MONOCYCLIC HETEROARYL COMPOUNDS

Abstract

This invention relates to compounds of the general formula:

##STR00001## in which the variable groups are as defined herein, and to their preparation and use.

Claims

1. A compound of the Formula I, a tautomer, an individual isomer, a mixture of isomers or a pharmaceutically acceptable salt, solvate or hydrate thereof: ##STR00058## wherein: Ring T represents a 5-membered monocyclic heteroaryl ring, comprising 1-3 heteroatoms selected from O, N and S and being optionally substituted with 1-3 R.sup.t groups; Ring A represents a 5- or 6-membered aryl or heteroaryl ring and is optionally substituted with 1-4 R.sup.a groups; Ring B represents a 5- or 6-membered aryl or heteroaryl ring and is optionally substituted with 1-5 R.sup.b groups; L.sup.1 is selected from NR.sup.1C(O) and C(O)NR.sup.1; At each occurrence, R.sup.a, R.sup.b and R.sup.t, is independently selected from the group consisting of halo, —CN, —NO.sub.2, —R.sup.4, —OR.sup.2, —NR.sup.2R.sup.3, —C(O)YR.sup.2, —OC(O)YR.sup.2, —NR.sup.2C(O)YR.sup.2, —SC(O)YR.sup.2, —NR.sup.2C(═S)YR.sup.2, —OC(═S)YR.sup.2, —C(═S)YR.sup.2, —YC(═NR.sup.3)YR.sup.2, —YP(═O)(YR.sup.4)(YR.sup.4), —Si(R.sup.4).sub.3, —NR.sup.2SO.sub.2R.sup.2, —S(O).sub.rR.sup.2, —SO.sub.2NR.sup.2R.sup.3 and —NR.sup.2SO.sub.2NR.sup.2R.sup.3, wherein Y is independently a bond, —O—, —S— or —NR.sup.3—; R.sup.1, R.sup.2 and R.sup.3 are independently selected from H, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heterocycle and heteroaryl; Alternatively, NR.sup.2R.sup.3 moiety may be 5- or 6-membered saturated, partially saturated or unsaturated ring, which can be optionally substituted and which contains 0-2 additional heteroatoms selected from N, O and S(O).sub.r; each occurrence of R.sup.4 is independently selected from alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heterocyclic and heteroaryl; each of the foregoing alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, heterocyclic and heteroaryl moieties is optionally substituted; m is 0, 1, 2, 3 or 4; n is 0, 1, 2 or 3; p is 0, 1, 2, 3, 4 or 5; r is selected 0, 1 or 2.

2. A compound according to claim 1 wherein Ring T is a monocyclic 5-membered ring selected from: ##STR00059## and is optionally substituted on carbon or heteroatom with 1-3 R.sup.t groups.

3. A compound of claim 1 having the Formula: ##STR00060## wherein: Ring C represents a 5- or 6-membered heterocyclic or heteroaryl ring, comprising carbon atoms and 1-3 heteroatoms selected from O, N and S(O).sub.r and is optionally substituted on carbon or heteroatom(s) with 1-5 R.sup.c groups; R.sup.c, at each occurrence, is independently selected from the group consisting of halo, ═O, ═S, —CN, —NO.sub.2, —R.sup.4, —OR.sup.2, —NR.sup.2R.sup.3, —C(O)YR.sup.2, —OC(O)YR.sup.2, —NR.sup.2C(O)YR.sup.2, —SC(O)YR.sup.2, —NR.sup.2C(═S)YR.sup.2, —OC(═S)YR.sup.2, —C(═S)YR.sup.2, —YC(═NR.sup.3)YR.sup.2, —YP(═O)(YR.sup.4)(YR.sup.4), —Si(R.sup.4).sub.3, —NR.sup.2SO.sub.2R.sup.2, —S(O).sub.rR.sup.2, —SO.sub.2NR.sup.2R.sup.3 and —NR.sup.2SO.sub.2NR.sup.2R.sup.3, wherein Y is independently a bond, —O—, —S— or —NR.sup.3—; R.sup.2 and R.sup.3 are independently selected from H, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heterocyclic and heteroaryl; alternatively, NR.sup.2R.sup.3 moiety may be a 5- or 6-membered saturated, partially saturated or unsaturated ring, which can be optionally substituted and which contains 0-2 additional heteroatoms selected from N, O and S(O).sub.r; each occurrence of R.sup.8 is independently selected from alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heterocyclic and heteroaryl; each of the foregoing alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heteroaryl and heterocycle moieties is optionally substituted; v is 0, 1, 2, 3, 4 or 5 t is 0, 1, 2, 3, or 4.

4. A compound according to claim 3, wherein Rings A and B are aryl.

5. A compound of claim 3 wherein Ring C is a heteroaryl ring.

6. A compound of claim 5 wherein Ring C is an imidazole ring.

7. A compound of claim 3 selected from the formulae IIa, IIb and IIc: ##STR00061##

8. A compound of claim 7 wherein R.sup.t is independently selected from —CH.sub.3, or —C(O)NH.sub.2, v is 1, n is 0 or 1, m is 1, t is 1, R.sup.a is —CH.sub.3, R.sup.b is CF.sub.3 and R.sup.c is —CH.sub.3.

9. A compound of claim 1 having the Formula: ##STR00062## wherein: Ring D represents a 5- or 6-membered heterocyclic or heteroaryl ring, comprising carbon atoms and 1-3 heteroatoms independently selected from N, O, S(O).sub.r and is optionally substituted with 1-5 R.sup.d groups; L.sup.2 is (CH.sub.2).sub.2, O(CH.sub.2).sub.x, NR.sup.3(CH.sub.2).sub.x, S(CH.sub.2).sub.x or (CH.sub.2).sub.xNR.sup.3C(O)(CH.sub.2).sub.x and the linkage unit can be used in either direction; R.sup.d, at each occurrence, is selected from the group consisting of halo, ═O, ═S, —CN, —NO.sub.2, —R.sup.4, —OR.sup.2, —NR.sup.2R.sup.3, —C(O)YR.sup.2, —OC(O)YR.sup.2, —NR.sup.2C(O)YR.sup.2, —SC(O)YR.sup.2, —NR.sup.2C(═S)YR.sup.2, —OC(═S)YR.sup.2, —C(═S)YR.sup.2, —YC(═NR.sup.3)YR.sup.2, —YP(═O)(YR.sup.4)(YR.sup.4), —Si(R.sup.4).sub.3, —NR.sup.2SO.sub.2R.sup.2, —S(O).sub.rR.sup.2, —SO.sub.2NR.sup.2R.sup.3 and —NR.sup.2SO.sub.2NR.sup.2R.sup.3, wherein Y is independently a bond, —O—, —S— or —NR.sup.3—; R.sup.2 .sub.and R.sup.3 are independently selected from H, alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heterocyclic and heteroaryl; alternatively, NR.sup.2R.sup.3 moiety may be a 5- or 6-membered saturated, partially saturated or unsaturated ring, which can be optionally substituted and which contains 0-2 additional heteroatoms selected from N, O and S(O).sub.r; each occurrence of R4 is independently selected from alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heterocyclic and heteroaryl; each of the foregoing alkyl, alkenyl, alkynyl, cycloalkyl, cycloalkenyl, cycloalkynyl, aryl, heteroaryl and heterocycle moieties is optionally substituted; w is selected from 0, 1, 2, 3, 4 or 5; x is 0, 1, 2 or 3; z is 1, 2, 3 or 4; and t is 0, 1, 2, 3, or 4.

10. A compound according to claim 9 wherein Rings A and B are aryl.

11. A compound of claim 9 wherein Ring D is a substituted or unsubstituted piperazine ring and L.sup.2 is CH.sub.2.

12. A compound of claim 9 wherein Ring D is a substituted or unsubstituted heteroaryl.

13. A compound of claim 11 selected from Formulae IIIa, IIIb and IIIc: ##STR00063##

14. A compound of claim 13 wherein R.sup.t is independently selected from —CH.sub.3 and —C(O)NH.sub.2, n is 0 or 1, m is 1, t is 1, R.sup.a is methyl, R.sup.b is CF.sub.3, one of R.sup.d is selected from methyl and CH.sub.2CH.sub.2OH.

15. A method for treating cancer in a mammal in need thereof, comprising administering to the mammal a therapeutically effective amount of a compound of any of claims 1-14 or a pharmaceutically acceptable salt, solvate or hydrate thereof.

16. A composition comprising a compound of any of claims 1-14 or a pharmaceutically acceptable salt, solvate or hydrate thereof and a pharmaceutically acceptable carrier, diluent or vehicle.

