Derivatives of indole for the treatment of cancer, viral infections and lung diseases

Abstract

The present invention relates to a new class of indole derivatives, having a particular MKlp2 inhibition profile and useful as a therapeutic agent, in particular for the treatment of cancer, viral infections and lung diseases.

Claims

1. A method for reducing tumor volume or ameliorating cancer in a subject comprising administering to said subject having a cancer selected from leukemia, acute myelogenous leukemia, lymphoma, breast cancer, pancreatic cancer, lung cancer and colon cancer an effective amount of a pharmaceutical composition comprising (E)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile, (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile, or pharmaceutically acceptable salts thereof.

2. The method according to claim 1, wherein said compound is (E)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile or a pharmaceutically acceptable salt thereof.

3. The method according to claim 1, wherein said compound is (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile or a pharmaceutically acceptable salt thereof.

4. The method according to claim 1, wherein said pharmaceutical composition further comprises an additional antitumoral drug.

5. The method according to claim 4, wherein said additional antitumoral drug is selected from the group consisting of an anti-mitotic agent, an inhibitor of topoisomerases I or II, a DNA alkylating agent, an anti-metabolic agent, a kinase inhibitor and/or a therapeutic antibody designed to mediate cytotoxicity against cells of said cancer or to modulate a key biological function of said cells.

6. The method according to claim 1, wherein said compound is administered in combination with radiotherapy, hyperthermia and/or other antitumoral therapies, or before, simultaneously and/or after surgery.

7. The method according to claim 1, wherein the cancer is lung cancer.

8. The method according to claim 1, wherein the cancer is pancreatic cancer.

9. The method according to claim 1, wherein the cancer is leukemia.

10. The method according to claim 1, wherein the cancer is acute myelogenous leukemia.

11. The method according to claim 1, wherein the cancer is lymphoma.

12. The method according to claim 1, wherein the cancer is breast cancer.

13. The method according to claim 1, wherein the cancer is colon cancer.

14. The method according to claim 1, said method reducing tumor volume in said subject.

15. The method according to claim 1, said method ameliorating cancer in said subject.

Description

FIGURES

(1) FIG. 1: Stability of compound 63 in mouse and human plasma.

(2) FIG. 2: Amount of formed compound 38 from compound 63 in mouse and human plasma.

(3) FIG. 3: Evaluation of anti-tumor activity of compound 38 in nude mice bearing subcutaneous human colon HCT-116 xenografts.

(4) FIG. 4: Evaluation of anti-tumor activity of compound 38 in nude mice bearing subcutaneous non small cell lung carcinoma NCl-H460 xenografts.

EXAMPLES

(5) The following examples illustrate in detail the preparation of compounds of formula (I) according to the invention. The structures of the products obtained have been confirmed by NMR spectra.

(6) Starting compounds and reactants, unless otherwise indicated, are commercially available or described in literature, or can be prepared according to methods described in literature or known to one skilled in the art.

Example 1: Preparation of Starting Indoles and Aldehydes

A) Syntheses of Starting Indoles

Tert-butyl-5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate

(7) ##STR00009##

(8) In a 250 mL pear flask, 2-(5-bromo-1H-indol-3-yl)acetonitrile (1.76 g, 7.49 mmol) was dissolved in 70 mL of acetonitrile to give a colorless solution. Di-tert-butyl-dicarbonate (1.922 mL, 8.98 mmol) and DMAP (0.091 g, 0.749 mmol) were added to the solution and the reaction mixture was stirred at RT for 1 h.

(9) TLC: 100% Dichloromethane showed no more starting material.

(10) Then the reaction mixture was poured into 50 mL of water, extracted with 250 mL of ethyl acetate and the combined organic layer was successively washed with 150 mL of Brine, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo to give a yellow oil which crystallized upon standing to give a yellow solid, m=2.59 g (Yield: 99%).

(11) APCI-MS: (MH)=234

(12) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 8.00 (d, J=8.8 Hz, 1H), 7.91 (d, J=1.8 Hz, 1H), 7.73 (s, 1H), 7.53 (dd, J=8.8, 2.0 Hz, 1H), 4.12 (d, J=0.9 Hz, 2H), 1.61 (s, 9H).

(13) Tert-butyl-5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate is obtained according to the same procedure as for Tert-butyl-5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate.

Tert-butyl 6-X-3-(cyanomethyl)-1H-indole-1-carboxylate

Synthesis of 2-(6-X-1H-indol-3-yl)acetonitrile (X=bromo, fluoro or chloro)

(14) A mixture of 6-X-1-H-indole-3-carbaldehyde (1 eq), formamide (9 mL/mmol), MeOH (9 mL/mmol) and NaBH.sub.4 (3 eq) were stirred 1 h at room temperature. KCN (10 eq) was then added and the resulting mixture was stirred 5 h at 60 C. The reaction was quenched with aqueous NaCl and extracted with CHCl.sub.3, dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was purified by silicagel chromatography (CH.sub.2Cl.sub.2/MeOH, 100:0 to 90:10) to give the title compound.

2-(6-bromo-1H-indol-3-yl)acetonitrile

(15) ##STR00010##

(16) 6-bromo-1H-indole-3-carbaldehyde (200.0 mg), formamide (8 mL), NaBH.sub.4 (101.0 mg), MeOH (8 ml). KCN (580.0 mg). Aspect of the pure product: white solid. (Yield: 67%).

(17) .sup.1H NMR (CDCl.sub.3, 300 MHz) ppm: 8.21 (s, 1H), 7.59 (s, 1H), 7.48 (d, 1H), 7.31 (d, 1H), 7.24 (s, 1H), 3.84 (s, 1H).

2-(6-fluoro-1H-indol-3-yl)acetonitrile

(18) ##STR00011##

(19) 6-fluoro-1H-indole-3-carbaldehyde (200.0 mg), formamide (10 mL), NaBH.sub.4 (138.0 mg), MeOH (10 ml). KCN (791.0 mg). Aspect of the pure product: white solid. (Yield: 75%).

(20) .sup.1H NMR (CDCl.sub.3, 300 MHz) ppm: 8.48 (s, 1H), 7.53-7.50 (m, 1H), 7.14 (s, 1H), 7.09 (d, 1H), 7.0-6.96 (m, 1H), 3.80 (s, 2H).

2-(6-chloro-1H-indol-3-yl)acetonitrile

(21) ##STR00012##

(22) 6-chloro-1H-indole-3-carbaldehyde (200.0 mg), formamide (10 mL), NaBH.sub.4 (126.0 mg), MeOH (10 ml). KCN (722.0 mg). Aspect of the pure product: white solid. (Yield: 81%).

(23) .sup.1H NMR (CDCl.sub.3, 300 MHz) ppm: 8.34 (s, 1H), 7.50 (d, 1H), 7.38 (s, 1H), 7.24-7.16 (m, 1H), 7.14 (d, 1H), 3.82 (s, 2H).

Synthesis of Tert-butyl 6-X-3-(cyanomethyl)-1H-indole-1-carboxylate

(24) To a solution of 2-(6-X-1H-indol-3-yl)acetonitrile in CH.sub.2Cl.sub.2, was added Boc.sub.2O (eq) and DMAP (eq). The resulting mixture was stirred 12 h at room temperature, then diluted with CH.sub.2Cl.sub.2, washed with water and concentrated to give the title compound.

Tert-butyl 6-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate

(25) ##STR00013##

(26) 2-(6-bromo-1H-indol-3-yl)acetonitrile (138.0 mg), DMAP (3.0 mg), Boc.sub.2O (152.0 mg). CH.sub.2Cl.sub.2 (2.8 mL). 12 h at room temperature. Aspect of the pure product: white solid. (Yield: 90%).

(27) .sup.1H NMR (CDCl.sub.3, 300 MHz) ppm: 8.41 (s, 1H), 7.62 (s, 1H), 7.47-7.36 (m, 2H), 3.78 (s, 2H), 1.70 (s, 9H).

Tert-butyl 6-fluoro-3-(cyanomethyl)-1H-indole-1-carboxylate

(28) ##STR00014##

(29) 2-(6-fluoro-1H-indol-3-yl)acetonitrile (160.0 mg), DMAP (4.3 mg), Boc.sub.2O (238.0 mg). CH.sub.2Cl.sub.2 (4.4 mL). 12 h at room temperature. Aspect of the pure product: white solid. (Yield: 98%).

(30) .sup.1H NMR (CDCl.sub.3, 300 MHz) ppm: 7.91 (d, 1H), 7.63 (s, 1H), 7.50-7.42 (m, 1H), 7.07 (td, 1H), 3.78 (s, 2H), 1.69 (s, 9H).

Tert-butyl 6-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate

(31) ##STR00015##

(32) 2-(6-chloro-1H-indol-3-yl)acetonitrile (168.0 mg), DMAP (6.0 mg), Boc.sub.2O (233.0 mg). CH.sub.2Cl.sub.2 (4.0 mL). Aspect of the pure product: white solid. (Yield: 86%).

(33) .sup.1H NMR (CDCl.sub.3, 300 MHz) ppm: 8.25 (s, 1H), 7.64 (s, 1H), 7.45 (d, 1H), 7.29 (d, 1H), 3.77 (s, 2H), 1.70 (s, 9H).

B) Syntheses of Starting Aldehydes

3-formyl-4-methoxybenzonitrile

(34) ##STR00016##

(35) In a 10 mL reactor flask, 4-fluoro-3-formylbenzonitrile (900 mg, 6.04 mmol) was dissolved under argon in 2 mL of methanol. Sodium methoxide (1.232 mL, 6.64 mmol) was added and the reaction mixture was heated at reflux for 2 h.

(36) TLC showed no more starting material.

(37) The reaction mixture was poured into 10 mL of water. The resulting solid was filtered, washed with water, DIPE and dried in vacuo. The residue was purified by flash chromatography, eluted with a gradient from petroleum ether to MTBE give 587 mg of a grey solid (Yield: 59%).

(38) LC-MS: 98%

(39) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.28 (s, 1H), 8.12 (dd, J=8.8, 2.2 Hz, 1H), 8.05 (d, J=2.1 Hz, 1H), 7.43 (d, J=8.8 Hz, 1H), 4.01 (s, 3H).

3-formyl-4-(1H-1,2,4-triazol-1-yl)benzonitrile

(40) ##STR00017##

(41) In sealed microwave reactors, 4-fluoro-3-formylbenzonitrile (850 mg, 5.7 mmol) was dissolved in acetonitrile (15 mL), 1H-1,2,4-triazol (590 mg, 8.55 mmol, 1.5 eq) and K.sub.2CO.sub.3 (1575 mg, 11.39 mmol, 2 eq) were added to give a colorless suspension. Then the reaction mixture was stirred and heated at 80 C. for 5 min.

(42) The reaction mixture was poured into 20 mL of water, extracted with 220 mL of EtOAc. The combined organic layers were washed with 120 mL of water, 120 mL of brine, dried over Na.sub.2SO.sub.4, filtered and concentrated in vacuo to give an orange solid, m=983 mg.

(43) The solid was triturated with dichloromethane and petroleum ether, filtered and dried in vacuo at 45 C. overnight to give 493 mg of a brown powder (Yield: 43%).

(44) APCI-MS: (M+H).sup.+=199

(45) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.01 (s, 1H), 9.33 (s, 1H), 8.46-8.29 (m, 3H), 8.05 (d, J=9.0 Hz, 1H).

