).

[0305] Step 7: Preparation of 2-[[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.02,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole. A solution of 2-[(9S)-7-(4- chlorophenyl)-5-ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]-N-(2,2-dimethoxyethyl)acetamide (6.11 g, 12.3 mmol, 1.0 equiv) in Eaton’s reagent (90.9 g, 381 mmol, 60 mL, 31.0 equiv) was stirred for 12 h. The mixture was slowly poured into a mixture of H2O (3.5 L) and NH4OH (1 L) at 0 °C, and the resulting mixture was extracted with ethyl acetate (200 mL x 3). The combined organic layers were filtered and concentrated under reduced pressure to give a residue. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 1 to 0 / 1) to give 2-[[(9S)-7-(4-chlorophenyl)-5-ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (2.65 g, 6.25 mmol, 51% yield).

[0306] Step 8: Preparation of 4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12- tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenol. To a solution of 2-[[(9S)-7-(4-chlorophenyl)-5- ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6A solution of [2-(2-aminophenyl)phenyl]-methylsulfonyloxy-palladium; dicyclohexyl-[3,6-dimethoxy-2-(2,4,6-triisopropylphenyl)phenyl]phosphane (641 mg, 707 μmol, 0.1 eq) and KOH (1.65 g, 29.4 mmol, 5.0 eq) in H2O (4 mL) was added to a solution of 4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenol (1.5 g, 3.7 mmol, 63% yield).

[0307] Step 9: Preparation of tert-butyl 2-[4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]nonane-7-carboxylate. To a solution of PPh3 (1.94 g, 7.42 mmol, 3.0 eq) in THF (10 mL) was added DIAD (997 mg, 4.93 mmol, 956 μL, 2.0 eq) at 0 °C. The mixture was stirred at 25 °C for 0.5 h. The mixture was added to a solution of 4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6[1.11 g, 2.47 mmol, 1.0 equivalent] phenol and tert-butyl 2-hydroxy-7-azaspiro[3.5]nonane-7-carboxylate (1.19 g, 4.93 mmol, 2.0 equivalent) in THF (10 mL). The mixture was stirred at 50 °C for 11.5 h. The reaction mixture was concentrated under reduced pressure. The residue was purified by column chromatography (SiO2, petroleum ether / ethyl acetate = 1 / 1 to 0 / 1) to give 2-[4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] as a white solid. 2,6 [1.55 g, 2.47 mmol, 100% yield] tert-butyl nonane-7-carboxylate (1.55 g, 2.47 mmol, 100% yield).

[0308] Step 10: Preparation of 2-[[(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-5-ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [Decadecyl-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole. To 2-[4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] 2,6 [1.54 g, 2.45 mmol, 1.0 equivalent]tetrazol-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]nonane-7-carboxylic acid tert-butyl ester (20 mL) was added to a solution of TFA (10 mL) in DCM (20 mL). The mixture was stirred at 25 °C for 0.5 h. The reaction mixture was concentrated under reduced pressure. The residue was made alkaline with an aqueous sodium bicarbonate solution and extracted with DCM / MeOH (10:1, 300 mL × 2). The combined organic layers were filtered and concentrated under reduced pressure to give 2-[[(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-5-ethyl-13-methyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0] as a yellow solid. 2,6 [Tetane-2(6),4,7,10,12-pentene-9-yl]methyl]oxazole (1.29 g, 2.44 mmol, crude material).