Description

EXAMPLES

[0165] Some of the following compounds have been converted into HCl salt. The general procedure for generating HCl salts is described below:

[0166] To the final product was added Just enough MeOH saturated with HCl (g) to dissolve, cooled to 0° C. for 0.5-1 h, filtered, washed solid with ice cold MeOH then Et.sub.2O, and the resulting solid dried in a vacuum desiccator to provide in most cases the tris HCl salt.

Example 1

1-(1H-Pyrazole-1-carboxamide-N-methyl)-4-methyl-N-(4-((4-methylpiperazin-1-yl)methyl)-3-(trifluoromethyl)phenyl)benzamide

[0167] ##STR00043##

[0168] 1-(Bromomethyl)-4-nitro-2-(trifluoromethyl)benzene: A suspension of 2-methyl-5-nitrobenzotrifluoride (3.90 g, 19 mmol), N-bromosuccinimide (NBS, 3.56 g, 20 mmol), 2,2′-azobis(2-methylpropionitrile) (AIBN, 94 mg, 0.6 mmol) in CCl.sub.4 (40 mL) was refluxed under N.sub.2 for 16 h. HPLC indicated ca. 50% conversion. More NBS (10 mmol) and AIBN (0.6 mmol) was added, and the mixture was refluxed for another 14 h. HPLC indicated ca. 80% conversion. The reaction mixture was cooled down, and the solid was filtered off and washed with EtOAc. The combined filtrate was washed with aq. NaHCO.sub.3, dried over Na.sub.2SO.sub.4, filtered, concentrated on rotovap and further dried under vacuum. 1H NMR shows the ratio of desired product to unreacted 2-methyl-5-nitrobenzotrifluoride is 75:25. This material was not purified but used directly in the next step.

[0169] 1-Methyl-4-(4-nitro-2-(trifluoromethyl)benzyl)piperazine: To a solution of crude 1-(bromomethyl)-4-nitro-2-(trifluoromethyl)benzene (13.33 mmol, 75% pure) in DCM (10 mL) was added Et.sub.3N (1.4 mL, 10 mmol) and 1-methylpiperazine (1.1 mL, 10 mmol). After stirring for 3 h at rt, aq. NaHCO.sub.3 was added, and the mixture was extracted with DCM. The combined organic layer was dried over Na.sub.2SO.sub.4, filtered, concentrated, and the resulting residue was purified by silica gel chromatography (eluted with 10% MeOH/DCM) to provide 2.21 g of product as a pale yellow oil.

[0170] 4((4-Methylpiperazin-1-yl)methyl-3-(trifluoromethyl)aniline: A suspension of 1-methyl-4-(4-nitro-2-(trifluoromethyl)benzyl)piperazine (1.23 g, 4 mmol) and sodium hydrosulfite (7.0 g, 85% pure from Aldrich, 40 mmol) in acetone and water (1:1, 20 mL) was refluxed for 3 h. Upon cooling, the volatile components (mainly acetone) were removed on rotavap, and the resulting mixture was subjected to filtration. The solid was thoroughly washed with EtOAc. The combined filtrate was extracted with n-BuOH (4×), and the combined organic layer was washed with saturated aq. NaHCO.sub.3, dried (Na.sub.2SO.sub.4), filtered, concentrated, and the resulting residue was purified by silica gel chromatography (eluted with 5% MeOH/DCM, MeOH was pre-saturated with ammonia gas) to provide 0.71 g of product as a pale yellow solid.

[0171] 3-iodo-4-methyl-N-(4-((4-methylpiperazin-1-yl)methyl)-3-(trifluoromethyl)phenyl) Benzamide: 3-iodo-4-methylbenzoyl chloride (0.48 g, 1.7 mmol), prepared from the reaction of 3-iodo-4-methylbenzoic acid and SOCl.sub.2 (as previously described), was added to a solution of 4-((4-methylpiperazin-1-yl)methyl)-3-(trifluoromethyl)aniline (0.47 g, 1.7 mmol), N,N-diisopropylethylamine (0.26 g, 2.0 mmol), and a catalytic amount of DMAP in THF (10 mL). After stirring at rt for 2 h, the reaction was quenched with water. EtOAc was added and the layers separated. The combined organic layers were concentrated to dryness and purified by silica gel chromatography (eluted with 5% MeOH/DCM, MeOH was pre-saturated with ammonia gas), to provide 0.51 g of product as an off-white solid.

[0172] 4-Methyl-N-[4-(4-methyl-piperazin-1-ylmethyl)-3-trifluoromethyl-phenyl]-3-trimethylsilanylethynyl-benzamide: 3-iodo-4-methyl-N-[4-(4-methyl-piperazin-1-ylmethyl)-3-trifluoromethyl-phenyl]-benzamide (2.59 g, 5 mmol), Pd[(PPh.sub.3)].sub.4 (289 mg, 0.25 mmol), Cul (71 mg, 0.375 mmol) was placed in a schlenk flask. The flask was subjected to 3 cycles of vacuum-refilling with N.sub.2. To this mixture was added anhydrous N,N-diisopropylethylamine (1.1 mL, 6 mmol), DMF (5 mL), and trimethylsilylacetylene (0.92 mL, 6.5 mmol). This solution was stirred at it for 24 h. Water and EtOAc were added to the reaction mixture to facilitate the extraction. The combined organic layers were dried over Na.sub.2SO.sub.4, filtered, and then concentrated on a rotavap and the residue was purified on a silica gel column (eluent: 5% MeOH in CH.sub.2Cl.sub.2, MeOH was pre-saturated with ammonia gas) to give the desired product as a light yellow solid in 82% yield (2.0 g).

[0173] 3-Ethynyl-4-methyl-N-[4-(4-methyl-piperazin-1-ylmethyl)-3-trifluoromethyl-phenyl]-benzamide: To a solution of 4-methyl-N-[4-(4-methyl-piperazin-1-ylmethyl)-3-trifluoromethyl-phenyl]-3-trimethylsilanylethynyl-benzamide (2.0 g, 4.1 mmol) in THF (15 mL) was added 5 mL of TBAF in THF (1.0M). After stirring at it for 1 h, the mixture was partitioned between H.sub.2O and EtOAc. The combined organic layers were dried over Na.sub.2SO.sub.4, filtered, and then concentrated on a rotavap and the residue was purified on a silica gel column (eluent: 10% MeOH in CH.sub.2Cl.sub.2, MeOH was pre-saturated with ammonia gas) to give the desired product as a light yellow solid in 78% yield (1.33 g).

[0174] 1H-Pyrazole-1-N-methyl carboxamide: To 4-iodopyrazole (3 g, 15.5 mmol) in dry CH.sub.2Cl.sub.2 (20 mL) was added p-nitrophenyl chloroformate (3.43 g, 17 mmol) in CH.sub.2Cl.sub.2 (30 mL) followed by TEA (2.5 mL, 18.6 mmol). This mixture was stirred at rt for 2 h. The mixture was then diluted with excess CH.sub.2Cl.sub.2 (60 mL) and washed with 10% aq. NaHCO.sub.3. The organic layer was dried over Na.sub.2SO.sub.4 and the solvent was evaporated to yield a solid which was recrystallized from Et.sub.2O to furnish a white solid (5 g). The resulting material (1 g, 2.79 mmol) in THF (12 mL) was treated with a solution of MeNH.sub.2 (2 mL, 2M in THF) for 10 minutes. The organic layer was evaporated to a solid which was dissolved in DCM and washed with 5% NaOH. The organic layer was dried, filtered and triturated with ether to afford the product as colorless shiny flakes (0.55 g).

[0175] 3-(1H-Pyrazole-1-carboxamide-N-methyl)-4-methyl-N-(4-((4-methylpiperazin-1-yl)methyl)-3-(trifluoromethyl)phenyl)benzamide: 3-Ethynyl-4-methyl-N-[4-(4-methyl-piperazin-1-ylmethyl)-3-trifluoromethyl-phenyl]-benzamide (91 mg, 0.22 mmol), 1H-Pyrazole-1-N-methyl carboxamide (0.2 mmol), Pd[(PPh.sub.3)].sub.4 (11.6 mg, 0.01 mmol), Cul (2.9 mg, 0.015 mmol) was placed in a vial capped with rubber septa. This vial was subjected to 3 cycles of vacuum-refilling with N.sub.2. To this mixture was added anhydrous N,N-diisopropylethylamine (0.1 mL, 0.6 mmol) and DMF (1.0 mL). The resulting solution was stirred at 80° C. for 24 h. After the reaction mixture was cooled, water and EtOAc were added to facilitate the extraction. The combined organic layers were dried over Na.sub.2SO.sub.4, filtered, and then concentrated on a rotavap and the residue was purified on a silica gel column (eluent: 10% MeOH in CH.sub.2Cl.sub.2, MeOH was pre-saturated with NH.sub.3 gas) to give the desired product as a light yellow solid in 56% yield (63.0 mg): MS (M+H).sup.+ 538.