4-(1H-1,2,4-triazol-1-yl)nicotinaldehyde

(46) ##STR00018##

(47) Aspect of the product: yellow solid (Yield: 49%) APCI-MS: (M+H)+=175

(48) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.20 (s, 1H), 9.39 (s, 1H), 8.99-8.91 (m, 2H), 8.39 (s, 1H), 7.90 (d, J=5.5 Hz, 1H).

3-formyl-4-(4-fluorophenylthio)benzonitrile

(49) ##STR00019##

(50) In a 50 mL round-bottomed flask, 4-fluoro-3-formylbenzonitrile (900 mg, 5.73 mmol) and potassium carbonate (872 mg, 6.31 mmol) were suspended in DMF (10 mL) to give a yellow suspension. Then 4-fluorobenzenethiol (0.654 mL, 6.02 mmol) was added and the reaction mixture heated at 70 C. for 18 h. The reaction mixture was poured into water. The solid was filtered, washed with water and with a few amount of DIPE then dried in vacuo to give 1.44 g of a pale yellow solid (Yield: 97%).

(51) APCI-MS: (M+H).sup.+=257

(52) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.12 (s, 1H), 8.49 (d, J=1.9 Hz, 1H), 7.85 (dd, J=8.5, 2.0 Hz, 1H), 7.71-7.62 (m, 2H), 7.43 (ddd, J=10.9, 6.0, 2.6 Hz, 2H), 6.80 (d, J=8.5 Hz, 1H).

(53) The following examples were prepared according to the previous method.

4-(ethylthio)-3-formylbenzonitrile

(54) ##STR00020##

(55) Aspect of the product: yellow solid (Yield: 83%)

(56) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.07 (s, 1H), 8.38 (d, J=1.9 Hz, 1H), 7.99 (dd, J=8.4, 1.9 Hz, 1H), 7.63 (d, J=8.5 Hz, 1H), 3.08 (q, J=7.4 Hz, 2H), 1.30 (t, J=7.3 Hz, 3H).

4-(dimethylamino)-3-formylbenzonitrile

(57) ##STR00021##

(58) Aspect of the product: orange solid (Yield: 96%)

(59) APCI-MS: (M+H).sup.+=175

(60) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 9.88 (s, 1H), 8.07 (d, J=2.2 Hz, 1H), 7.72 (dd, J=8.9, 2.2 Hz, 1H), 7.09 (d, J=8.9 Hz, 1H), 2.99 (s, 6H).

4-(diethylamino)-3-formylbenzonitrile

(61) ##STR00022##

(62) Aspect of the product: yellow solid (Yield: 83%)

(63) APCI-MS: (M+H).sup.+=203

(64) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 9.91 (s, 1H), 8.00 (d, J=2.2 Hz, 1H), 7.78 (dd, J=8.9, 2.2 Hz, 1H), 7.21 (d, J=8.9 Hz, 1H), 3.44-3.35 (m, 4H), 1.11 (t, J=7.0 Hz, 6H).

4-dimethylamino-3-formyl-pyridine

(65) ##STR00023##

(66) A mixture of 4-chloronicotinaldehyde (500 mg, 3.53 mmol), potassium carbonate (976 mg, 7.06 mmol) and dimethylamine in THF (2.65 mL, 5.30 mmol) was heated at 80 C. for 3 hours.

(67) TLC (eluent EtOAc) showed no more starting material.

(68) The reaction mixture was concentrated under pressure. Purification by flash chromatography on silica gel column (eluant: CH.sub.2Cl.sub.2/MeOH 90/10) yielded 0.48 g of a pale yellow solid (Yield: 90%).

(69) APCI-MS: (M+H).sup.+=151

(70) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 9.93 (s, 1H), 8.60 (s, 1H), 8.23 (d, J=6.1 Hz, 1H), 6.87 (d, J=6.2 Hz, 1H), 2.99 (s, 6H).

4-fluorophenoxy-3-formylpyridine

(71) ##STR00024##

(72) To a solution of 4-fluorophenol (455 mg, 4.06 mmol) in THF [5 mL] was added HNa (162 mg, 4.06 mmol). After stirring for 0.5 hour, 4-chloronicotinaldehyde (500 mg, 3.53 mmol) was added and the reaction mixture was heated at 65 C. for 3 hours.

(73) Then the reaction mixture was diluted with water and brine, extracted with MTBE, dried over MgSO.sub.4 and concentrated under reduced pressure to give 617 mg of an oily compound (Yield: 64%).

(74) APCI-MS: (M+H).sup.+=218

(75) The following example was prepared as the previous method.

4-fluorophenoxy-3-formylbenzonitrile

(76) ##STR00025##

(77) Aspect of the product: yellow solid (Yield: 64%)

(78) APCI-MS: (MH)=240

(79) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.41 (s, 1H), 8.23 (d, J=2.2 Hz, 1H), 8.03 (dd, J=8.8, 2.2 Hz, 1H), 7.40-7.30 (m, 4H), 6.95 (d, J=8.8 Hz, 1H).

4-fluorophenylthio-3-formylpyridine

(80) ##STR00026##

(81) A mixture of 4-chloronicotinaldehyde (500 mg, 3.53 mmol), potassium carbonate (537 mg, 3.89 mmol) and 4-fluorobenzenethiol (0.403 mL, 3.71 mmol) in DMF (10 mL) was heated at 70 C. for 1 h.

(82) The reaction mixture was quenched with water, extracted with 320 mL of AcOEt, the combined organic were washed with brine, dried over Na.sub.2SO.sub.4, filtered and concentrated under vacuum to give a brown oil. The oil was triturated with 5 mL of DIPE to give 0.46 g of a beige solid (Yield: 55%).

(83) APCI-MS: (M+H).sup.+=234

(84) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.19 (s, 1H), 9.03 (s, 1H), 8.45 (d, J=5.6 Hz, 1H), 7.69 (dd, J=8.5, 5.5 Hz, 2H), 7.45 (t, J=8.7 Hz, 2H), 6.61 (d, J=5.6 Hz, 1H).

(85) The following examples were prepared as the previous method.

4-(pyridin-2-ylthio)nicotinaldehyde

(86) ##STR00027##

(87) Aspect of the product: yellow solid (Yield: 61%)

(88) APCI-MS: (M+H).sup.+=217

(89) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.18 (s, 1H), 9.04 (s, 1H), 8.68 (d, J=3.7 Hz, 1H), 8.51 (d, J=5.5 Hz, 1H), 7.95 (td, J=7.7, 1.9 Hz, 1H), 7.74 (d, J=7.8 Hz, 1H), 7.51 (dd, J=6.5, 4.8 Hz, 1H), 7.00 (d, J=5.5 Hz, 1H).

3-formyl-4-(4-trifluorophenyl)benzonitrile

(90) ##STR00028##

(91) In a 50 mL pear flask, magnetic stirrer, 4-chloronicotinaldehyde (500 mg, 3.53 mmol), 4-trifluoromethyl)phenylboronic acid (671 mg, 3.53 mmol), triphenylphosphine (55.6 mg, 0.212 mmol), palladium(II) acetate (47.6 mg, 0.212 mmol) and potassium carbonate (976 mg, 7.06 mmol) were added successively followed by 1,2-dimethoxyethane (10 mL) and water (2.5 mL). The reaction mixture was stirred and heated at 85 C. for 18 hours (LC/MS showed no starting material).

(92) 20 mL of water and 20 mL of ethyl acetate were added. The mixture was filtered over Celite and the cake rinsed with 20 mL of ethyl acetate. Organic phases were washed twice with brine, dried over sodium sulfate, filtered and the solvent was removed to give 890 mg of an oil.

(93) The crude oil was purified by flash chromatography on SiO.sub.2, eluted with 100% dichloromethane then 95/5 dichloromethane/acetone to give 370 mg of a grey solid (Yield: 42%).

(94) APCI-MS: (M+H).sup.+=252

(95) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.00 (s, 1H), 9.08 (s, 1H), 8.90 (d, J=5.1 Hz, 1H), 7.92 (d, J=8.1 Hz, 2H), 7.78 (d, J=8.0 Hz, 2H), 7.61 (dd, J=5.1, 0.6 Hz, 1H).

(96) The following example was prepared as the previous method.

4-(furan-3-yl)nicotinaldehyde

(97) ##STR00029##

(98) Aspect of the product: yellow solid (Yield: 64%)

(99) LC-MS: (M+H).sup.+=174

(100) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 10.24 (s, 1H), 8.96 (s, 1H), 8.78 (s, 1H), 8.25 (s, 1H), 7.90 (s, 1H), 7.62 (s, 1H), 6.99 (s, 1H).

Preparation of Examples 2 to 53

Method A: Knoevenagel Condensation

(101) ##STR00030##

(102) To a solution of cyanomethyl-indole-1-carboxylic acid tert-butyl ester (1 eq.) in THF was added sodium hydride (1.5 eq.) under an argon atmosphere. The reaction apparatus was protected from light and the mixture stirred at room temperature for 1 hour. Then the reaction mixture was cooled to 0 C. and the aldehyde (1.2 eq.) was added in portions.

(103) The mixture was stirred at room temperature for 24 or 48 hours, then quenched with saturated ammonium chloride aqueous solution and extracted with AcOEt. The combined organic layers were dried over MgSO.sub.4 and evaporated under vacuo. When the desired compound protected with a Boc group remained as a side product of the reaction, the reaction mixture was treated with a solution of HCl in dioxane or an aqueous solution of NaOH 1N to complete the Boc deprotection. Then the crude residue was triturated with a minimum of solvent (MeOH or CH.sub.2Cl.sub.2 or Et.sub.2O), filterated, washed with Et.sub.2O, and dried in vacuo for 12 hours (in dark) to afford the corresponding acrylonitrile.

Method B: SN.SUB.AR .reactions from (Z)-2-(5-methoxy-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile or (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile

(104) To a solution of 4-chloropyridine derivative (Z)-2-(5-methoxy-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile or (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (1 eq.) in MeOH, EtOH or isopropanol was added KOH (2 to 5 eq.) under an argon atmosphere. The reaction apparatus was protected from light and the mixture was refluxed overnight. The mixture was diluted with AcOEt, washed with water then brine. The organic layer was dried over MgSO.sub.4 and reduced in vacuo. The crude product was purified by trituration with AcOEt or by flash chromatography to afford the SN.sub.AR derivative.

Method C: SN.SUB.AR .reactions from (Z)-2-(5-methoxy-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile or (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile

(105) To a solution of 4-chloropyridine derivative-(Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (1 eq.) in DMF was added NaSMe or NaSEt (2 eq.) under an argon atmosphere. The reaction apparatus was protected from light and the mixture was stirred overnight at room temperature. The mixture was diluted with AcOEt, washed with water then brine. The organic layer was dried over MgSO.sub.4 and reduced in vacuo. The crude product was purified by trituration with AcOEt to afford the SN.sub.AR derivative.

Method D: SN.SUB.AR .reactions from (Z)-2-(5-methoxy-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile or (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile

(106) To a solution of 4-chloropyridine derivative (Z)-2-(5-methoxy-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile or (1 eq.) in DMF were added arylthiol (1.1 eq.) and sodium or potassium carbonate (2 eq.) under an argon atmosphere. The reaction apparatus was protected from light and the mixture was stirred overnight at room temperature. The mixture was diluted with AcOEt, washed with water then brine. The organic layer was dried over MgSO.sub.4 and reduced in vacuo. The crude product was purified by trituration with AcOEt to afford the SN.sub.AR derivative.