[0309] Step 11: Preparation of N-[(lr,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3- thia- 1,8, 11, 12-tetraazatricyclo [8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide (III-1). To a solution of 2-[[(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]- 5-ethyl-13-methyl-3-thia-l,8,l l,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (1.28 g, 2.44 mmol, 1.0 equiv) and 2-chloro-N-[(lr,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]pyrimidine-5-carboxamide (1.21 g, 2.92 mmol, 1.2 equiv) in NMP (5 mL) was added DIEA (629 mg, 4.87 mmol, 848 μL, 2.0 equiv). The mixture was stirred at 25 °C for 12 h. The reaction mixture was concentrated under reduced pressure. The residue was purified by preparative HPLC (column: Waters Xbridge BEH C18 250 x 50 mm x 10 μm; mobile phase: [water (NH4HCO3) - ACN]; gradient: 60% - 90% B, 20 min) to give N-[(lr,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]-2-[2-[4-[(9S)-5-ethyl-13-methyl-9-(oxazol-2-ylmethyl)-3-thia-l,8,l l,12- tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide (1.44 g, 1.57 mmol, 64% yield). 1H NMR (400 MHz, CD3OD) δ 8.72 (s, 2H), 7.89 (s, 1H), 7.52 (d, J = 8.56 Hz, 1H), 7.27-7.42 (m, 3H), 7.12 (s, 1H), 6.83 (d, J = 8.68 Hz, 2H), 6.62 (s, 1H), 6.55 (dd, J = 8.68, 1.83 Hz, 1H), 4.77-4.84 (m, 1H), 4.70-4.76 (m, 1H), 4.24 (s, 1H), 4.12 (s, 1H), 3.88-4.02 (m, 7H), 3.81-3.87 (m, 2H), 2.73 (s, 3H), 2.48-2.58 (m, 2H), 2.21-2.32 (m, 1H), 2.02-2.12 (m, 1H), 1.97 (dd, J = 12.78, 6.17 Hz, 2H), 1.66-1.71 (m, 4H), 1.28 (s, 6H), 1.22 (s, 6H), 0.97-1.01 (m, 3H). LC-MS: MS (ES + ): RT = 2.750 min, m / z = 907.6 [M+H + ]; LC-MS Method 25.

[0310] Example 8 - Synthesis of N-[(1 r,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]-2-[2-[4-[(9S)-4-(hydroxymethyl)-5,13-dimethyl-9-(oxazol-2-ylmethyl)-3- thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]nonan-7-yl]pyrimidine-5- carboxamide (III-2)

[0311]

[0312] Step 1 : Preparation of tert-butyl 2-[4-[(9S)-4-formyl-5,13-dimethyl-9-(oxazol-2-ylmethyl)- 3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]nonane-7-carboxylate. To a 15-mL vial equipped with a stir bar was added tert-butyl 2-[4-[(9S)-4,5,13-trimethyl-9- (oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6tert-butyl 2-[4-[(9S)-4-formyl-5, 13-dimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 10 O 32 (803mg, 242pmol, 0.2 eq). The mixture was stirred for 4 h and irradiated with a 365 nm LED lamp (7 cm distance) while the reaction temperature was maintained at 25 °C with a cooling fan. The mixture was filtered and concentrated. To the residue was added (n-Bu4N)4W 10 O 32 (80.3mg, 24.2pmol, 0.02 eq) and acetonitrile (3 mL) and the mixture was stirred for 4 h and irradiated with a 365 nm LED lamp (7 cm distance) while the reaction temperature was maintained at 25 °C with a cooling fan. The mixture was filtered, concentrated, and repeated once more in the same manner. The mixture was filtered and concentrated to give a residue. The residue was purified by preparative HPLC (column: Phenomenex luna C18 150 x 40 mm x 15 pm; mobile phase: [water (FA)-ACN]; gradient: 58-88% B, 10 min) to give tert-butyl 2-[4-[(9S)-4-formyl-5, 13-dimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]thirteen-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]nonane-7-carboxylate (75 mg, 117 pmol, 10% yield).