Example 2

1Methyl-5-(2-methyl-5-[4-(4-methyl-piperazin-1-ylmethyl)-3-trifluoromethyl-phenylcarbamoyl]-phenylethynyl)-1H-imidazole-2-carboxylic acid amide

[0176] ##STR00044##

[0177] 5-Ethynyl-1-methyl-1H-imidazole-2-carboxylic acid amide: To a solution of 1-methyl-5-trimethylsilanylethynyl-1H-imidazole (1.78 g, 10 mmol) in THF (30 mL) was slowly added a solution of n-BuLi in hexanes (2.5M, 4.4 mL) at −78° C. and the resulting suspension was stirred at the same temperature for 1 h. Trimethylsilyl isocyanate (from Aldrich, 85% pure, 11.76 mmol, 1.6 mL) was slowly added to this suspension at −78° C. After stirring at −78° C. for another 30 minutes the reaction mixture was allowed to warm up slowly by removing the cooling bath. Water (2 mL) and MeOH (1 mL) was added to quench the reaction and the mixture was stirred overnight. More water and EtOAc was added to facilitate the extraction. The combined organic layers were dried over Na.sub.2SO.sub.4, filtered, and then concentrated on a rotavap and the residue was purified on a silica gel column (eluent: 40-60% EtOAc in hexanes) to give the desired product as a white solid in 26% yield (387 mg).

[0178] 1-Methyl-5-(2-methyl-5-[4-(4-methyl-piperazin-1-ylmethyl)-3-trifluoromethyl-phenylcarbamoyl]-phenylethynyl)-1H-imidazole-2-carboxylic acid amide: 5-Ethynyl-1-methyl-1H-imidazole-2-carboxylic acid amide (33 mg, 0.22 mmol), 3-iodo-4-methyl-N-(4-((4-methylpiperazin-1-yl)methyl)-3-(trifluoromethyl)phenyl)benzamide as prepared in example 1 (103.4 mg, 0.2 mmol), Pd[(PPh.sub.3).sub.4] (11.6 mg, 5 mol %), and Cul (2.9 mg, 7.5 mmol %) were placed in a vial with rubber septum. The mixture underwent 3 cycles of vacuum/filling with N.sub.2, and DMF (1.5 ml) and N,N-diisopropylethylamine (53 μL, 0.3 mmol) were added. The mixture was stirred at rt for 16 h, and the reaction was quenched with H.sub.2O. EtOAc and more water were added for extraction. The combined organic layer was dried (Na.sub.2SO.sub.4), filtered, concentrated, and the resulting residue was purified by silica gel chromatography (eluent: 5% MeOH in methylene chloride, MeOH was pre-saturated with ammonia gas), giving the titled compound as an off-white solid (65%, 70 mg): MS (M+H).sup.+ 539.

Example 3

1Methyl-5-(2-methyl-5-[3-(4-methyl-imidazol-1-yl)-5-trifluoromethyl-phenylcarbamoyl]-phenylethynyl)-1H-imidazole-2-carboxylic acid amide

[0179] ##STR00045##

[0180] The title compound was made as for example 2 using 5-ethynyl-1-methyl-1H-imidazole-2-carboxylic acid amide and 3-iodo-4-methyl-N-(3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl)benzamide: MS (M+H).sup.+ 507.

[0181] 3-(4-Methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)benzenamine: A suspension of 3-bromo-5-(trifluoromethyl)aniline (4.8 g, 20 mmol), 4-methylimidazole (1.97 g, 24 mmol), potassium carbonate (3.04 g, 22 mmol), Cul (0.57 g, 3 mmol), and 8-hydroxyquinoline (0.44 g, 3 mmol,) in dry DMSO (20 mL) in a pressure tube was degassed by bubbling N.sub.2 into the suspension for 10 minutes while stirring. The tube was sealed tightly. The mixture was heated at 120° C. (oil bath temperature) for 15 h. The mixture was cooled down to 45-50° C. and 14% aq. NH.sub.4OH (20 mL) was added. The mixture was maintained at this temperature for 1 h. After cooling to rt, water and ethyl acetate were added. The aqueous layer was extracted with ethyl acetate and the combined organic layers were passed through a short silica gel column to remove most of green/blue Cu salts. The filtrate was dried over sodium sulfate and concentrated on a rotavap. The crude product was recrystallized from EtOAc/hexanes, giving pure pale yellow needles. The mother liquor was concentrated and the residue was purified on silica gel column (5% methanol/methylene chloride), yielding a second crop as pale yellow needles.

[0182] 3-iodo-4-methyl-N-(3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl) Benzamide: 3-iodo-4-methylbenzoic acid (2.62 g, 10 mmol) was refluxed in SOCl.sub.2 (10 mL) for 1 h. The volatile components were removed on a rotavap and the residue was dissolved in benzene (10 mL), concentrated to dryness on a rotavap and further dried under vacuum. The resulting acyl chloride was added to a solution 3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)benzenamine (2.46 g, 10.2 mmol), N,N-diisopropylethylamine (1.56 g, 12 mmol), and a catalytic amount of DMAP in THF (20 mL). After stirring at rt for 2 h, the reaction was quenched with water. EtOAc was added and the layers separated. The combined organic layers were concentrated to dryness and used without purification in the coupling step.

Example 4

5-[5-(3-imidazol-1-yl-5-trifluoromethylphenylcarbamoyl)-2-methyl-phenylethynyl]-1-methyl-1H-imidazole-2-carboxylic acid amide

[0183] ##STR00046##

[0184] This was made as for example 2 using 5-ethynyl-1-methyl-1H-imidazole-2-carboxylic acid amide and N-(3-(1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl)-3-iodo-4-methylbenzamide: MS (M+H).sup.+ 493.

[0185] 3-(1H-imidazol-1-yl)-5-(trifluoromethyl)aniline: A mixture of 3-Amino-5-bromobenzotrifluoride (4.0 g, 0.0167 mol), 8-hydroxy quinoline (0.362 g, 0.0025 mol), Cul (0.476 g. 0.025 mol), imidazole (1.36 g, 0.0199 mol), and potassium carbonate (2.52 g, 0.0183 mol) in 17 mL of DMSO (degassed with argon for −10 min) was heated at 120° C. under an atmosphere of argon for 15 h; the HPLC indicated no starting material. A 14% aqueous solution of ammonium hydroxide was added to the cooled mixture and this was stirred for 1 h at ambient temperature. Water (50 mL) and EtOAc (200 mL) were added and the aqueous layer was extracted with EtOAc (3×30 mL). The combined organic layers were dried over Na.sub.2SO.sub.4 and concentrated. The crude product was purified by silica gel flash chromatography (eluted with EtOAc/hexanes) to provide 2.51 g of product.

[0186] N-(3-(1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl)-3-iodo-4-methylbenzamide: To 3-iodo-4-methylbenzoic acid (3.07 g, 0.0117 mol) was added thionyl chloride (10 mL) and refluxed for 2 h. The excess thionyl chloride was carefully removed and the resulting acid chloride was dried in vacuo for 2 h. The residue was then dissolved in DCM (anhydrous, 25 mL) and cooled on ice. To the cooled solution was added 3-(1H-imidazol-1-yl)-5-(trifluoromethyl)aniline (3.46 g, 0.0152 mol) in DCM followed by the dropwise addition of diisopropylethylamine (8.2 mL, 0.047 mol). This was stirred at ambient temperature for 21 h. The white solid that separated was filtered and washed with water and dried to provide 4.65 g of product. Additional product could be obtained from the filtrate following concentration and purification by silica gel flash chromatography in EtOAc/hexanes.

Example 5

3-[(2-amino-1,3-thiazol-5yl)ethynyl]-4-methyl-N-[3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl]benzamide

[0187] ##STR00047##

[0188] tert-butyl {5-[(trimethylsilyl)ethynyl]-1,3-thiazol-2-yl}carbamate: A mixture of tert-butyl (5-bromo-1,3-thiazol-2-yl)carbamate (2.79 g, 10 mmol), ethynyltrimethylsilane (1.27 g, 13 mmol), Pd(PPh.sub.3).sub.4 (578 mg, 0.5 mmol), Cul (143 mg, 0.75 mmol), and diisopropylethylamine (1.94 g, 15 mmol) in DMF (10 mL) was heated at 50° C. overnight under an atmosphere of N.sub.2. Upon cooling to ambient temperature, the reaction mixture was concentrated and the crude product was purified by silica gel flash chromatography (eluted with 20% EtOAc/hexanes) to give a yellow solid (2.55 g, 86%).