Method E: E Isomers

(107) Z isomers were dissolved in ethanol and subjected to a 150 W halogen lamp with a continuous argon flux until there was no more starting material (TLC). The solution was then concentrated and the residue was purified by C18 chromatography to give the title compound.

Method F: Synthesis of (Z)-2-(6-X-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(108) To a solution of tert-butyl 6-X-3-(cyanomethyl)-1H-indole-1-carboxylate (1 eq) in THF, was added NaH (eq). The resulting mixture was stirred 10 min at room temperature and 4-methoxynicotinaldehyde (1.3 eq) was added with one drop of DMF. The mixture was stirred at room temperature hidden from light. The reaction was quenched with aqueous NH.sub.4Cl and extracted with AcOEt, dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was dissolved with THF and NaOH 2.5 M was added. The system was stirred at room temperature hidden from light, diluted with AcOEt, dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was taken off with a minimal amount of AcOEt and filtrated to give the title compound.

Method G: Synthesis of (Z)-3-(2-(5-X-1H-indol-3-yl)-2-cyanovinyl)-4-methoxypyridine 1-oxide

(109) To a solution of (Z)-2-(5-X-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile in THF was added m-CPBA (1 eq), the resulting mixture was stirred 12 h at room temperature, hidden from light and a new portion of m-CPBA (0.5 eq) was added. After additional 4 h of stirring, the mixture was concentrated and the residue was triturated in AcOEt and filtrated to give the title compound.

Method H: Synthesis of (Z)-3-(2-(5-X-1H-indol-3-yl)-2-cyanovinyl)-4-(trifluoromethoxy) benzonitrile

(110) To a solution of tert-butyl 5-X-3-(cyanomethyl)-1H-indole-1-carboxylate (1 eq) in THF, was added NaH (3 eq). The resulting mixture was stirred 10 min at room temperature and 3-formyl-4-(trifluoromethoxy)benzonitrile (1 eq) was added with one drop of DMF. The mixture was stirred at room temperature hidden from light. The reaction was quenched with aqueous NH.sub.4Cl and extracted with AcOEt, dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was dissolved with THF and NaOH 2.5 M was added. The system was stirred at room temperature hidden from light, diluted with AcOEt, dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was purified by silicagel chromatography (CH.sub.2Cl.sub.2/MeOH, 100:0 to 90:10) to give the title compound.

Example 2

(Z)-3-(4-ethoxypyridin-3-yl)-2-(5-methoxy-1H-indol-3-yl)acrylonitrile

(111) ##STR00031##

Method B

(112) (Z)-2-(5-methoxy-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (50 mg), KOH (45 mg) and EtOH (1.5 mL). Trituration with AcOEt. Aspect of the pure product: yellow solid. (Yield: 67%).

(113) ESI-MS: (M+H).sup.+=320

(114) .sup.1H NMR (Acetone-d.sub.6, 300 MHz) ppm: 9.05 (s, 1H), 8.47 (d, J=5.8 Hz, 1H), 7.81 (s, 1H), 7.75 (s, 1H), 7.49 (d, J=2.3 Hz, 1H), 7.47 (d, J=9.0 Hz, 1H), 7.11 (d J=5.8 Hz, 1H), 6.93 (dd, J=9.0 Hz, J=2.3 Hz, 1H), 4.40 (q, J=7.0 Hz, 1H), 3.88 (s, 3H), 1.49 (t, J=7.0 Hz, 1H).

Example 3

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-chloropyridin-3-yl)acrylonitrile

(115) ##STR00032##

Method A

(116) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (670 mg). Sodium hydride (138 mg). THF 17 mL. 4-chloronicotinaldehyde (457 mg). Trituration of the crude product with MeOH. Aspect of the pure product orange solid. (Yield: 38%) ESI-MS: (M+H).sup.+=314

(117) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 12.1 (s, 1H), 9.05 (s, 1H), 8.6 (d, 1H), 8.06 (d, 1H), 8.03 (d, 1H), 7.8 (s, 1H), 7.75 (d, 1H), 7.57 (dd, 1H), 7.3 (dd, 1H).

Example 4

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-chloropyridin-3-yl)acrylonitrile

(118) ##STR00033##

Method A

(119) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (600 mg). Sodium hydride (100 mg). THF 17 mL. 4-chloronicotinaldehyde (355 mg). Trituration of the crude product with dichloromethane and then washed with methanol and ether. Aspect of the pure product orange brown solid. (Yield: 50%).

(120) ESI-MS: (M+H).sup.+=358

(121) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 12.1 (s, 1H), 9.03 (s, 1H), 8.62 (d, 1H), 8.2 (d, 1H), 8.0 (d, 1H), 7.8 (s, 1H), 7.76 (d, 1H), 7.55 (d, 1H), 7.4 (dd, 1H).

Example 5

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(122) ##STR00034##

Method B

(123) (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (80 mg), KOH (25 mg), MeOH (5 mL) and THF (2 mL). The mixture was refluxed for 24 hours. Purification by flash chromatography (CH.sub.2Cl.sub.2/MeOH 100/0 to 96/3). Aspect of the pure product: yellow solid. (Yield: 66%).

(124) ESI-MS: (M+H).sup.+=354

(125) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 11.96 (s, 1H), 8.83 (s, 1H), 8.52 (d, J=8.8 Hz, 1H), 8.09 (d, J=1.9 Hz, 1H), 7.90 (d, J=2.6 Hz, 1H), 7.68 (s, 1H), 7.48 (d, J=8.7 Hz, 1H), 7.36 (dd, J=8.7 Hz, J=1.9 Hz, 1H), 7.20 (d, J=5.8 Hz, 1H), 3.95 (s, 3H).

Example 5b

(E)-2-(5-bromo-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(126) ##STR00035##

Method E

(127) (Z)-2-(5-bromo-1H-indol-3-yl)-3-(2-methoxyphenyl)acrylonitrile (30 mg). EtOH (40 mL). Reaction time: 18 h. Aspect of the pure product: yellow solid. (Yield: 40%).

(128) ESI-MS: (M+H)=354

(129) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.31 (d, 1H), 8.00 (s, 1H), 7.55 (s, 1H), 7.42 (s, 1H), 7.35 (d, 1H), 7.26-7.14 (m, 2H), 6.96 (d, 1H), 4.00 (s, 3H).

Example 6

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-(dimethylamino)pyridin-3-yl)acrylonitrile

(130) ##STR00036##

Method A

(131) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). Sodium hydride (28.9 mg). THF 2 mL. 4-dimethylaminonicotinaldehyde (93 mg). Trituration of the crude product with water and diisopropylether. Aspect of the product pale yellow solid (Yield: 82%).

(132) APCI-MS: (M+H).sup.+=323

(133) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.92 (s, 1H), 8.52 (s, 1H), 8.26 (d, J=5.8 Hz, 1H), 7.98 (d, J=1.9 Hz, 1H), 7.90 (s, 1H), 7.59 (s, 1H), 7.51 (d, J=8.7 Hz, 1H), 7.23 (dd, J=8.7, 1.9 Hz, 1H), 6.93 (d, J=5.9 Hz, 1H), 1.16-1.10 (m, 0.33H), 1.07 (t, J=7.0 Hz, 5.6H).

Example 7

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-(dimethylamino)pyridin-3-yl)acrylonitrile, hydrochloride

(134) ##STR00037##

(135) In a 25 mL pear flask, (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-(dimethylamino)pyridin-3-yl)acrylonitrile (60 mg, 0.186 mmol) was dissolved in ethanol (1 mL) and dichloromethane (0.5 mL) to give a yellow solution followed by addition of HCl 37% in water (0.015 mL, 0.186 mmol).

(136) The reaction mixture was concentrated under reduced pressure to give 67 mg of a yellow solid (Yield: 100%).

(137) APCI-MS: (M+H).sup.+=323

(138) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 14.47-13.49 (m, 1H), 12.09 (s, 1H), 8.53 (s, 1H), 8.28 (d, J=7.3 Hz, 1H), 8.08 (s, 1H), 7.91 (s, 2H), 7.53 (d, J=8.7 Hz, 1H), 7.25 (d, J=10.6 Hz, 1H), 7.14 (d, J=7.3 Hz, 1H), 3.25 (s, 6H).

Example 8

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(dimethylamino)pyridin-3-yl)acrylonitrile

(139) ##STR00038##

Method A

(140) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). Sodium hydride (25 mg). THF 2 mL. 4-dimethylaminonicotinaldehyde (81 mg). Trituration of the crude product with water and ethanol. Aspect of the product pale yellow solid (Yield: 61%)

(141) APCI-MS: (M+H).sup.+=367

(142) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.92 (s, 0.9H), 11.85-11.77 (m, 0.1H), 8.51 (s, 1H), 8.26 (d, J=5.9 Hz, 1H), 8.10 (s, 1H), 7.88 (d, J=2.8 Hz, 1H), 7.62 (s, 1H), 7.46 (d, J=8.7 Hz, 1H), 7.39-7.28 (m, 1H), 6.89 (d, J=5.9 Hz, 1H), 2.92 (s, 5.6H), 2.85 (s, 0.4H).

Example 9

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(143) ##STR00039##

Method A

(144) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (276 mg). THF 7 mL. Sodium hydride (57 mg), 4-methoxypyridine-3-carboxaldehyde (156 mg). Reaction time 24 hours. Trituration of the crude product with MeOH. Aspect of the pure product: orange solid. (Yield: 35%).

(145) ESI-MS: (M+H).sup.+=310

(146) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 11.97 (s, 1H), 8.82 (s, 1H), 8.51 (d, J=5.8 Hz, 1H), 7.94 (d, J=1.8 Hz, 1H), 7.91 (s, 1H), 7.68 (s, 1H), 7.52 (d, J=8.7 Hz, 1H), 7.24 (dd, J=8.7, 1.9 Hz, 1H), 7.19 (d, J=5.8 Hz, 1H), 3.94 (s, 3H).

Example 9b

(E)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(147) ##STR00040##

Method E

(148) (Z)-2-(5-chloro-1H-indol-3-yl)-3-(2-methoxyphenyl)acrylonitrile (20 mg). EtOH (35 mL). Reaction time: 8 h. Aspect of the pure product: yellow solid. (Yield: 30%).

(149) ESI-MS: (M+H).sup.+=310

(150) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.31 (d, 1H), 8.01 (s, 1H), 7.56 (s, 1H), 7.42 (s, 1H), 7.38 (d, 1H), 7.16 (d, 1H), 7.10 (d, 1H), 6.83 (d, 1H), 4.00 (s, 3H).

Example 10

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-phenoxypyridin-3-yl)acrylonitrile

(151) ##STR00041##

Method A

(152) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (190 mg). THF 6 mL. Sodium hydride (40 mg), 4-phenoxypyridine-3-carboxaldehyde (160 mg). Reaction time 24 hours. The reaction mixture was not extracted. A precipitate was formed in the reaction mixture, filtered and washed with ether. Trituration of the precipitate with MeOH. Aspect of the pure product: yellow orange solid. (Yield: 73%).

(153) ESI-MS: (MH)=370

(154) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 8.98 (s, 1H), 8.46 (d, J=5.7 Hz, 1H), 7.99 (s, 1H), 7.87 (s, 1H), 7.73 (s, 1H), 7.62 (d, 1H), 7.50 (m, 2H), 7.35-7.2 (m, 3H), 7.17 (dd, J=8.7, 1H), 6.75 (d, J=5.7 Hz, 1H).