[0313] Step 2: Preparation of tert-butyl 2-[4-[(9S)-4-(hydroxymethyl)-5, 13-dimethyl-9- (oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]thirteen-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]nonane-7-carboxylate (75 mg, 117 pmol, 10% yield). 2,6To a solution of tert-butyl 2-[4-[(9S)-4-(hydroxymethyl)-5,13-dimethyl-9-(oxazol-2-ylmethyl)- 3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 tert-butyl 2-[4-[(9S)-4-(hydroxymethyl)-5,13-dimethyl-9-(oxazol-2-ylmethyl)- 3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0

[0314] Step 3: Preparation of [(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-5,13-dimethyl-9- (oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraaza-tricyclo[8.3.0.0 2,6 tert-butyl 2-[4-[(9S)-4-(hydroxymethyl)-5,13-dimethyl-9-(oxazol-2-ylmethyl)- 3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 To a solution of tert-butyl 2-[4-[(9S)-4-(hydroxymethyl)-5,13-dimethyl-9-(oxazol-2-ylmethyl)- 3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 tert-butyl 2-[4-[(9S)-4-(hydroxymethyl)-5,13-dimethyl-9-(oxazol-2-ylmethyl)- 3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0

[0315] Step 4: Preparation of N-[(lr,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4- tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4-(hydroxymethyl)-5,13-dimethyl-9-(oxazol-2- ylmethyl)-3-thia- 1,8, 11, 12-tetraazatricyclo [8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide (III-2). To a solution of [(9S)-7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]-5,13- dimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8, 11, 12-tetraazatricyclo [8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-4-yl]methanol (63.6 mg, 117 pmol, 1.0 equiv) and 2-chloro-N-[(lr,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]- pyrimidine-5-carboxamide (48.4 mg, 116.7 pmol, 1.0 equiv) in NMP (2 mL) was added DIEA (45.3 mg, 350 pmol, 61.0 pL, 3.0 equiv). The reaction mixture was stirred at 25 °C for 12 h. The residue was purified by preparative HPLC (column: Phenomenex luna C18 150 x 25 mm x 10 pm; mobile phase: [water (FA)-ACN]; gradient: 53-83% B, 10 min) to give N-[(lr,3r)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4-[(9S)-4- (hydroxymethyl)-5,13-dimethyl-9-(oxazol-2-ylmethyl)-3-thia- 1,8, 11, 12-tetraazatricyclo [8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide (50 mg, 54.17 pmol, 46% yield). 1H NMR (400 MHz, CD3OD δ 8.72 (s, 2H), 7.89 (s, 1H), 7.52 (d, J = 8.6 Hz, 1H), 7.36-7.29 (m, 2H), 7.12 (s, 1H), 6.85-6.80 (m, 2H), 6.62 (d, J = 2.1 Hz, 1H), 6.57-6.53 (m, 1H), 4.81-4.71 (m, 4H), 4.24 (s, 1H), 4.17-4.08 (m, 1H), 4.03-3.82 (m, 9H), 2.72 (s, 3H), 2.58-2.46 (m, 2H), 2.03-1.89 (m, 2H), 1.74 (s, 3H), 1.73-1.64 (m, 4H), 1.28 (s, 6H), 1.22 (s, 6H). LC-MS: MS (ES + ): RT = 2.45 min, m / z = 923.6 [M+H + ]; LC-MS Method 10.

[0316] Example 9 - Synthesis of N-[(1 r,3s)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl- cyclobutyl]-2-[2-[4-[4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6 ]trideca-2(6),4,7,10,12-pentaen-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5- carboxamide (III-3)

[0317]