[0189] tert-butyl {5-[(2-methyl-5-{[3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl]carbamoyl}phenyl)ethynyl]-1,3-thiazol-2-yl}-carbamate: A mixture of tert-butyl {5-[(trimethylsilyl)ethynyl]-1,3-thiazol-2-yl}carbamate (166 mg, 0.56 mmol), 3-iodo-4-methyl-N-(3-(4-methyl-1H-imidazol-1-yl)-5- (trifluoromethyl)phenyl)benzamide (247 mg. 0.51 mmol), Pd(PPh.sub.3).sub.4 (29 mg, 0.025 mmol), Cul (7.1 mg, 0.0375 mmol), TBAF (1.0 M in THF, 0.62 mL, 0.62 mmol), and diisopropylethylamine (0.14 mL, 0.78 mmol) in 3.0 mL of DMF was stirred at ambient temperature overnight under an atmosphere of N.sub.2. The reaction was quenched with H.sub.2O. EtOAc and more water were added for extraction. The combined organic layers were dried (Na.sub.2SO.sub.4), filtered, concentrated, and the resulting residue was purified by silica gel flash chromatography (eluted with 10% MeOH in CH.sub.2Cl.sub.2) to give the desired product as a brownish solid (216 mg, 73%).

[0190] 3-[(2-amino-1,3-thiazol-5-yl)ethynyl]-4-methyl-N-[3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl]benzamide: The solution of tert-butyl {5-[(2-methyl-5-{[3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl]carbamoyl}phenyl)ethynyl]-1,3-thiazol-2-yl}carbamate (216 mg) in TFA/CH.sub.2Cl.sub.2 (10 mL, 1:1) was stirred at room temperature for 1 hr. The volatile components were removed on rotavap and the residue was partitioned between EtOAc and aq. NaHCO.sub.3. The combined organic layer was dried (Na.sub.2SO.sub.4), filtered, concentrated, and the resulting residue was purified by silica gel flash chromatography (eluted with 10% MeOH in CH.sub.2Cl.sub.2) to give the desired product as a brownish solid (170 mg, 95%): 482 m/z (M+H).

Example 6

3-[(2-(acetylamino)-1,3-thiazol-5-yl]ethynyl)-4-methyl-N-[3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl]benzamide

[0191] ##STR00048##

A solution of 3-[(2-amino-1,3-thiazol-5-yl)ethynyl]-4-methyl-N-[3-(4-methyl-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl]benzamide (100 mg) in Ac.sub.2O (10 mL) was heated at 130° C. for 2 hr. Upon cooling to room temperature the volatile components were removed under vacuum and the residue was partitioned between EtOAc and aq. NaHCO.sub.3. The combined organic layers were dried (Na.sub.2SO.sub.4), filtered, concentrated, and the resulting residue was purified by silica gel flash chromatography (eluted with 10% MeOH in CH.sub.2Cl.sub.2) to give the desired product as a brownish solid (105 mg, 96%): 524 m/z (M+H).

Example 7

Potential Synthesis of 4-[(5-([3-(2-[(dimethylamino)methyl]-1H-imidazol-1yl)-5-trifluoromethyl)phenyl]carbamoyl)-2-methylphenyl)ethynyl]-N-methyl-1H-pyrazole- 1-carboxamide

[0192] ##STR00049##

The title compound can be synthesized from 1H-pyrazole-1-N-methylcaboxamide and N-[3-{2-[(dimethylamino)methyl]-1H-imidazol-1-yl}-5-(trifluoromethyl)phenyl]-3- ethynyl-4-methylbenzamide in a manner similar to that described for Example 1.

[0193] 1-(1H-imidazol-2-yl)-N,N-dimethylmethanathine: To a two-necked round-bottomed flask equipped with a reflux condenser and a pressure-equalizing addition funnel, was added 2-imidazolecarboxaldehyde (6 g, 62.5 mmol) in MeOH (60 mL). To this suspension (ambient temperature) was added a solution of dimethylamine (40% aqueous, 60 mL) at a fast dropping rate (20 min). After the addition was complete, solid sodium borohydride (7 g, 186.8 mmol,) was CAUTIOUSLY added portionwise over 45 min. Foaming occurred after each portion, and the internal temperature was allowed to maintain −50° C. without external cooling. The reaction mixture was then heated to 65° C. for 3 h and allowed to cool to ambient temperature for overnight. The reaction contents were concentrated in vacuo and the resultant residue was taken up in EtOAc (2×30 mL) washed with brine and with CHCl.sub.3 (4×100 mL). The EtOAc extract was discarded. The CHCl.sub.3 extract was dried over (NaSO.sub.4), filtered, and concentrated in vacuo to give 3.7 g of the desired product as a waxy solid.

[0194] 3-(2-((Dimethylamino)methyl)-1H-imidazol-1-yl)-5-(trifluoromethyl)aniline: 3-Amino-5-bromobenzotrifluoride (6 g, 25 mmol) and 1-(1H-imidazol-2-yl)-N,N-dimethylmethanamine (3.7 g, 29.8 mmol) were dissolved in anhydrous DMSO (25 mL). To this was added Cul (0.95 g, 7.5 mmol), 8-hydroxy quinoline (0.72 g, 7.5 mmol) and K.sub.2CO.sub.3 (6.9 g, 50 mmol). The mixture was stirred vigorously and degassed with N.sub.2 for 15 minutes. The flask was then equipped with a condenser and heated at 120° C. for 18 h. The resultant heterogeneous mixture was cooled to rt, poured into 14% aq. NH.sub.4OH (100 mL) and extracted with EtOAc (3×300 ml). The combined extracts were dried over NaSO.sub.4 and concentrated in vacuo. The residue was chromatograhed over silica gel eluting with MeOH/DCM (5:95) to furnish 3.5 g of the desired product as a tan colored material: 285 m/z (M+H).

[0195] N-(3-(2((dimethylamino)methyl)-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl)-3-iodo-4-methylbenzamide: 3-iodo-4-methylbenzoyl chloride (2.2 g, 7.88 mmol), dissolved in anhydrous THF (13 mL), was added dropwise to a solution of 3-(2-((dimethylamino)methyl)-1H-imidazol-1-yl)-5-(trifluoromethyl)aniline (1.5 g, 5.5 mmol), DIPEA (2.1 mL, 11.8 mmol) in THF (30 mL) at ˜5° C. The resultant solution was stirred at ambient temperature overnight. The solvent was removed in vacuo and the crude residue was redissolved in CH.sub.2Cl.sub.2 and washed with 1N NaOH. The organic layer was then washed with water, and brine then dried over NaSO.sub.4 before being concentrated in vacuo. The brown colored residue was then triturated in a mixture of hexanes/DCM to precipitate 1.4 g of the desired product as an off-white powder: 529 m/z (M+H).

[0196] Potential synthesis of N-[3-(2-[(dimethylamino)methyl]-1H-imidazol-1yl)-5-(trifluoromethyl)phenyl]-4-methyl-3((trimethylsilyl)ethynyl)benzamide: N-(3-(2-((dimethylamino)methyl)-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl)-3-iodo-4-methylbenzamide (5 mmol), Pd[(PPh.sub.3)].sub.4 (289 mg, 0.25 mmol), Cul (71 mg, 0.375 mmol) are placed in a schlenk flask. The flask is subjected to 3 cycles of vacuum -refilling with N.sub.2. To this mixture is added anhydrous N,N-diisopropylethylamine (1.1 mL, 6 mmol), DMF (5 mL), and trimethylsilylacetylene (0.92 mL, 6.5 mmol). This solution is stirred at rt for 24 h. Water and EtOAc are added to the reaction mixture to facilitate the extraction. The combined organic layers are dried over Na.sub.2SO.sub.4, filtered, and then concentrated on a rotavap and the residue is purified on a silica gel column (eluent: 5% MeOH in CH.sub.2Cl.sub.2, MeOH was pre-saturated with ammonia gas) to give the desired product.