Example 11

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-phenoxypyridin-3-yl)acrylonitrile

(155) ##STR00042##

Method A

(156) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (224 mg). THF 4.9 mL. Sodium hydride (40 mg), 4-phenoxypyridine-3-carboxaldehyde (160 mg). Reaction time 24 hours. Trituration of the crude product with MeOH. Aspect of the pure product: yellow solid. (Yield: 18.7%).

(157) ESI-MS: (MH)=414

(158) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 12.01 (s, 1H), 9.0 (s, 1H), 8.48 (d, J=5.7 Hz, 1H), 8.11 (d, J=1.7 Hz, 1H), 7.94 (s, 1H), 7.83 (s, 1H), 7.55-7.48 (m, 3H), 7.4-7.25 (m, 4H), 6.76 (d, J=5.7 Hz, 1H).

Example 12

(Z)-2-(5-methoxy-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(159) ##STR00043##

Method B

(160) (Z)-2-(5-methoxy-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (50 mg), KOH (45 mg) and MeOH (1.5 mL). Trituration with AcOEt. Aspect of the pure product: yellow solid. (Yield: 71%).

(161) ESI-MS: (M+H).sup.+=306

(162) .sup.1H NMR (Acetone-d.sub.6, 300 MHz) ppm: 9.02 (s, 1H), 8.50 (d, J=5.7 Hz, 1H), 7.77 (s, 1H), 7.75 (s, 1H), 7.50 (d, J=2.3 Hz, 1H), 7.46 (d, J=8.9 Hz, 1H), 7.14 (d, J=5.7 Hz, 1H), 6.93 (dd, J=8.9 Hz, J=2.3 Hz, 1H), 4.03 (s, 3H), 3.87 (s, 3H).

Example 13

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-ethoxypyridin-3-yl)acrylonitrile

(163) ##STR00044##

Method B

(164) (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (40 mg), KOH (31 mg), EtOH (5 mL). Trituration with AcOEt. Aspect of the pure product: yellow solid. (Yield: 76%).

(165) ESI-MS: (M+H).sup.+=368

(166) .sup.1H NMR (acetone-d.sub.6, 300 MHz) ppm: 9.06 (s, 1H), 8.49 (d, J=5.5 Hz, 1H), 8.20 (s, 1H), 7.85 (s, 1H), 7.55 (d, J=8.5 Hz, 1H), 7.40 (d, J=8.2 Hz, 1H), 7.12 (d, J=5.8 Hz, 1H), 4.31 (q, J=7.0 Hz, 2H), 1.54 (t, J=7.0 Hz, 3H).

Example 14

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-isopropoxypyridin-3-yl)acrylonitrile

(167) ##STR00045##

Method B

(168) (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (70 mg), KOH (55 mg), isopropanol (2 mL). Purification by flash chromatography (CH.sub.2Cl.sub.2/MeOH 100/0 to 95/5). Aspect of the pure product: yellow solid. (Yield: 20%).

(169) ESI-MS: (M+H).sup.+=382

(170) .sup.1H NMR (MeOD, 300 MHz) ppm: 9.02 (s, 1H), 8.42 (d, J=5.8 Hz, 1H), 8.09 (d, J=1.5 Hz, 1H), 7.75 (s, 1H), 7.69 (s, 1H), 7.42 (d, J=8.7 Hz, 1H), 7.36 (dd, J=8.7 Hz, J=1.5 Hz, 1H), 7.19 (d, J=5.8 Hz, 1H), 4.93 (sept, J=6.0 Hz, 1H), 1.50 (d, J=6.0 Hz, 6H).

Example 15

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(methylthio)pyridin-3-yl)acrylonitrile

(171) ##STR00046##

Method C

(172) (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (80 mg), NaSMe (31 mg), DMF (1 mL). Aspect of the pure product: yellow solid. (Yield: 61%).

(173) ESI-MS: (M+H).sup.+=370

(174) .sup.1H NMR (acetone-d.sub.6, 300 MHz) ppm: 11.12 (s, 1H), 8.83 (s, 1H), 8.48 (d, J=5.3 Hz, 1H), 8.24 (d, J=1.9 Hz, 1H), 7.91 (s, 1H), 7.67 (s, 1H), 7.57 (d, J=8.7 Hz, 1H), 7.41 (dd, J=8.7 Hz, J=1.9 Hz, 1H), 7.39 (d, J=5.3 Hz, 1H), 2.66 (s, 3H).

Example 16

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(ethylthio)pyridin-3-yl)acrylonitrile

(175) ##STR00047##

Method C

(176) (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (80 mg), NaSEt (37 mg), DMF (1 mL). Aspect of the pure product: yellow solid. (Yield: 70%).

(177) ESI-MS: (M+H).sup.+=384

(178) .sup.1H NMR (acetone-d.sub.6, 300 MHz) ppm: 11.13 (s, 1H), 8.86 (s, 1H), 8.47 (d, J=5.5 Hz, 1H), 8.26 (d, J=1.7 Hz, 1H), 7.91 (s, 1H), 7.68 (s, 1H), 7.56 (d, J=8.7 Hz, 1H), 7.45-7.39 (m, 2H), 3.21 (q, J=7.5 Hz, 2H), 1.42 (t, J=7.5 Hz, 3H).

Example 17

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(3-bromophenyl)pyridin-3-yl)acrylonitrile

(179) ##STR00048##

Method A

(180) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (300 mg). THF 6.6 mL. Sodium hydride (54 mg), 4-(3-bromophenyl)-3-pyridinecarboxaldehyde (328 mg). Reaction time 24 hours. Trituration of the crude product with MeOH. Aspect of the pure product: yellow solid. (Yield: 65%).

(181) ESI-MS: (M+H).sup.+=478

(182) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 11.96 (s, 1H), 9.04 (s, 1H), 8.72 (d, 1H), 7.84 (s, 1H), 7.79-7.68 (m, 3H), 7.65-7.43 (m, 5H), 7.32 (dd, 1H).

Example 18

(Z)-3-(4-(3-bromophenyl)pyridin-3-yl)-2-(5-chloro-1H-indol-3-yl)acrylonitrile

Method A

(183) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (266 mg). THF 6.8 mL. Sodium hydride (55 mg), 4-(3-bromophenyl)-3-pyridinecarboxaldehyde (336 mg). Reaction time 24 hours. Trituration of the crude product with MeOH. Aspect of the pure product: yellow solid. (Yield: 52%).

(184) ##STR00049##

(185) ESI-MS: (M+H).sup.+=434

(186) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 11.96 (s, 1H), 9.04 (s, 1H), 8.72 (d, 1H), 7.86 (s, 1H), 7.81-7.68 (m, 3H), 7.66-7.46 (m, 5H), 7.22 (dd, 1H).

Example 19

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(phenylthio)pyridin-3-yl)acrylonitrile

(187) ##STR00050##

Method D

(188) (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (80 mg), thiophenol (25 l), sodium carbonate (47 mg), DMF (1 mL). Aspect of the pure product: yellow solid. (Yield: 53%).

(189) ESI-MS: (M+H).sup.+=432

(190) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 12.05 (s, 1H), 8.81 (s, 1H), 8.40 (d, J=5.3 Hz, 1H), 8.15 (d, J=1.7 Hz, 1H), 7.96 (s, 1H), 7.73 (s, 1H), 7.62 (m, 2H), 7.55 (m, 3H), 7.50 (d, J=8.7 Hz, 1H), 7.38 (dd, J=8.7 Hz, J=1.7 Hz, 1H), 6.81 (d, J=5.3 Hz, 1H).

Example 20

(Z)-3-(4-(benzylthio)pyridin-3-yl)-2-(5-bromo-1H-indol-3-yl)acrylonitrile

(191) ##STR00051##

Method D

(192) (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (50 mg), benzyl mercaptan (18 L), sodium carbonate (30 mg), DMF (1 mL). Aspect of the pure product: yellow solid. (Yield: 47%).

(193) ESI-MS: (M+H).sup.+=446

(194) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 12.01 (s, 1H), 8.73 (s, 1H), 8.58 (d, J=5.2 Hz, 1H), 8.12 (d, J=1.2 Hz, 1H), 7.92 (s, 1H), 7.60 (s, 1H), 7.56 (d, J=5.2 Hz, 1H), 7.49-7.47 (m, 3H), 7.37-7.33 (m, 3H), 7.28 (m, 1H), 4.47 (s, 2H).

Example 21

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-((3,4-dimethoxyphenyl)thio)pyridin-3-yl)acrylonitrile

(195) ##STR00052##

Method D

(196) (Z)-2-(5-Bromo-1H-indol-3-yl)-3-(4-chloro-3-yl)acrylonitrile (61 mg), 3,4-dimethoxythiophenol (37 L), sodium carbonate (36 mg), DMF (1 mL). Aspect of the pure product: yellow solid. (Yield: 50%).

(197) ESI-MS: (M+H).sup.+=492

(198) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 12.05 (s, 1H), 8.76 (s, 1H), 8.36 (d, J=5.2 Hz, 1H), 8.17 (s, 1H), 7.96 (s, 1H), 7.70 (s, 1H), 7.51 (d, J=8.5 Hz, 1H), 7.39 (d, J=8.5 Hz, 1H), 7.22 (d, J=8.2 Hz, 1H), 7.19 (s, 1H), 7.13 (d, J=8.5 Hz, 1H), 6.74 (d, J=5.2 Hz, 1H), 3.82 (s, 3H), 3.76 (s, 3H).

Example 22

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(4-fluorophenoxy)pyridin-3-yl)acrylonitrile

(199) ##STR00053##

Method A

(200) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (200 mg). THF 2 mL. Sodium hydride (31.7 mg), 4-(4-fluorophenyl)-3-pyridinecarboxaldehyde (148 mg). Reaction time 1 h 30. Trituration of the crude product with DCM. Aspect of the product: yellow solid. (Yield: 10%).

(201) LC-MS: (M+H).sup.+=434

(202) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 12.01 (s, 1H), 8.98 (s, 1H), 8.46 (d, J=5.7 Hz, 1H), 8.11 (d, J=1.7 Hz, 1H), 7.94 (s, 1H), 7.83 (s, 1H), 7.48 (d, J=8.7 Hz, 1H), 7.38-7.31 (m, 5H), 6.73 (d, J=5.7 Hz, 1H).

Example 23

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-(4-fluorophenoxy)pyridin-3-yl)acrylonitrile

(203) ##STR00054##

Method A

(204) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (200 mg). THF 2 mL. Sodium hydride (35.1 mg), 4-(4-fluorophenyl)-3-pyridinecarboxaldehyde (163 mg). Reaction time 1 hour 30 minutes. Trituration of the crude product with DCM. Aspect of the product: yellow solid. (Yield: 82%).

(205) APCI-MS: (M+H).sup.+=390

(206) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 8.98 (s, 1H), 8.46 (d, J=5.7 Hz, 1H), 7.99-7.92 (m, 2H), 7.83 (s, 1H), 7.52 (d, J=8.7 Hz, 1H), 7.34 (d, J=5.9 Hz, 4H), 7.24 (dd, J=8.7, 1.8 Hz, 1H), 6.73 (d, J=5.7 Hz, 1H).