[0318] The starting material was prepared analogously to the procedure described above. To 2-chloro-N-[(1 r,3s)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]pyrimidine-5- carboxamide (540 mg, 1.04 mmol, 1.0 equiv) and 2-[[7-[4-(7-azaspiro[3.5]non-2-yloxy)phenyl]- 4,5,13-trimethyl-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0 2,6[Decadecano-2(6),4,7,10,12-pentaen-9-yl]methyl]oxazole (550 mg, 1.04 mmol, 1.0 equivalent) was added to a solution of NMP (5.0 mL) with DIEA (269 mg, 2.08 mmol, 362 μL, 2.0 equivalent). The mixture was stirred at 40 °C for 12 hours. Water (30 mL) was added to the reaction mixture, and the mixture was extracted with ethyl acetate (50 mL × 3). The combined organic phases were washed with brine (30 mL), dried over anhydrous Na₂SO₄, filtered, and concentrated under vacuum. The residue was purified by preparative HPLC (column: Waters Xbridge C18 150×50mm×10μm; mobile phase: [water(NH4HCO3)-ACN]; gradient: 57%-87% B, 10 min) to obtain a white solid N-[(1r,3s)-3-(4-cyano-3-methoxy-phenoxy)-2,2,4,4-tetramethyl-cyclobutyl]-2-[2-[4-[4,5,13-trimethyl-9-(oxazol-2-ylmethyl)-3-thia-1,8,11,12-tetraazatricyclo[8.3.0.0]). 2,6 [13C-2(6),4,7,10,12-pentene-7-yl]phenoxy]-7-azaspiro[3.5]non-7-yl]pyrimidine-5-carboxamide (450 mg, 47% yield). 1 HNMR (400MHz, CDCl3): δ = 8.70 (s, 2H), 7.64 (s, 1H), 7.47-7.45 (m, 1H), 7.36-7.34 (m, 2H), 7.05 ( s,1H),6.76-6.74(m,2H),6.54-6.38(m,2H),5.94-5.92(m,1H),4.82-4.63(m,2H),4.16-4.04(m ,3H),4.00-3.98(m,1H),3.92(s,3H),3.89(s,2H),3.83-3.82(m,2H),2.68(s,3H),2.53-2.44(m ,2H),2.41(s,3H),2.05-1.98(m,2H),1.95-1.78(m,3H),1.71(s,4H),1.41(s,6H),1.09(s,6H). LC-MS:MS(ES + ): RT=2.704min, m / z=907.0[M+H + ]; LC-MS method 5-95.

[0319] Incorporation

[0320] The full disclosure of each of the patent documents and scientific articles referenced in this article is incorporated herein by reference for all purposes.

[0321] equivalents

[0322] The application may be embodied in other specific forms without departing from the spirit or essential characteristics thereof. The foregoing embodiments are therefore to be considered in all respects as illustrative only and not restrictive of the application described herein. The scope of the application is thus indicated by the appended claims, rather than by the foregoing description, and all changes that come within the meaning and range of equivalents are intended to be embraced therein.

Claims

1. A crystalline compound of Formula II-5:

2. The compound of claim 1, wherein the compound exhibits an X-ray powder diffraction pattern comprising peaks at the following diffraction angles (2Q): 13.1 ± 0.2, 17.6 ± 0.2, 18.3 ± 0.2, 18.7 ± 0.2, 19.5 ± 0.2, 23.8 ± 0.2, and 25.6 ± 0.

2.

3. The compound of claim 2, wherein the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 8.1 ± 0.

2.

4. The compound of claim 2 or 3, wherein the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 9.9 ± 0.

2.

5. The compound of any one of claims 2-4, wherein the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 10.5 ± 0.

2.

6. The compound of any one of claims 2-5, wherein the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 15.8 ± 0.

2.

7. The compound of any one of claims 2-6, wherein the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 19.8 ± 0.

2.

8. The compound of any one of claims 2-7, wherein the X-ray powder diffraction pattern further comprises a peak at the following diffraction angle (2Q): 25.3 ± 0.

2.

9. The compound of any one of claims 2-8, wherein the relative intensity of the peak at the diffraction angle (2Q) is at least 20%.

10. The compound of any one of claims 2-8, wherein the relative intensity of the peak at the diffraction angle (2Q) is at least 30%.

11. The compound of claim 1, characterized by the following X-ray powder diffraction pattern expressed in terms of diffraction angle 2Q, interplanar spacing d, and relative intensity (expressed as a percentage relative to the most intense peak):

12. The compound of claim 1, wherein the compound has an X-ray powder diffraction pattern substantially as shown in FIG.

1.

13. The compound of any one of claims 1-12, wherein the compound has a melting point onset in the range of about 180 °C to about 200 °C as determined by differential scanning calorimetry.

14. The compound of any one of claims 1-12, wherein the compound has a melting point onset of about 192 °C as determined by differential scanning calorimetry.