[0197] Potential Synthesis of N-[3-(2-[(dimethylamino)methyl]-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl]-3-ethynyl-4-methylbenzamide: To a solution of N-[3-(2-[(dimethylamino)methyl]-1H-imidazol-1-yl)-5-(trifluoromethyl)phenyl]-4-methyl-3-[(trimethylsilyl)ethynyl]benzamide (4.1 mmol) in THF (15 mL) is added 5 mL of TBAF in THF (1.0M). After stirring at rt for 1 h, the mixture is partitioned between H.sub.2O and EtOAc. The combined organic layers are dried over Na.sub.2SO.sub.4, filtered, and then concentrated on a rotavap and the residue is purified on a silica gel column (eluent: 10% MeOH in CH.sub.2Cl.sub.2, MeOH was pre-saturated with ammonia gas) to give the desired product.

Example 8

Potential Synthesis of N-(3-chloro-4-[(4-methylpiperazin-1-yl)methyl]phenyl)-4-methyl-3-[(1-methyl-1H-imidazol-5-yl)ethynyl]benzamide:

[0198] ##STR00050##

The title compound can be synthesized according to Example 2, from 5-ethynyl-1-methyl-1H-imidazole (prepared from deprotection of 1-methyl-5-trimethylsilanylethynyl-1H-imidazole) and N-(3-chloro-4-((4-methylpiperazin-1-yl)methyl)phenyl)-3-iodo-4-methylbenzamide.

[0199] 1-(Bromomethyl)-2-chloro-4-nitro-benzene: A suspension of 2-chloro-4-nitrotoluene (10.0 g, 58.3 mmol), N-bromosuccinimide (NBS, 10.9 g, 61.2 mmol), and 2,2′-azobis(2-methylpropionitrile) (AIBN, 0.29 g, 1.75 mmol) in 120 mL of CCl.sub.4 was heated at reflux under an atmosphere of N.sub.2 for 12 h. The reaction mixture was cooled to ambient temperature, and the solid was filtered and washed with EtOAc. The combined filtrate was washed with aq. NaHCO.sub.3, dried over Na.sub.2SO.sub.4, filtered, concentrated on rotovap, and further dried under vacuum. .sup.1H NMR indicated the ratio of desired product to unreacted 2-chloro-4-nitrotoluene to be 50:50. This material was used directly in the next step.

[0200] 1-(2-Chloro-4-nitrobenzyl)-4-methylpiperazine: To a solution of crude 1-(bromomethyl)-2-chloro-4-nitro-benzene (29.1 mmol; 50% pure) in 30 mL of DCM was added Et.sub.3N (4.2 mL, 30 mmol) and 1-methylpiperazine (3.4 mL, 30 mmol). After stirring for 3 h at ambient temperature, aq. NaHCO.sub.3 was added and the mixture was extracted with DCM. The combined organic layer was dried over Na.sub.2SO.sub.4, filtered, concentrated, and the resulting residue was purified by silica gel chromatography (eluted with 5% MeOH/DCM) to provide 6.80 g of product as a dark yellow oil.

[0201] 3-Chloro-4-((4-methylpiperazin-1-yl)methyl)aniline: To a solution of 1-(2-chloro-4-nitrobenzyl)-4-methylpiperazine (0.96 g, 3.6 mmol) in MeOH/water (4:1, 50 mL) was added 1.80 g (33.7 mmol) of NH.sub.4Cl and 1.47 g (26.3 mmol) of Fe dust and the mixture heated at reflux under an atmosphere of N.sub.2 for 2 h (HPLC indicated no progress). To this was added 4 mL of glacial acetic acid and the mixture heated at reflux for an additional 2 h. The reaction mixture was cooled to ambient temperature, filtered, and the filtrate concentrated. The residue was partitioned between EtOAc and saturated aq. NaHCO.sub.3, the separated aqueous layer was extracted with EtOAc, and the combined organics washed with brine and dried over Na.sub.2SO.sub.4. Upon concentration, the crude product was purified by silica gel chromatography (eluted with 5-7% MeOH/DCM; silica gel deactivated with 1% triethylamine/DCM) to provide 0.53 g of product.

Alternative Potential Synthesis of N-(3-chloro-4-[(4-methylpiperazin-1-yl)methyl]phenyl)-4-methyl-3-[(1-methyl-1H-imidazol-5-yl)ethynyl]benzamide

[0202] 5-ethynyl-1-methyl-1H-imidazole: To a solution of 1-methyl-5-trimethylsilanylethynyl-1H-imidazole (1.39 mol) in 10× volume of Ethyl acetate and 1.5× volume of Methanol is added two and a half equivalents of potassium carbonate at ambient temperature and the solution stirred for 1 hour. Potassium carbonate is filtered off and the organic stream is washed with water and with saturated sodium chloride solution (two or more times). Aqueous phases can be combined and re-extracted with ethyl acetate. Organic streams can than be combined and concentrated under vacuum to about 0.5 L. Solids can be allowed to precipitate out upon concentration. Slurry is cooled, e.g. to about −5° C., stored overnight, filtered, and washed with about 0.3 L of cold ethyl acetate. The solids can then be dried under vacuum.

[0203] 4-methyl-3-[(1-methyl-1H-imidazol-5-yl)ethynyl]benzoic acid can be prepared in a manner similar to that described above for the Sonogashira reaction. 5-ethynyl-1-methyl-1H-imidazole and 3-iodo-4-methylbenzoic acid can be used as coupling partners. Alternatively, the solvent (DMF) can be replaced by ethyl acetate and the base (Hunig base) can be replaced by triethylamine. The product can be isolated by filtration of the crude reaction mixture. The filter cake is washed sequentially with a solvent such as ethyl acetate and then water, then dried in a vacuum oven. Further purification can be achieved by slurrying the solids in water adjusted to pH 3 with the addition of concentrated HCl. After filtration and water wash, the product can be dried in a vacuum oven.

[0204] N-(3-chloro-4-[(4-methylpiperazin-1-yl)methyl]phenyl)-4-methyl-3-[(1-methyl-1H-imidazol-5-yl)ethynyl]benzamide: 4-methyl-3-[(1-methyl-1H-imidazol-5-yl)ethynyl]benzoic acid (18 mmol) is dissolved in methylene chloride (100 mL). To this solution is added 3 equivalents of 4-methylmorpholine (NMM) followed by 1.05 equivalents of oxalyl chloride. After stirring at ambient temperature for 30 minutes, 0.8 equivalents of 3-Chloro-4-((4-methylpiperazin-1-yl)methyl)aniline (prepared as above) is added along with 5 mole % of DMAP. After initially stirring at ambient temperature, the mixture is brought to reflux and stirred overnight. After 16 h an additional 0.2 equivalents of the aniline is added, bringing the total charge to 1 equivalent. The mixture can then be stirred for an additional 2 h, quenched with water, and the layers separated. The aqueous layer can be extracted with methylene chloride (2×50 mL) and the combined extracts can be washed with water. The combined methylene chloride layers can then be evaporated and the residue dissolved in 100 mL of ethyl acetate (20 mL). After standing for 1 h, the product is allowed to crystallize. The mixture is cooled, e.g. to 0° C., filtered, and the solid product is washed with cold ethyl acetate.

Example 9

Potential Synthesis of 3-[(2-amino-1,3-thiazol-5-yl)ethynyl]-N-(3-cyclopropyl-4-[(4-methylpiperazin-1-yl)methyl]phenyl)-4-methylbenzamide

[0205] ##STR00051##

The title compound can be synthesized from tert-butyl (5-ethynyl-1,3-thiazol-2-yl)carbamate and N-(3-cyclopropyl-4-((4-methylpiperazin-1-yl)methyl)phenyl)-3-iodo-4-methylbenzamide in a manner similar to that described for Example 2 (nitro reduction performed in a manner similar to that described for Example 8; 0.25M in MeOH/10% AcOH). t-BOC deprotection can be performed after coupling as described in Example 5.

[0206] Potential Synthesis of tert-butyl (5-ethynyl-1,3-thiazol-2-yl)carbamate: To a solution of tert-butyl (5-[(trimethylsilyl)ethynyl]-1,3-thiazol-2-yl)carbamate (prepared as in Example 5, 1.39 moil) in 10× volume of Ethyl acetate and 1.5× volume of Methanol is added two and a half equivalents of potassium carbonate at ambient temperature and the solution stirred for 1 hour. Potassium carbonate is filtered off and the organic stream is washed with water and with saturated sodium chloride solution (two or more times). Aqueous phases can be combined and re-extracted with ethyl acetate. Organic streams can then be combined and concentrated under vacuum to about 0.5 L. Solids can be allowed to precipitate out upon concentration. Slurry is cooled, e.g. to about −5° C., stored overnight, filtered, and washed with about 0.3 L of cold ethyl acetate. The solids can then be dried under vacuum.