Example 24

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(diethylamino)pyridin-3-yl)acrylonitrile

(207) ##STR00055##

Method A

(208) Aspect of the product: yellow solid (Yield: 60%)

(209) APCI-MS: (M+H).sup.+=395

(210) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.93 (s, 0.83H), 11.88-11.76 (m, 0.12H), 8.52 (s, 1H), 8.27 (d, J=5.8 Hz, 1H), 8.12 (d, J=1.7 Hz, 1H), 7.88 (s, 1H), 7.59 (s, 1H), 7.51-7.21 (m, 2H), 6.93 (d, J=5.9 Hz, 1H), 3.32-3.24 (m, 4H), 1.07 (t, J=7.0 Hz, 6H).

Example 25

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(4-(trifluoromethyl)phenyl)pyridin-3-yl)acrylonitrile

(211) ##STR00056##

Method A

(212) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (120 mg). THF 3 mL. Sodium hydride (20.05 mg), 4-(4-trifluoromethylphenyl)-3-pyridinecarboxaldehyde (108 mg). Reaction time 16 hours. Purification by chromatography on 24 g Redisep column 20-40 m, eluted with a gradient of CH.sub.2Cl.sub.2/MeOH from 100/00 to 95/05. Dissolution of the solid in EtOH (3 mL) and water (0.3 mL) and concentration in a Genevac evaporator to remove the traces of solvent. Aspect of the product: yellow solid. (Yield: 57%).

(213) APCI-MS: (M+H).sup.+=468

(214) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 12.19-11.57 (s, 1H), 9.11 (s, 1H), 8.80-8.61 (d, 1H), 7.66 (m, 11H).

Example 26

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-(4-(trifluoromethyl)phenyl)pyridin-3-yl)acrylonitrile

(215) ##STR00057##

Method A

(216) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (120 mg). THF 3 mL. Sodium hydride (23.11 mg), 4-(4-trifluoromethylphenyl)-3-pyridinecarboxaldehyde (124 mg). Reaction time 16 hours. Purification by chromatography on 24 g Redisep column 20-40 m, eluted with a gradient of CH.sub.2Cl.sub.2/MeOH from 100/00 to 95/05. Dissolution of the solid in EtOH (3 mL) and water (0.3 mL) and concentration in a Genevac evaporator to remove the traces of solvent. Aspect of the product: yellow solid. (Yield: 68%).

(217) APCI-MS: (M+H).sup.+=424

(218) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 12.40-11.44 (m, 1H), 9.10 (s, 1H), 8.73 (d, J=5.1 Hz, 1H), 7.91-7.16 (m, 11H).

Example 27

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-((4-fluorophenyl)thio)pyridin-3-yl)acrylonitrile

(219) ##STR00058##

Method A

(220) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). THF 2 mL. Sodium hydride (28.9 mg), 4-(4-fluorophenylthio)-3-pyridinecarboxaldehyde (144 mg). Reaction time 16 hours. Trituration of the crude product with heptane and diisopropylether. Aspect of the product: yellow solid. (Yield: 42%).

(221) APCI-MS: (M+H).sup.+=406

(222) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 8.79 (s, 1H), 8.38 (d, J=5.4 Hz, 1H), 7.97 (s, 2H), 7.68 (s, 4H), 7.56 (d, J=8.6 Hz, 1H), 7.42 (d, J=8.6 Hz, 3H), 7.26 (s, 1H), 6.76 (d, J=5.4 Hz, 1H).

Example 28

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-((4-fluorophenyl)thio)pyridin-3-yl)acrylonitrile

(223) ##STR00059##

Method A

(224) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). THF 2 mL. Sodium hydride (25.06 mg), 4-(4-fluorophenylthio)-3-pyridinecarboxaldehyde (125 mg). Reaction time 16 hours. Trituration of the crude product with heptane and ether.

(225) Aspect of the product: yellow solid. (Yield: 32%).

(226) APCI-MS: (M+H).sup.+=450

(227) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.98 (sl, 1H), 8.82-8.74 (m, 1H), 8.43-8.35 (m, 1H), 8.18-8.13 (m, 1H), 7.99-7.93 (m, 1H), 7.70 (dd, J=8.6, 2.7 Hz, 3H), 7.44 (qd, J=8.6, 7.1 Hz, 4H), 6.80-6.73 (m, 1H).

Example 29

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-(furan-3-yl)pyridin-3-yl)acrylonitrile

(228) ##STR00060##

Method A

(229) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). THF 3 mL. Sodium hydride (17.33 mg), 4-(furan-3-yl)nicotinaldehyde 107 mg). Reaction time 16 hours. Trituration of the crude product with heptane and diisopropylether. Aspect of the product: yellow solid. (Yield: 21%).

(230) APCI-MS: (M+H).sup.+=346

(231) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.99 (s, 1H), 8.93 (s, 1H), 8.64 (d, J=5.2 Hz, 1H), 8.10 (s, 1H), 8.01-7.92 (m, 2H), 7.87 (s, 1H), 7.77 (s, 1H), 7.64 (d, J=5.1 Hz, 1H), 7.53 (d, J=8.7 Hz, 1H), 7.25 (dd, J=8.7, 1.9 Hz, 1H), 6.97 (s, 1H).

Example 30

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-(pyridin-2-ylthio)pyridin-3-yl)acrylonitrile

(232) ##STR00061##

Method A

(233) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). THF 2 mL. Sodium hydride (28.9 mg), 4-(piridin-2-yl)thio-nicotinaldehyde (134 mg). Reaction time 16 hours. Trituration of the crude product with heptane and diisopropylether.

(234) Aspect of the product: yellow solid. (Yield: 55%).

(235) APCI-MS: (M+H).sup.+=389

(236) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 9.00 (s, 1H), 8.60-8.52 (m, 1H), 8.48 (d, J=3.8 Hz, 1H), 7.86 (s, 1H), 7.81-7.73 (m, 1H), 7.68 (s, 1H), 7.59 (s, 1H), 7.52 (d, J=5.3 Hz, 2H), 7.43 (d, J=7.9 Hz, 1H), 7.28 (dd, J=7.0, 5.2 Hz, 1H), 7.20-7.12 (m, 1H).

Example 31

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(pyridin-2-ylthio)pyridin-3-yl)acrylonitrile

(237) ##STR00062##

Method A

(238) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). THF 2 mL. Sodium hydride (25.06 mg), 4-(piridin-2-yl)thio-nicotinaldehyde (116 mg). Reaction time 16 hours. Trituration of the crude product with heptane and ether. Aspect of the product: yellow solid. (Yield: 65%).

(239) APCI-MS: (M+H).sup.+=433

(240) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 9.00 (s, 1H), 8.62-8.57 (m, 1H), 8.52-8.46 (m, 1H), 7.91-7.86 (m, 2H), 7.78 (ddd, J=9.5, 7.7, 1.8 Hz, 1H), 7.68 (s, 1H), 7.56-7.52 (m, 1H), 7.49-7.42 (m, 2H), 7.37-7.26 (m, 2H).

Example 32

(Z)-3-(4-(1H-1,2,4-triazol-1-yl)pyridin-3-yl)-2-(5-bromo-1H-indol-3-yl)acrylonitrile

(241) ##STR00063##

Method A

(242) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). THF 2 mL. Sodium hydride (25.06 mg), 4-(1H-1,2,4-triazol-1-yl)nicotinaldehyde (118 mg). Reaction time 16 hours. Trituration of the crude product with ethanol. Aspect of the product: yellow solid. (Yield: 32%).

(243) APCI-MS: (M+H).sup.+=391

(244) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 12.00 (s, 1H), 9.28 (s, 1H), 9.14 (s, 1H), 8.81 (d, 1H), 8.37 (s, 1H), 8.10 (s, 1H), 7.87 (s, 3H), 7.52-7.29 (m, 2H).

Example 33

(Z)-3-(4-(1H-1,2,4-triazol-1-yl)pyridin-3-yl)-2-(5-chloro-1H-indol-3-yl)acrylonitrile

(245) ##STR00064##

Method A

(246) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). THF 5 mL. Sodium hydride (28.9 mg), 4-(1H-1,2,4-triazol-1-yl)nicotinaldehyde (136 mg). Reaction time 16 hours. Purification by flash chromatography on 24 g Redisep column 20-40 m, gradient 100% CH.sub.2Cl.sub.2 to CH.sub.2Cl.sub.2/MeOH (90/10). Aspect of the product: yellow solid. (Yield: 33%).

(247) Aspect of the product: yellow solid (Yield: 33%)

(248) APCI-MS: (M+H).sup.+=347

(249) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 12.32-11.49 (m, 1H), 9.27 (s, 1H), 9.15 (d, J=4.0 Hz, 1H), 8.82 (d, J=5.4 Hz, 1H), 8.66-8.57 (m, 0.1H), 8.46-8.25 (m, 1H), 8.03-7.71 (m, 4H), 7.59-7.35 (m, 1H), 7.25 (dd, J=8.7, 2.0 Hz, 1H).

Example 34

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(furan-3-yl)pyridin-3-yl)acrylonitrile

(250) ##STR00065##

Method A

(251) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg). THF 3 mL. Sodium hydride (15.03 mg), 4-(furan-3-yl)nicotinaldehyde (93 mg). Reaction time 16 hours. Trituration of the crude product with water and NaOH. Aspect of the product: yellow solid. (Yield: 27%).

(252) APCI-MS: (M+H).sup.+=390

(253) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.98 (s, 1H), 8.92 (s, 1H), 8.64 (d, J=5.2 Hz, 1H), 8.13-8.06 (m, 2H), 7.92 (d, J=2.7 Hz, 1H), 7.87 (t, J=1.7 Hz, 1H), 7.76 (s, 1H), 7.63 (d, J=5.1 Hz, 1H), 7.48 (d, J=8.7 Hz, 1H), 7.35 (dd, J=8.7, 1.8 Hz, 1H), 6.96 (d, J=1.0 Hz, 1H).

Example 34b

(E)-2-(5-bromo-1H-indol-3-yl)-3-(4-(furan-3-yl)pyridin-3-yl)acrylonitrile

(254) ##STR00066##

Method E

(255) (Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(furan-3-yl)pyridin-3-yl)acrylonitrile (20 mg). EtOH (40 mL). Reaction time: 8 h. Aspect of the pure product: yellow solid. (Yield: 60%).

(256) ESI-MS: (M+H).sup.+=390

(257) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.37 (d, 1H), 8.19 (s, 1H), 8.0 (s, 1H), 7.72 (s, 1H), 7.60 (d, 1H), 7.45 (s, 1H), 7.38 (s, 1H), 7.30 (d, 1H), 7.19 (d, 1H), 6.98 (s, 1H), 6.89 (s, 1H).

Example 35

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile, hydrochloride

(258) ##STR00067##

Method A

(259) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (300 mg). THF 5 mL. Sodium hydride (57.8 mg), 4-methoxynicotinaldehyde (170 mg). Reaction time 16 hours. Trituration of the crude product with DCM and 4N HCl in dioxane. Aspect of the product: yellow solid. (Yield: 81%).

(260) APCI-MS: (M+H).sup.+=310

(261) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 12.18 (s, 1H), 9.12 (s, 1H), 8.86 (d, J=6.8 Hz, 1H), 7.99 (d, J=3.1 Hz, 2H), 7.71 (d, J=6.8 Hz, 1H), 7.64 (s, 1H), 7.55 (d, J=8.7 Hz, 1H), 7.27 (dd, J=8.7, 1.9 Hz, 1H), 4.16 (s, 3H).