15. The compound of any one of claims 1-12, wherein the compound has a differential scanning calorimetry curve substantially the same as shown in FIG.

2.

16. The compound of claim 1, wherein the compound exhibits an X-ray powder diffraction pattern comprising peaks at the following diffraction angles (2Q): 9.4 ± 0.2, 12.8 ± 0.2, 15.4 ± 0.2, 18.2 ± 0.2, 18.7 ± 0.2, 20.2 ± 0.2, and 24.6 ± 0.

2.

17. The compound of claim 16, wherein the X-ray powder diffraction pattern further comprises one or more peaks at diffraction angles (2 theta) of 8.5 ± 0.2, 11.7 ± 0.2, 12.5 ± 0.2, 17.6 ± 0.2, 19.6 ± 0.2, 21.9 ± 0.2, and 23.5 ± 0.

2.

18. The compound of claim 16 or 17, wherein the relative intensity of the peaks at diffraction angles (2 theta) is at least 20%.

19. The compound of claim 16 or 17, wherein the relative intensity of the peaks at diffraction angles (2 theta) is at least 30%.

20. The compound of claim 1, characterized by the following X-ray powder diffraction pattern expressed in terms of diffraction angle 2 theta, interplanar spacing d, and relative intensity (expressed as a percentage relative to the most intense peak):

21. The compound of claim 1, wherein the compound has an X-ray powder diffraction pattern substantially as shown in FIG.

4.

22. The compound of any one of claims 16-21, wherein the compound has a melting point onset in the range of about 180 °C to about 200 °C as determined by differential scanning calorimetry.

23. The compound of any one of claims 16-21, wherein the compound has a melting point onset of about 192 °C as determined by differential scanning calorimetry.

24. The compound of any one of claims 16-21, wherein the compound has a differential scanning calorimetry curve substantially the same as shown in FIG.

5.

25. The compound of any one of claims 1-24, wherein the compound has a purity greater than 98% by weight.

26. The compound of any one of claims 1-24, wherein the compound has a purity greater than 99% by weight.

27. The compound of any one of claims 1-24, wherein the compound has a purity greater than 99.5% by weight.

28. A compound in Table 3, or a pharmaceutically acceptable salt thereof.

29. A pharmaceutical composition comprising a compound of any one of claims 1-28 and a pharmaceutically acceptable carrier.

30. A method of treating cancer, the method comprising administering to a patient in need thereof a therapeutically effective amount of a compound of any one of claims 1-28 to treat the cancer.

31. The method of claim 30, wherein the cancer is ovarian cancer, uterine cancer, endometrial cancer, cervical cancer, prostate cancer, testicular cancer, breast cancer, brain cancer, lung cancer, oral cancer, esophageal cancer, head and neck cancer, stomach cancer, colon cancer, rectal cancer, skin cancer, sebaceous gland cancer, bile duct cancer, gallbladder cancer, liver cancer, pancreatic cancer, bladder cancer, urinary tract cancer, kidney cancer, eye cancer, thyroid cancer, lymphoma, or leukemia.

32. The method of claim 30, wherein the cancer is prostate cancer.

33. The method of any one of claims 30-32, wherein the patient is a human.

34. A method of causing death of a cancer cell, the method comprising contacting a cancer cell with an effective amount of a compound of any one of claims 1-28 to cause death of the cancer cell.

35. The method of claim 34, wherein the cancer cell is selected from an ovarian cancer, uterine cancer, endometrial cancer, cervical cancer, prostate cancer, testicular cancer, breast cancer, brain cancer, lung cancer, oral cancer, esophageal cancer, head and neck cancer, stomach cancer, colon cancer, rectal cancer, skin cancer, sebaceous gland cancer, bile duct cancer, gall bladder cancer, liver cancer, pancreatic cancer, bladder cancer, urinary tract cancer, kidney cancer, eye cancer, thyroid cancer, lymphoma, or leukemia cell.

36. The method of claim 34, wherein the cancer cell is a prostate cancer cell.

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