[0207] 1-(2-Cyclopropyl-4-nitrobenzyl)-4-methylpiperazine: A mixture of 1-(2-bromo-4-nitrobenzyl)-4-methylpiperazine (0.94 g, 3.0 mmol), 0.77 g (9.0 mmol) of cyclopropylboronic acid, 0.057 g (0.30 mmol) of Pd(OAc).sub.2, 2.87 g (13.5 mmol) of K.sub.3PO.sub.4, and 0.168 g (0.60 mmol) of tricyclohexylphosphine in 18 mL of toluene/water (5:1) was heated at reflux under an atmosphere of N.sub.2 for 19 h. The reaction mixture was concentrated and the crude product was purified by silica gel chromatography (eluted with 5% MeOH/DCM; MeOH was pre-saturated with ammonia gas) to provide 0.80 g of product.

Example 10

Potential Synthesis of 1-methyl-5-((2-methyl-5[((4-[(4-methylpiperazin-1-yl)methyl]-3-(trifluoromethyl)phenyl)carbonyl)amino]phenyl)ethynyl)-1H-imidazole-2- carboxamide

[0208] ##STR00052##

The title compound can be synthesized from 5-ethynyl-1-methyl-1H-imidazole-2-carboxylic acid amide and N-(3-iodo-4-methylphenyl)-4-((4-methylpiperazin-1-yl)methyl)-3-(trifluoromethyl)benzamide in a manner similar to that described for Example 2. 5-ethynyl-1-methyl-1H-imidazole-2-carboxylic acid amide is prepared as in Example 2.

[0209] N-(3-iodo-4-methylphenyl)-4-((4-methylpiperazin-1-yl)methyl)-3-(trifluoromethyl)benzamide: To a flask containing 1.0 g (2.67 mmol) of 4-[(4-methyl-1-piperazinyl)methyl]-3-(trifluoromethyl)-benzoic acid (CAS #859027-02-4; prepared according to Asaki, T. et al. Bioorg. Med. Chem. Lett. (2006), 16, 1421-1425), 0.62 .sub.g (2.67 mmol) of 3-iodo-4-methylaniline, 0.77 g (4.0 mmol) of N-(3-dimethylaminopropyl)-N′-ethylcarbodiimide hydrochloride (EDAC), and 0.43 g (3.2 mmol) of N-hydroxybenzotriazole monohydrate (HOBt.H.sub.2O) was added 5 mL of DCM and 5 mL of triethylamine. The solution was stirred at ambient temperature under an atmosphere of N.sub.2 for 3 days, concentrated, and the crude product purified by silica gel chromatography (eluted with 100% EtOAc then 10% MeOH/EtOAc), to provide 0.69 g of product as a white solid.

Example 11

Potential Synthesis of 5-[(5-[(4-[((3R)-3(dimethylamino)pyrrolidin-1-yl]methyl)-3-(trifluoromethyl)phenyl]carbamoyl)-2-methylphenyl)ethynyl]-1-methyl-1H-imidazole-2-carboxamide

[0210] ##STR00053##

The title compound can be synthesized from 5-ethynyl-1-methyl-1H-imidazole-2-carboxylic acid amide and (R)-N-(4-((3-(Dimethylamino)pyrrolidin-1-yl)methyl)-3-(trifluoromethyl)phenyl)-3-iodo-4-methylbenzamide in a manner similar to that described for Example 2. 5-ethynyl-1-methyl-1H-imidazole-2-carboxylic acid amide is prepared as in Example 2.

[0211] 1-(Bromomethyl)-4-nitro-2-(trifluoromethyl)benzene: A suspension of 2-methyl-5-nitrobenzotrifluoride (3.90 g, 19 mmol), N-bromosuccinimide (NBS, 3.56 g, 20 mmol), and 2,2′-azobis(2-methylpropionitrile) (AIBN, 0.094 g, 0.6 mmol) in 40 mL of CCl.sub.4 was heated at reflux under N.sub.2 for 16 h. HPLC indicated ca. 50% conversion. Additional NBS (10 mmol) and AIBN (0.6 mmol) were added and the mixture was heated at reflux for another 14 h. HPLC indicated ca. 80% conversion. The reaction mixture was cooled to ambient temperature, and the solid was filtered and washed with EtOAc. The combined filtrate was washed with aq. NaHCO.sub.3, dried over Na.sub.2SO.sub.4, filtered, concentrated on rotovap, and further dried under vacuum. .sup.1H NMR indicated the ratio of desired product to unreacted 2-methyl-5-nitrobenzotrifluoride to be 75:25. This material was used directly in the next step.

[0212] (R)-N,N-Dimethyl-1-(4-nitro-2-(trifluoromethyl)benzyl)pyrrolidin-3-amine: To a solution of crude 1-(bromomethyl)-4-nitro-2-(trifluoromethyl)benzene (17.5 mmol, 75% pure) in 40 mL of DCM was added Et.sub.3N (2.69 mL, 19.3 mmol) and (R)-(+)-3-(dimethylamino)pyrrolidine (2.0 g, 17.5 mmol). After stirring overnight at ambient temperature under an atmosphere of N.sub.2, the reaction solution was concentrated, added aq. NaHCO.sub.3 (100 mL), and the resulting mixture extracted with DCM (4×50 mL). The combined organic layer was dried over Na.sub.2SO.sub.4, filtered, concentrated, and the resulting residue was purified by silica gel chromatography (eluted with 0-10% MeOH/DCM) to provide 3.35 g of product as a yellow oil.

[0213] (R)-1-(4-Amino-2-(trifluoromethyl)benzyl)-N,N-dimethylpyrrolidin-3-amine: To a solution of (R)-N,N-dimethyl-1-(4-nitro-2-(trifluoromethyl)benzyl)pyrrolidin-3-amine (1.20 g, 3.79 mmol) in 20 mL of wet EtOH was added 0.26 g of Pd/C (10% Pd on C) and the mixture shaken in a Parr apparatus (pressure reaction vessel purged thoroughly with H.sub.2 and pressure regulated at 45 psi throughout) for 2-3 h. The reaction mixture was filtered through a small pad of celite, washed with EtOAc, and the combined organics concentrated to provide a quantitative yield of a light yellow oil. This material was used directly in the next step.

[0214] (R)-N-(4-((3-(Dimethylamino)pyrrolidin-1-yl)methyl)-3-(trifluoromethyl)phenyl)-3-iodo-4-methylbenzamide: To a cooled (0° C.) solution of (R)-1-(4-amino-2-(trifluoromethyl)benzyl)-N,N-dimethylpyrrolidin-3-amine (3.79 mmol) in 14 mL DCM, under an atmosphere of N.sub.2, was added 3-iodo-4-methylbenzoyl chloride (1.17 g, 4.17 mmol; CAS #52107-98-9, prepared from the reaction of 3-iodo-4-methylbenzoic acid and SOCl.sub.2) followed by dropwise addition of N,N-diisopropylethylamine (2.64 mL, 15.2 mmol). After stirring to ambient temperature over 1.5 h, the reaction mixture was concentrated and the crude product was purified by silica gel chromatography (eluted with 0-8% MeOH/DCM; MeOH was pre-saturated with ammonia gas), to provide 0.71 g of product as a thick yellow oil.

[0215] Potential Synthesis of 5-[(5-((4-(((3R)-3-(dimethylamino)pyrrolidin-1-yl)methyl)-3-(trifluoromethyl)phenyl]carbamoyl)-2-methylphenyl)ethynyl)-1-methyl-1H- imidazole-2-carboxamide: A mixture of 5-ethynylpyrimidne (0.34 mmol), 0.150 g (0.28 mmol) of (R)-N-(4-((3-(dimethylamino)pyrrolidin-1-yl)methyl)-3-(trifluoromethyl)phenyl)-3-iodo-4- methylbenzamide, 0.016 g (0.014 mmol) of Pd(PPh.sub.3).sub.4, 0.004 g (0.021 mmol) of Cul, and 0.09 mL (0.51 mmol) of N,N-diisopropylethylamine in 3.5 mL of DMF is stirred at ambient temperature, under an atmosphere of N.sub.2, for 3 days (reaction pushed to completion with additional equivalents of reagents and heating to 80° C.). The reaction mixture is concentrated and the crude product is purified by silica gel chromatography (eluted with 0-10% MeOH/DCM; MeOH was pre-saturated with ammonia gas) to provide the title compound.