Example 36

(Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-((2-dimethylamino)ethyl)thio)pyridin-3-yl)acrylonitrile

(262) ##STR00068##

Method D

(263) (Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-chloropyridin-3-yl)acrylonitrile (50 mg). DMF 2.0 mL. Potassium carbonate (64 mg), 2-(dimethyl)aminoethanethiol hydrochloride (25 mg). Reaction time 12 hours at 60 C. Trituration of the crude product with AcOEt. Aspect of the pure product: yellow solid. (Yield: 50%).

(264) ESI-MS: (M+H).sup.+=428

(265) .sup.1H NMR (acetone-d.sub.6, 300 MHz) ppm: 11.14 (s, 1H), 8.86 (s, 1H), 8.48 (s, 1H), 8.28 (d, 1H), 7.92 (s, 1H), 7.74 (s, 1H) 7.57 (d, 1H), 7.48 (d, 1H), 7.44 (dd, 1H), 3.33 (t, 2H), 2.70 (t, 2H), 2.28 (s, 6H).

Example 37

(Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-(4-fluorophenoxy)benzonitrile

(266) ##STR00069##

Method A

(267) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (100 mg). THF 2.0 mL. Sodium hydride (16.71 mg), 3-cyano-4-fluorophenoxy-benzaldehyde (102 mg). Reaction time 1 hour 30 minutes. Purification by flash chromatography, eluent petroleum ether/MTBE. Aspect of the pure product: yellow solid. (Yield: 48%).

(268) APCI-MS: (MH)=456

(269) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 12.02 (s, 1H), 8.39 (s, 1H), 8.05 (s, 1H), 7.93 (s, 1H), 7.86 (d, J=6.6 Hz, 1H), 7.81 (s, 1H), 7.47 (d, J=8.7 Hz, 1H), 7.32 (dd, J=15.3, 7.0 Hz, 5H), 6.95 (d, J=8.7 Hz, 1H).

Example 38

(Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(270) ##STR00070##

Method A

(271) Tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (150 mg, 0.447 mmol) THF 2 mL, NaH (25.06 mg, 0.626 mmol), 3-formyl-4-methoxybenzonitrile (88 mg, 0.537 mmol). Reaction time 1 hour 30 minutes. The reaction mixture diluted with water (20 mL). The resulting solid was filtered, washed successively with water, CH.sub.2Cl.sub.2, DIPE and acetonitrile and dried in vacuo to give 55 mg of a yellow solid (Yield: 30%).

(272) APCI-MS: (MH)=376

(273) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.99 (s, 1H), 8.22 (s, 1H), 8.08 (s, 1H), 7.91 (s, 2H), 7.70 (s, 1H), 7.60-7.15 (m, 3H), 3.97 (s, 3H).

Example 38b

(E)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(274) ##STR00071##

Method E

(275) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (20 mg). EtOH (40 mL). Reaction time: 8 h. Aspect of the pure product: yellow solid. (Yield: 60%).

(276) ESI-MS: (M+H).sup.+=379

(277) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 7.61 (d, 1H), 7.49 (s, 1H), 7.36 (s, 1H), 7.32 (d, 1H), 7.26-7.13 (m, 3H), 6.85 (d, 1H), 3.91 (s, 3H).

Example 39

(Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-(dimethylamino)benzonitrile

(278) ##STR00072##

Method A

(279) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (120 mg). THF 2.5 mL. Sodium hydride (11.55 mg), 3-cyano-4-dimethylamino-benzaldehyde (73.3 mg). Reaction time 16 hours. Silical gel flash-column chromatography (eluent heptane/ethyl acetate. Aspect of the purified product: yellow solid. (Yield: 33%).

(280) APCI-MS: (M+H).sup.+=391

(281) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.98 (s, 1H), 8.10 (d, J=1.6 Hz, 1H), 8.05 (d, J=1.7 Hz, 1H), 7.91 (s, 1H), 7.75 (dd, J=8.6, 2.1 Hz, 1H), 7.56 (s, 1H), 7.48 (d, J=8.7 Hz, 1H), 7.35 (dd, J=8.7, 1.8 Hz, 1H), 7.16 (d, J=8.7 Hz, 1H), 2.89 (s, 6H).

Example 40

(Z)-3-(2-(5-chloro-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(282) ##STR00073##

Method A

(283) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (200 mg). THF 2.0 mL. Sodium hydride (35.1 mg), 3-cyano-4-methoxybenzaldehyde (124 mg). Reaction time 1 hour 30 minutes. Silical gel flash-column chromatography (petroleum ether/DIPE) and trituration of the purified product with acetonitrile. Aspect of the pure product: yellow solid. (Yield: 40%).

(284) APCI-MS: (MH)=332

(285) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 8.21 (s, 1H), 7.93 (d, J=6.5 Hz, 3H), 7.70 (s, 1H), 7.52 (d, J=8.7 Hz, 1H), 7.29 (dd, J=22.2, 9.6 Hz, 2H), 3.96 (s, 3H).

Example 41

(Z)-3-(2-(5-chloro-1H-indol-3-yl)-2-cyanovinyl)-4-(dimethylamino)benzonitrile

(286) ##STR00074##

Method A

(287) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (200 mg). THF 2.0 mL. Sodium hydride (35.1 mg), 3-cyano-4-dimethylaminobenzaldehyde (134 mg). Reaction time 1 hour 30 minutes. Silical gel flash-column chromatography (petroleum ether/DIPE) and trituration of the purified product with acetonitrile. Aspect of the pure product: yellow solid. (Yield: 33%).

(288) APCI-MS: (M+H).sup.+=347

(289) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 11.95 (s, 1H), 8.04 (d, J=1.7 Hz, 1H), 8.00-7.89 (m, 2H), 7.75 (dd, J=8.6, 2.0 Hz, 1H), 7.62-7.45 (m, 2H), 7.30-7.09 (m, 2H), 2.89 (s, 6H).

Example 42

(Z)-3-(2-(5-chloro-1H-indol-3-yl)-2-cyanovinyl)-4-(ethylthio)benzonitrile

(290) ##STR00075##

Method A

(291) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (200 mg). THF 2.0 mL. Sodium hydride (35.1 mg), 3-cyano-4-ethylthiobenzaldehyde (145 mg). Reaction time 1 hour 30 minutes. Silical gel flash-column chromatography (petroleum ether/DIPE) and trituration of the purified product with acetonitrile. Aspect of the pure product: yellow solid. (Yield: 36%).

(292) APCI-MS: (MH)=362

(293) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm: 12.04 (s, 1H), 8.12 (s, 1H), 8.04-7.92 (m, 2H), 7.87 (dd, J=8.3, 1.7 Hz, 1H), 7.72-7.58 (m, 2H), 7.54 (d, J=8.7 Hz, 1H), 7.27 (dd, J=8.7, 1.9 Hz, 1H), 3.18 (q, J=7.3 Hz, 2H), 1.31 (t, J=7.3 Hz, 3H).

Example 43

(Z)N-(3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)pyridin-4-yl)pivalamide

(294) ##STR00076##

Method A

(295) tert-butyl 5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (300 mg). THE 6 mL. Sodium hydride (54 mg), N-(formyl-pyridin-4-yl)-2,2-dimethyl-propionamide (258 mg). Reaction time 24 hours. Silical gel flash-column chromatography (elution with cycloheptane/AcOEt: 1/1 to 1/9) and trituration with methanol. Aspect of the pure product: yellow solid. (Yield: 13%).

(296) ESI-MS: (MH)=421

(297) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 11.98 (s, 1H), 9.49 (s, 1H), 8.96 (s, 1H), 8.55 (s, 1H), 8.17 (d, 1H), 7.88 (d, 1H), 7.68 (s, 1H), 7.56 (d, 1H), 7.50 (d, 1H), 1.25 (s, 9H).

Example 44

(Z)N-(3-(2-(5-chloro-1H-indol-3-yl)-2-cyanovinyl)pyridin-4-yl)pivalamide

(298) ##STR00077##

Method A

(299) tert-butyl 5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (300 mg). THF 6 mL. Sodium hydride (62 mg), N-(formyl-pyridin-4-yl)-2,2-dimethyl-propionamide (298 mg). Reaction time 24 hours. Silical gel flash-column chromatography (elution with cycloheptane/AcOEt: 1/1 to 1/9) of the residue afforded the corresponding acrylonitrile as a yellow solid (Yield: 18%).

(300) ESI-MS: (M+H).sup.+=379

(301) .sup.1H NMR (DMSO-d.sub.6, 300 MHz) ppm: 11.98 (s, 1H), 9.49 (sl, 1H), 8.96 (s, 1H), 8.55 (s, 1H), 8.17 (s, 1H), 7.85 (s, 1H), 7.68 (s, 1H), 7.56 (m, 1H), 7.25 (d, 1H), 1.22 (s, 9H).

Example 45

(Z)-2-(6-bromo-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(302) ##STR00078##

Method F

(303) Tert-butyl 6-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (84.0 mg), NaH (10.0 mg). THF (2 mL). 10 min at rt. 4-methoxynicotinaldehyde (41.0 mg). 12 h at rt.

(304) Trituration of the crude in AcOEt. Aspect of the pure product: yellow solid. (Yield: 35%).

(305) ESI-MS: (M+H).sup.+=356

(306) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.95 (s, 1H), 8.47 (d, 1H), 7.87 (d, 1H), 7.76 (s, 1H), 7.70-7.65 (m, 2H), 7.33 (d, 1H), 7.21 (d, 1H), 4.06 (s, 3H).

Example 46

(Z)-2-(6-fluoro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(307) ##STR00079##

Method F

(308) Tert-butyl 6-fluoro-3-(cyanomethyl)-1H-indole-1-carboxylate (98.0 mg), NaH (18.0 mg). THF (3 mL). 10 min at rt. 4-methoxynicotinaldehyde (58.0 mg). 12 h at rt.

(309) Trituration of the crude in AcOEt. Aspect of the pure product: yellow solid. (Yield: 43%).

(310) ESI-MS: (M+H).sup.+=294

(311) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.30 (d, 1H), 8.00 (s, 1H), 7.55 (s, 1H), 7.41 (s, 1H), 7.38 (d, 1H), 7.16 (d, 1H), 7.10 (dd, 1H), 6.82 (d, 1H), 4.00 (s, 3H).

Example 47

(Z)-2-(6-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(312) ##STR00080##

Method F

(313) Tert-butyl 6-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (100.0 mg), NaH (18.0 mg). THF (3 mL). 10 min at rt. 4-methoxynicotinaldehyde (56.0 mg). 12 h at rt. NaOH 2.5 M (1.5 mL). 12 h at rt. Trituration of the crude in AcOEt. Aspect of the pure product: yellow solid. (Yield: 42%).

(314) ESI-MS: (M+H).sup.+=310

(315) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.95 (s, 1H), 8.48 (d, 1H), 7.92 (d, 1H), 7.76 (s, 1H), 7.71 (s, 1H), 7.50 (d, 1H), 7.24-7.18 (m, 2H), 4.06 (s, 3H).

Example 48

(Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-(furan-3-yl)pyridine-1-oxide

(316) ##STR00081##

Method G

(317) (Z)-2-(5-bromo-1H-indol-3-yl)-3-(4-(furan-3-yl)pyridin-3-yl)acrylonitrile (30.0 mg), m-CPBA (28.0+15.0 mg). THF (1.0 mL). 16 h at room temperature. Trituration of the crude in AcOEt. Aspect of the pure product: yellow solid. (Yield: 72%).

(318) ESI-MS: (M+H).sup.+=408

(319) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.83 (s, 1H), 8.37 (d, 1H), 8.03 (s, 1H), 8.01-7.92 (m, 2H), 7.82 (s, 1H), 7.79-7.72 (m, 1H), 7.66-7.56 (m, 2H), 7.54-7.33 (m, 1H), 6.86 (s, 1H).

Example 49

(Z)-3-(2-(5-chloro-1H-indol-3-yl)-2-cyanovinyl)-4-methoxypyridine 1-oxide

(320) ##STR00082##

Method G

(321) (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile (30.0 mg), m-CPBA (33.0+16.0 mg). THF (1.5 mL). 16 h at room temperature. Trituration of the crude in AcOEt. Aspect of the pure product: yellow solid. (Yield: 80%).

(322) ESI-MS: (M+H).sup.+=326

(323) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.84 (s, 1H), 8.33 (d, 1H), 7.94 (s, 1H), 7.81 (s, 1H), 7.59 (s, 1H), 7.50 (d, 1H), 7.35 (d, 1H), 7.26 (d, 1H), 4.11 (s, 3H).

Example 50

(Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-hydroxypyridin-3-yl)acrylonitrile

(324) ##STR00083##

(325) To a solution of (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile (30.0 mg, 1 eq) in NMP (0.2 mL) were added LiCl (41.0 mg, 10 eq) and p-Toluenesulfonic acid (166.0 mg, 10 eq). The resulting mixture was stirred 1 h 30 at 180 C. then cooled to room temperature and extracted with AcOEt. The combined organic layers were washed with water and dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was taken off with a minimal amount of AcOEt and filtrated to give 18.0 mg of the title compound (Yield: 95%).

(326) ESI-MS: (M+H).sup.+=297

(327) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 9.02 (s, 1H), 8.58 (d, 1H), 8.35 (s, 1H), 8.19 (s, 1H), 8.08 (d, 1H), 7.50-7.38 (m, 2H), 7.23 (d, 1H).

Example 51

(Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-hydroxybenzonitrile

(328) ##STR00084##

(329) To a solution of (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (30.0 mg, 1 eq) in NMP (0.2 mL) were added LiCl (33.0 mg, 10 eq) and p-Toluenesulfonic acid (136.0 mg, 10 eq). The resulting mixture was stirred 1 h 30 at 180 C. then cooled to room temperature and extracted with AcOEt. The combined organic layers were washed with water and dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was purified by silicagel chromatography (CH.sub.2Cl.sub.2/MeOH, 100:0 to 90:10) to give 15.0 mg of the title compound (Yield: 52%).

(330) ESI-MS: (M+H).sup.+=366

(331) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.27 (s, 1H), 8.18 (s, 1H), 7.90-7.53 (m, 2H), 7.53 (d, 1H), 7.45-7.13 (m, 3H).

Example 52

(Z)-3-(2-(5-chloro-1H-indol-3-yl)-2-cyanovinyl)-4-(trifluoromethoxy)benzonitrile

(332) ##STR00085##

Method H

(333) tert-butyl-5-chloro-3-(cyanomethyl)-1H-indole-1-carboxylate (1 eq), NaH (3 eq), 3-formyl-4-(trifluoromethoxy)benzonitrile (1 eq). Room temperature hidden from light.

(334) Aspect of the pure product: yellow solid. (Yield: 20%).

(335) ESI-MS: (M+H).sup.+=387

(336) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.49 (d, 1H), 7.92 (d, 1H), 7.83 (s, 1H), 7.68 (s, 1H), 7.82 (s, 1H), 7.58-7.42 (m, 2H), 7.28 (d, 1H).

Example 53

(Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-(trifluoromethoxy)benzonitrile

(337) ##STR00086##

Method H

(338) tert-butyl-5-bromo-3-(cyanomethyl)-1H-indole-1-carboxylate (1 eq), NaH (3 eq), 3-formyl-4-(trifluoromethoxy)benzonitrile (1 eq). Room temperature hidden from light. Aspect of the pure product: yellow solid. (Yield: 39%).

(339) ESI-MS: (M+H).sup.+=433

(340) .sup.1H NMR (methanol-d4, 300 MHz) ppm: 8.48 (s, 1H), 8.07 (s, 1H), 7.91 (d, 1H), 7.80 (s, 1H), 7.66 (s, 1H), 7.49-7.35 (m, 3H).

Example 54

(Z)-3-(2-cyano-2-(6-methoxy-1H-indol-3-yl)vinyl)-4-methoxybenzonitrile

(341) ##STR00087##

Method A

(342) tert-butyl 3-(cyanomethyl)-6-methoxy-1H-indole-1-carboxylate (175 mg), 3-formyl-4-methoxybenzonitrile (103 mg), NaH (34 mg), THF (2 ml). Reaction time 2 hours at RT. Poured in water, extracted with Ethyl acetate and trituration with diethyl ether. Aspect of the pure product: yellow solid. (Yield: 18%).

(343) APCI-MS: (M+H).sup.+=330

(344) .sup.1H NMR (300 MHz, CDCl.sub.3) ppm 11.61 (s, 1H), 8.23 (d, J=1.7 Hz, 1H), 7.93 (dd, J=8.7, 2.0 Hz, 1H), 7.79 (d, J=8.8 Hz, 1H), 7.69 (s, 2H), 7.32 (d, J=8.7 Hz, 1H), 6.98 (d, J=2.2 Hz, 1H), 6.84 (dd, J=8.8, 2.2 Hz, 1H), 3.97 (s, 3H), 3.80 (s, 3H).

(345) Some compounds of the previous examples have been the subject of tests which have demonstrated their specific relevance as inhibitors of MKlp2, and their cytotoxic effects on human cancer cells.

Preparation of Examples 55 to 79

Example 55

(Z)-2-(1-acetyl-5-bromo-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(346) ##STR00088##

(347) To a solution of (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile (50.0 mg, 1 eq) in THF (4 mL) were added pyridine (1 mL), NEt.sub.3 (0.085 mL, 4.5 eq), DMAP (8.8 mg, 0.5 eq) and acetyl chloride (0.044 mL, 3.6 eq). The resulting mixture was stirred 48 h at RT, then neutralized with saturated NH.sub.4Cl and extracted with AcOEt. The combined organic layers were washed with water and dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was taken off with a minimal amount of MeOH and filtrated to give the title compound as an orange solid (36.0 mg, 65%).

(348) ESI-MS: (M+H).sup.+=396

(349) .sup.1H NMR (300 MHz, CDCl.sub.3) ppm 9.12 (s, 1H), 8.62 (d, J=5.3 Hz, 1H), 8.47 (d, J=8.9 Hz, 1H), 8.05 (d, J=1.9 Hz, 1H), 7.84 (s, 1H), 7.81 (s, 1H), 7.58 (dd, J=8.9 Hz, 1.9 Hz, 1H), 6.98 (s, 1H), 4.04 (s, 3H), 2.73 (s, 3H).

(350) The following example was prepared as the previous method.

Example 56

(Z)-3-(2-(1-acetyl-5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(351) ##STR00089##

(352) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (30 mg), THF (2.4 ml), pyridine (0.6 mL), NEt.sub.3 (0.051 mL, 4.5 eq), DMAP (5.3 mg, 0.5 eq) and acetyl chloride (0.027 mL, 3.6 eq). Reaction time 48 hours. Extracted with AcOEt, precipitated with MeOH. Aspect of the pure product: yellow solid. (Yield: 40%).

(353) ESI-MS: (2M).sup.+=839

(354) .sup.1H NMR (300 MHz, CDCl.sub.3) ppm 8.30 (d, J=1.7 Hz, 1H), 8.08 (d, J=1.7 Hz, 1H), 7.83-7.79 (m, 1H), 7.78 (s, 1H), 7.73 (s, 1H), 7.45-7.35 (m, 2H), 7.30 (s, 1H), 7.27 (s, 1H), 4.06 (s, 3H), 2.76 (s, 3H).

Example 57

(Z)-2-(5-bromo-1-pivaloyl-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(355) ##STR00090##

(356) To a solution of (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile (30.0 mg, 1 eq) in THF (1 mL) was added NaH (6.7 mg, 2 eq), The mixture was stirred 10 min at room temperature and pivaloyl chloride (0.011 mL, 1.1 eq) was added. The resulting solution was stirred 3 h at room temperature, poured in saturated NH.sub.4Cl and extracted with CH.sub.2Cl.sub.2. The combined organic layers were dried over Na.sub.2SO.sub.4, filtrated and concentrated to give the title compound as a white solid (37.0 mg, 100%).

(357) ESI-MS: (M+H).sup.+=438

(358) .sup.1H NMR (300 MHz, CD.sub.3OD) ppm 9.20 (s, 1H), 8.82 (d, J=6.8 Hz, 1H), 8.45 (d, J=9.0 Hz, 1H), 8.35 (s, 1H), 8.13 (d, J=1.9 Hz, 1H), 7.88 (s, 1H), 7.68 (d, J=6.8 Hz, 1H), 7.60 (dd, J=9.0 Hz, 1.9 Hz, 1H), 4.30 (s, 3H), 1.57 (s, 9H).

(359) The following examples were prepared as the previous method.

Example 58

(Z)-3-(2-(5-bromo-1-pivaloyl-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(360) ##STR00091##

(361) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (500 mg), THF (16 ml), NaH 60% in oil (0.105 g, 2 eq), pivaloyl chloride (0.216 mg, 1.35 eq). Reaction time 12 hours. Poured into AcOEt. Aspect of the pure product: white solid. (Yield: 100%).

(362) ESI-MS: (M+H).sup.+=462

(363) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.39 (s, 1H), 8.36 (s, 1H), 8.27 (s, 1H), 8.12 (s, 1H), 8.01 (dd, J=8.9 Hz, 1.5 Hz, 1H), 7.97 (s, 1H), 7.61 (dd, J=8.9 Hz, 1.5 Hz, 1H), 7.38 (d, J=8.9 Hz, 1H), 3.98 (s, 3H), 1.50 (s, 9H).

Example 59

(Z)-methyl-3-(5-bromo-3-(1-cyano-2-(4-methoxypyridin-3-yl)vinyl)-1H-indol-1-yl)-3-oxopropanoate

(364) ##STR00092##

(365) (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile (100.0 mg), THF (4 mL), NaH 60% in oil (0.01 g, 1 eq), methylmalonyl chloride (0.042 g, 1.1 eq).

(366) Reaction time: 12 hours. Purification by silicagel chromatography CH.sub.2Cl.sub.2/MeOH (100:0 to 90:10). Aspect of the pure product: yellow solid. (Yield: 72%).

(367) ESI-MS: (M+H).sup.+=454

(368) .sup.1H NMR (300 MHz, CD.sub.3OD) ppm 8.95 (s, 1H), 8.49 (s, 1H), 8.07 (s, 1H), 7.76-7.60 (m, 2H), 7.48-7.29 (m, 2H), 7.22 (s, 1H), 4.07 (s, 3H), 3.96 (s, 3H), 3.85 (s, 2H).