Example 12: Biological Evaluation of Compounds

[0216] Compounds of this invention are evaluated in a variety of assays to determine their biological activities. For example, the compounds of the invention can be tested for their ability to inhibit various protein kinases of interest. Some of the compounds tested displayed potent nanomolar activity against the following kinases: Abl, Abl T315I, Src and FGFR. Furthermore, several of these compounds were screened for antiproliferative activity in BaF3 cells transfected with either wild-type Bcr-Abl or the Bcr-Abl T315I mutant and demonstrated activity in the range of 1-100 nM.

[0217] The compounds can also be evaluated for their cytotoxic or growth inhibitory effects on tumor cells of interest, e.g., as described in more detail below and as shown above for some representative compounds. See e.g., WO 03/000188, pages 115-136, the full contents of which are incorporated herein by reference.

[0218] Some representative compounds are depicted below:

##STR00054## ##STR00055## ##STR00056## ##STR00057##

Kinase Inhibition

[0219] More specifically, the compounds described herein are screened for kinase inhibition activity as follows. Kinases suitable for use in the following protocol include, but are not limited to: Abl, Lck, Lyn, Src, Fyn, Syk, Zap-70, Itk, Tec, Btk, EGFR, ErbB2, Kdr, Flt1, Flt-3, Tek, c-Met, InsR, and AKT.

[0220] Kinases are expressed as either kinase domains or full length constructs fused to glutathione S-transferase (GST) or polyHistidine tagged fusion proteins in either E. coli or Baculovirus-High Five expression systems. They are purified to near homogeneity by affinity chromatography as previously described (Lehr et al., 1996; Gish et al., 1995). In some instances, kinases are co-expressed or mixed with purified or partially purified regulatory polypeptides prior to measurement of activity.

[0221] Kinase activity and inhibition can be measured by established protocols (see e.g., Braunwalder et al., 1996). In such cases, the transfer of .sup.33PO.sub.4 from ATP to the synthetic substrates poly(Glu, Tyr) 4:1 or poly(Arg, Ser) 3:1 attached to the bioactive surface of microtiter plates is taken as a measure of enzyme activity. After an incubation period, the amount of phosphate transferred is measured by first washing the plate with 0.5% phosphoric acid, adding liquid scintillant, and then counting in a liquid scintillation detector. The IC50 is determined by the concentration of compound that causes a 50% reduction in the amount of .sup.33P incorporated onto the substrate bound to the plate.

[0222] In one method, the activated kinase is incubated with a biotinylated substrate peptide (containing tyrosine) with or without the presence of a compound of the invention. After the kinase assay incubation period, excess kinase inhibitor is added to kill the kinase reaction along with Europium-labeled anti-phosphotyrosine antibody (Eu-Ab) and Aliophycocyanin-Streptavidin (SA-APC). The biotinylated substrate peptide (with or without phosphorylated Tyrosine) in solution binds to the SA-APC via Biotin-Avidin binding. The Eu-Ab binds only to substrate with phosphorylated tyrosine. When the solution is excited at 615 nm, there is an energy transfer from the Europium to the APC when they are in close proximity (i.e. attached to the same molecule of biotinylated and phosphorylated substrate peptide). The APC then fluoresces at a wavelength of 665 nm. Excitation and emission take place in a Wallac Victor.sup.2 V plate reader where the plate is read fluorometrically and absorbances at 615 and 665 nm are recorded. These data are then processed by an Excel plate processor which calculates IC50s of test compounds by converting the fluorescence into amounts of phosphorylated substrate made and determining the concentration of test compound that would be required to inhibit the development of phosphorylated substrate by 50% (IC50).

[0223] Other methods relying upon the transfer of phosphate to peptide or polypeptide substrate containing tyrosine, serine, threonine or histidine, alone, in combination with each other, or in combination with other amino acids, in solution or immobilized (i.e., solid phase) are also useful.

[0224] For example, transfer of phosphate to a peptide or polypeptide can also be detected using scintillation proximity, Fluorescence Polarization or homogeneous time-resolved fluorescence. Alternatively, kinase activity can be measured using antibody-based methods in which an antibody or polypeptide is used as a reagent to detect phosphorylated target polypeptide.

[0225] For additional background information on such assay methodologies, see e.g., Braunwalder et al., 1996, Anal. Biochem. 234(I):23; Cleaveland et al., 1990, Anal Biochem. 190(2):249 Gish et al. (1995). Protein Eng. 8(6):609 Kolb et al. (1998). Drug Discov. Toda V. 3:333 Lehr et al. (1996). Gene 169(2):27527-87 Seethaia et al. (1998). Anal Biochem. 255(2):257 Wu et al. (2000).

[0226] IC50 values in the low nanomolar range have been observed for compounds of this invention against various kinases, including Src, Abl and kdr.

Cell-Based Assays

[0227] Certain compounds of this invention have also been demonstrated cytotoxic or growth inhibitory effects on tumor and other cancer cell lines and thus may be useful in the treatment of cancer and other cell proliferative diseases. Compounds are assayed for anti-tumor activity using in vivo and in vitro assays which are well known to those skilled in the art. Generally, initial screens of compounds to identify candidate anti-cancer drugs are performed in cellular assays. Compounds identified as having anti-proliferative activity in such cell-based assays can then be subsequently assayed in whole organisms for anti-tumor activity and toxicity. Generally speaking, cell-based screens can be performed more rapidly and cost-effectively relative to assays that use whole organisms. For purposes of this invention, the terms “anti-tumor” and “anti-cancer” activity are used interchangeably.

[0228] Cell-based methods for measuring antiproliferative activity are well known and can be used for comparative characterization of compounds of this invention. In general, cell proliferation and cell viability assays are designed to provide a detectable signal when cells are metabolically active. Compounds may be tested for antiproliferative activity by measuring any observed decrease in metabolic activity of the cells after exposure of the cells to compound. Commonly used methods include, for example, measurement of membrane integrity (as a measure of cell viability) (e.g. using trypan blue exclusion) or measurement of DNA synthesis (e.g. by measuring incorporation of BrdU or 3H-thymidine).

[0229] Some methods for assaying cell proliferation use a reagent that is converted into a detectable compound during cell proliferation. Particularly preferred compounds are tetrazolium salts and include without limitation MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; Sigma-Aldrich, St. Louis, Mo.), MTS (3-(4,5-dimethylthiazol-2yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium), XTT (2,3-bis(2-Methoxy-4-nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide), INT, NBT, and NTV (Bernas et al. Biochim Biophys Acta 1451(1):73-81, 1999). Preferred assays utilizing tetrazolium salts detect cell proliferation by detecting the product of the enzymatic conversion of the tetrazolium salts into blue formazan derivatives, which are readily detected by spectroscopic methods (Mosman. J. Immunol. Methods. 65:55-63, 1983).

[0230] Generally, preferred methods for assaying cell proliferation involve incubating cells in a desired growth medium with and without the compounds to be tested. Growth conditions for various prokaryotic and eukaryotic cells are well-known to those of ordinary skill in the art (Ausubel et al. Current Protocols in Molecular Biology. Wiley and Sons. 1999; Bonifacino et al. Current Protocols in Cell Biology. Wiley and Sons. 1999 both incorporated herein by reference). To detect cell proliferation, the tetrazolium salts are added to the incubated cultured cells to allow enzymatic conversion to the detectable product by active cells. Cells are processed, and the optical density of the cells is determined to measure the amount of formazan derivatives. Furthermore, commercially available kits, including reagents and protocols, are available for examples, from Promega Corporation (Madison, Wis.), Sigma-Aldrich (St. Louis, Mo.), and Trevigen (Gaithersburg, Md.).

[0231] More specifically, the cell proliferation assay we currently perform is using CellTiter 96 AQueous One Solution Cell Proliferation assay kit (Promaga, Cat #G3581). This assay is a colorimetric method for determining the number of alive cells in proliferation or cytotoxicity assays. The assay utilizing terazolium salts detect cell proliferation by detecting the product of the enzymatic conversion of the tetrazolium salts into blue formazan derivatives, which can be measured by the absorbance at 490 nm in a plate reader, Wallac Victor.sup.2V (PerkinElmer).

[0232] An example of cell-based assay is shown as below. The cell lines used in the assay are Ba/F3, a murine pro-B cell line, which have been stably transfected with full-length wild-type Bcr-Abl or Bcr-Abl with various kinase domain point mutations (including T351I, Y253F, E255K, H396P, M351T etc) constructs. Parental Ba/F3 cell line is used as control. These cell lines were obtained from Brian J. Druker (Howard Hughes Medical Institute, Oregon Health and Science University, Portland, Oreg., USA). Ba/F3 cell expressing Bcr-Abl or Bcr-Abl mutants were maintained in PRMI 1640 growth medium with 200 μM L-gultamine, 10% FCS, penicillin (200 U/ml), and streptomycin (200 μg/ml). Parental Ba/F3 cells were culture in the same medium supplemented with 10 ng/ml IL-3.

[0233] Parental Ba/F3 cells (supplemented with IL-3) or Ba/F3 cells expressing WT or mutant Bcr-Abl are plated in duplicate at 1×10.sup.4 cells/well in 96-well plates with the compounds in different concentrations in the media. The compounds are first dissolved and diluted in DMSO by preparation of 4-fold dilution; next equal volumes of compounds with DMSO are transferred to medium and then transferred to cell plates. The final compound concentrations start from 10 μM to 6 nM. DMSO at same percentage is used as control. After compound was incubated with cells for 3 days, the numbers of active cells are measured using CellTiter 96 AQueous One Solution Cell Proliferation assay kit following the kit instruction. Basically, the tetrazolium salts are added to the incubated cultured cells to allow enzymatic conversion to the detectable product by active cells. Cells are processed, and the optical density of the cells is determined to measure the amount of formazan derivatives. Mean+/−SD are generated from duplicated wells and reported as the percentage absorbance of control. IC50s are calculated in best-fit curves using Micorsoft Excel-fit software.

[0234] In addition, a wide variety of cell types may be used to screen compounds for antiproliferative activity, including the following cell lines, among others: COLO 205 (colon cancer), DLD-1 (colon cancer), HCT-15 (colon cancer), HT29 (colon cancer), HEP G2 (Hepatoma), K-562 (Leukemia), A549 (Lung), NCI-H249 (Lung), MCF7 (Mammary), MDA-MB-231 (Mammary), SAOS-2 (Osteosarcoma), OVCAR-3 (Ovarian), PANC-1 (Pancreas), DU-145 (Prostate), PC-3 (Prostate), ACHN (Renal), CAKI-1 (Renal), MG-63 (Sarcoma).

[0235] While the cell line is preferably mammalian, lower order eukaryotic cells such as yeast may also be used to screen compounds. Preferred mammalian cell lines are derived from humans, rats, mice, rabbits, monkeys, hamsters, and guinea pigs since cells lines from these organisms are well-studied and characterized. However, others may be used as well.

[0236] Suitable mammalian cell lines are often derived from tumors. For example, the following tumor cell-types may be sources of cells for culturing cells: melanoma, myeloid leukemia, carcinomas of the lung, breast, ovaries, colon, kidney, prostate, pancreas and testes), cardiomyocytes, endothelial cells, epithelial cells, lymphocytes (T-cell and B cell), mast cells, eosinophils, vascular intimal cells, hepatocytes, leukocytes including mononuclear leukocytes, stem cells such as haemopoetic, neural, skin, lung, kidney, liver and myocyte stem cells (for use in screening for differentiation and de-differentiation factors), osteoclasts, chondrocytes and other connective tissue cells, keratinocytes, melanocytes, liver cells, kidney cells, and adipocytes. Non-limiting examples of mammalian cells lines that have been widely used by researchers include HeLa, NIH/3T3, HT1080, CHO, COS-1, 293T, WI-38 and CV1/EBNA-1.

[0237] Other cellular assays may be used which rely upon a reporter gene to detect metabolically active cells. Non-limiting examples of reporter gene expression systems include green fluorescent protein (GFP), and luciferase. As an example of the use of GFP to screen for potential antitumor drugs, Sandman et al. (Chem Biol. 6:541-51; incorporated herein by reference) used HeLa cells containing an inducible variant of GFP to detect compounds that inhibited expression of the GFP, and thus inhibited cell proliferation.

[0238] Compounds identified by such cellular assays as having anti-cell proliferation activity are then tested for anti-tumor activity in whole organisms. Preferably, the organisms are mammalian. Well-characterized mammalians systems for studying cancer include rodents such as rats and mice. Typically, a tumor of interest is transplanted into a mouse having a reduced ability to mount an immune response to the tumor to reduce the likelihood of rejection. Such mice include for example, nude mice (athymic) and SCID (severe combined immunodeficiency) mice. Other transgenic mice such as oncogene containing mice may be used in the present assays (see for example U.S. Pat. No. 4,736,866 and U.S. Pat. No. 5,175,383). For a review and discussion on the use of rodent models for antitumor drug testing see Kerbel (Cancer Metastasis Rev. 17:301-304, 1998-99).

[0239] In general, the tumors of interest are implanted in a test organism preferably subcutaneously. The organism containing the tumor is treated with doses of candidate anti-tumor compounds. The size of the tumor is periodically measured to determine the effects of the test compound on the tumor. Some tumor types are implanted at sites other than subcutaneous sites (e.g. intraperitoneal sites) and survival is measured as the endpoint. Parameters to be assayed with routine screening include different tumor models, various tumor and drug routes, and dose amounts and schedule. For a review of the use of mice in detecting antitumor compounds see Corbett et al. (Invest New Drugs. 15:207-218, 1997; incorporated herein by reference).

Example 8: Pharmaceutical Compositions

[0240] Representative pharmaceutical dosage forms of the compounds of this invention (the active ingredient being referred to as “Compound”), are provided for therapeutic or prophylactic use in humans:

TABLE-US-00001 (a) Tablet I mg/tablet Compound 100 Lactose Ph. Eur 182.75 Croscarmellose sodium 12.0 Maize starch paste (5% w/v paste) 2.25 Magnesium stearate 3.0

TABLE-US-00002 (b) Tablet II mg/tablet Compound 50 Lactose Ph. Eur 223.75 Croscarmellose sodium 6.0 Maize starch 15.0 Polyvinylpyffolidone (5% w/v paste) 2.25 Magnesium stearate 3.0

TABLE-US-00003 (c) Tablet III mg/tablet Compound 1.0 Lactose Ph. Eur 93.25 Croscarmellose sodium 4.0 Maize starch paste (5% w/v paste) 0.75 Magnesium stearate 1.0-76

TABLE-US-00004 (d) Capsule mg/capsule Compound 10 Lactose Ph. Eur 488.5 Magnesium 1.5

TABLE-US-00005 (e) Injection I (50 mg/ml) Compound 5.0% w/v 1M Sodium hydroxide solution 15.0% w/v  0.1M Hydrochloric acid (to adjust pH to 7.6) Polyethylene glycol 400 4.5% w/v Water for injection to 100%

TABLE-US-00006 (f) Injection II (10 mg/ml) Compound 1.0% W/v Sodium phosphate BP 3.6% w/v O.1M Sodium hydroxide solution 15.0% v/v  Water for injection to 100%

TABLE-US-00007 (g) Injection III (1 mg/ml, buffered to pH 6) Compound 0.1% w/v Sodium phosphate BP 2.26% w/v  Citric acid 0.38% w/v  Polyethylene glycol 400 3.5% w/v Water for injection to 100%

TABLE-US-00008 (h) Aerosol 1 mg/ml Compound 10.0 Sorbitan trioleate 13.5 Trichlorofluoromethane 910.0 Dichlorodifluoromethane 490.0

TABLE-US-00009 (i) Aerosol II mg/ml Compound 0.2 Sorbitan trioleate 0.27 Trichlorofluoromethane 70.0 Dichlorodifluoromethane 280.0 Dichlorotetrafluoroethane 1094.0

TABLE-US-00010 (j) Aerosol III mg/ml Compound 2.5 Sorbitan trioleate 3.38 Trichlorofluoromethane 67.5 Dichlorodifluoromethane 1086.0 Dichlorotetrafluoroethane 191.6

TABLE-US-00011 (k) Aerosol IV mg/ml Compound 2.5 Soya lecithin 2.7 Trichlorofluoromethane 67.5 Dichlorodifluoromethane 1086.0 Dichlorotetrafluoroethane 191.6

TABLE-US-00012 (l) Ointment ml Compound 40 mg Ethanol 300 μl Water 300 μl 1-Dodecylazacycloheptanone 50 μl Propylene glycol to 1 ml
Note: These formulations may be prepared using conventional procedures well known in the pharmaceutical art. The tablets (a)-(c) may be enteric coated by conventional means, if desired to provide a coating of cellulose acetate phthalate, for example. The aerosol formulations (h)-(k) may be used in conjunction with standard, metered dose aerosol dispensers, and the suspending agents sorbitan trioleate and soya lecithin may be replaced by an alternative suspending agent such as sorbitan monooleate, sorbitan sesquioleate, polysorbate 80, polyglycerol oleate or oleic acid.