Example 60

(Z)-5-bromo-3-(1-cyano-2-(4-methoxypyridin-3-yl)vinyl)-N,N-dimethyl-1H-indole-1-carbohydrazide

(369) ##STR00093##

(370) To a mixture of (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile (50.0 mg, 1 eq) in CH.sub.2Cl.sub.2 was added DIPEA (0.024 mL, 1 eq) and triphosgene (64.0 mg, 0.37 eq). The mixture was stirred 20 min at room temperature and a solution of dimethylhydrazine (0.011 mL, 1 eq), DIPEA (0.024 mL, 1 eq) in CH.sub.2Cl.sub.2 was added. The mixture was stirred 2 h at room temperature and concentrated. The residue was purified by silicagel chromatography (CH.sub.2Cl.sub.2/MeOH (100:0 to 90:10) to give the title compound as a yellow solid (61%).

(371) ESI-MS: (M+H).sup.+=440

(372) .sup.1H NMR (300 MHz, CD.sub.3OD) ppm 9.00 (s, 1H), 8.53 (d, J=5.8 Hz, 1H), 8.34-8.27 (m, 1H), 8.14-8.10 (m, 2H), 7.88 (s, 1H), 7.52 (d, J=8.5 Hz, 1H), 7.25 (d, J=6.0 Hz, 1H), 4.08 (s, 3H), 2.56 (s, 6H).

Example 61

(Z)-2-(5-bromo-1-(2-(dimethylamino)acetyl)-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile

(373) ##STR00094##

(374) To a mixture of (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxypyridine (60 mg) in DMF (2 ml) was added 2-(dimethylamino)acetic acid (28 mg), PyBOP (132 mg), TEA (48 l). The mixture was stirred at RT for 2 hours, poured in water and filtered. Aspect of the pure product: yellow solid. (Yield: 87%).

(375) APCI-MS: (M+H).sup.+=438

(376) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.89 (s, 1H), 8.58 (d, J=5.8 Hz, 1H), 8.38 (d, J=8.9 Hz, 1H), 8.33 (s, 1H), 8.19 (s, 1H), 7.94 (s, 1H), 7.66 (d, J=7.3 Hz, 1H), 7.25 (d, J=5.8 Hz, 1H), 4.26 (s, 2H), 3.97 (s, 3H), 2.54 (s, 6H).

Example 62

(Z)-2-(4-methylpiperazin-1-yl)ethyl 5-bromo-3-(1-cyano-2-(4-methoxypyridin-3-yl)vinyl)-1H-indole-1-carboxylate

(377) ##STR00095##

(378) To a mixture of (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile (30.0 mg, 1 eq) in CH.sub.2Cl.sub.2 (1 mL) was added NaH (7.0 mg, 1.2 eq). The mixture was stirred 10 min at room temperature and triphosgene (15.6 mg, 0.37 eq) was added. The mixture was stirred 3 h at room temperature and a solution of 2-(4-methylpiperazin-1-yl)ethanol (11.0 mg, 1 eq), DIPEA (0.024 mL, 1 eq) in CH.sub.2Cl.sub.2 was added. The mixture was stirred 2 h at room temperature and concentrated. The residue was purified by silicagel chromatography (CH.sub.2Cl.sub.2/MeOH (100:0 to 90:10) to give the title compound as a yellow solid (41%).

(379) ESI-MS: (M+H).sup.+=524

(380) .sup.1H NMR (300 MHz, CD.sub.3OD) ppm 8.95 (s, 1H), 8.48 (d, J=6.0 Hz, 1H), 8.07 (s, 1H), 7.73 (s, 1H), 7.69 (s, 1H), 7.48-7.29 (m, 2H), 7.21 (d, J=6.0 Hz, 1H), 4.07 (s, 3H), 3.69 (t, J=6.0 Hz), 2.74-2.40 (m, 6H), 2.29 (s, 4H).

Example 63

((Z)-3-(2-(5-bromo-1-(2-(dimethylamino)acetyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(381) ##STR00096##

(382) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (100 mg), DMF (2 ml), 2-(dimethylamino)acetic acid (34 mg), PyBOP (206 mg), TEA (74 l). Reaction time 2 hours. Poured in water and diisopropylether. Aspect of the pure product: yellow solid. (Yield: 82%).

(383) APCI-MS: (M+H).sup.+=462

(384) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.38 (d, J=10.8 Hz, 2H), 8.29 (s, 1H), 8.16 (s, 1H), 8.00 (d, J=10.7 Hz, 1H), 7.94 (s, 1H), 7.64 (d, J=10.6 Hz, 1H), 7.37 (d, J=8.8 Hz, 1H), 3.96 (d, J=10.2 Hz, 5H), 2.38 (s, 6H).

Example 64

(Z)-tert-butyl 5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indole-1-carboxylate

(385) ##STR00097##

(386) A mixture of (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (150 mg), Di-tert-butyldicarbonate (104 mg), DMAP (5 mg) in acetonitrile (3 ml) was stirred at RT for 0.25 h. The mixture was poured in water and filtered. Aspect of the pure product: yellow solid. (Yield: 88%).

(387) APCI-MS: (MH-Boc)=376

(388) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.26 (d, J=1.8 Hz, 1H), 8.16 (d, J=1.8 Hz, 1H), 8.11 (d, J=8.9 Hz, 1H), 8.06 (s, 1H), 8.00 (dd, J=8.7, 2.1 Hz, 1H), 7.92 (s, 1H), 7.63 (dd, J=8.9, 1.9 Hz, 1H), 7.36 (d, J=8.8 Hz, 1H), 3.97 (s, 3H), 1.65 (s, 9H).

(389) The following examples were prepared as the previous method.

Example 65

(R,Z)-benzyl-4-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-2-(tert-butoxycarbonylamino)-4-oxobutanoate

(390) ##STR00098##

(391) ((Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (150 mg), DMF (4.5 ml), (S)-4-(benzyloxy)-3-(tert-butoxycarbonylamino)-4-oxobutanoic acid (160 mg), PyBOP (310 mg), TEA (1111). Reaction time 3 hours. Poured in water and diisopropylether. Aspect of the pure product: yellow solid. (Yield: 61%).

(392) APCI-MS: (MH-Boc)=583

(393) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.37 (d, J=7.6 Hz, 2H), 8.30 (s, 1H), 8.17 (s, 1H), 8.08-7.90 (m, 2H), 7.64 (d, J=8.8 Hz, 1H), 7.46 (d, J=7.5 Hz, 1H), 7.43-7.25 (m, 6H), 5.15 (s, 2H), 4.64 (d, J=5.5 Hz, 1H), 3.98 (s, 3H), 3.60 (s, 2H), 1.36 (s, 9H).

Example 66

(R,Z)-tert-butyl-5-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-2-(tert-butoxycarbonylamino)-5-oxopentanoate

(394) ##STR00099##

(395) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (200 mg), DMF (5 ml), (S)-5-tert-butoxy-4-(tert-butoxycarbonylamino)-5-oxopentanoic acid (201 mg), PyBOP (413 mg), TEA (147 l). Reaction time 2 hours. Poured in water, extracted with AcOEt and trituration of the purified product with diisopropylether. Aspect of the pure product: yellow solid. (Yield: 53%).

(396) APCI-MS: (M+H).sup.+=507

(397) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.46-8.33 (m, 2H), 8.29 (d, J=1.8 Hz, 1H), 8.17 (d, J=1.7 Hz, 1H), 8.01 (dd, J=8.7, 2.0 Hz, 1H), 7.96 (s, 1H), 7.64 (dd, J=8.9, 1.7 Hz, 1H), 7.38 (d, J=8.8 Hz, 1H), 7.22 (d, J=7.9 Hz, 1H), 3.99 (s, 4H), 3.22 (d, J=6.1 Hz, 2H), 2.13 (m, 1H), 1.99 (m, 1H), 1.39 (d, J=9.2 Hz, 18H).

Example 67

(R,Z)-benzyl-2-amino-4-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-4-oxobutanoate

(398) ##STR00100##

(399) A mixture of (R,Z)-benzyl 4-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-2-(tert-butoxycarbonylamino)-4-oxobutanoate (116 mg), HCl-Dioxanne 4M (1.5 ml) in EtOH (2 ml) was stirred at RT for 24 hours. After concentration to dryness, the residue was triturated with water and filtered. Aspect of the pure product: yellow solid. (Yield: 62%).

(400) APCI-MS: (M+H).sup.+=583

(401) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.39 (s, 1H), 8.35-8.30 (m, 2H), 8.20 (d, J=1.8 Hz, 1H), 8.03 (dd, J=8.7, 2.1 Hz, 1H), 7.98 (s, 1H), 7.68 (dd, J=8.9, 1.9 Hz, 1H), 7.37 (dd, J=10.4, 6.3 Hz, 3H), 7.33-7.25 (m, 3H), 5.25 (s, 2H), 4.62 (t, J=4.9 Hz, 1H), 3.99 (s, 3H), 3.86 (d, J=3.9 Hz, 2H).

Example 68

(Z)-3-(2-(5-bromo-1-(2-(4-methylpiperazin-1-yl)acetyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(402) ##STR00101##

(403) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (150 mg), 2-(4-methylpiperazin-1-yl)acetic acid (78 mg), PyBOP (310 mg), Triethylamine (0.11 ml), DMF (2 ml). Reaction time 5 h at RT. Poured in water and tritured with methylene chloride. Aspect of the product: yellow solid. (Yield: 92%).

(404) APCI-MS: (M+H).sup.+=518

(405) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.43-8.32 (m, 2H), 8.30 (d, J=1.8 Hz, 1H), 8.19 (d, J=1.7 Hz, 1H), 8.02 (dd, J=8.8, 2.0 Hz, 1H), 7.95 (s, 1H), 7.65 (dd, J=8.9, 1.8 Hz, 1H), 7.38 (d, J=8.8 Hz, 1H), 4.13 (s, 2H), 3.98 (s, 3H), 2.97 (m, 8H), 2.68 (s, 3H).

Example 69

(Z)-3-(2-(5-bromo-1-(2-(4-methylpiperazin-1-yl)acetyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile hydrochloride

(406) ##STR00102##

(407) A suspension of (Z)-3-(2-(5-bromo-1-(2-(4-methylpiperazin-1-yl)acetyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (114 mg), HCl-Dioxane 4M (0.3 ml) in dioxane (1 ml) was stirred few minutes and concentrated under vacuum. Aspect of the product: yellow solid. (Yield: 95%).

(408) APCI-MS: (M+H).sup.+=518

(409) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 10.47 (s, 1H), 8.43-8.33 (m, 2H), 8.30 (d, J=1.8 Hz, 1H), 8.19 (d, J=1.8 Hz, 1H), 8.02 (dd, J=8.7, 2.1 Hz, 1H), 7.96 (s, 1H), 7.66 (dd, J=8.9, 1.9 Hz, 1H), 7.39 (d, J=8.8 Hz, 1H), 4.33 (s, 2H), 3.98 (s, 3H), 3.44 (d, J=11.5 Hz, 2H), 3.19 (s, 4H), 2.91 (s, 2H), 2.78 (s, 3H).

(410) The following examples were prepared as the previous method.

Example 70

(S,Z)-3-(2-(1-(3-aminobutanoyl)-5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile hydrochloride

(411) ##STR00103##

(412) (S,Z)-tert-butyl 4-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-4-oxobutan-2-ylcarbamate (125 mg), HCl 37% (0.092 ml), EtOH (2 ml). Reaction time 1 h at reflux. Aspect of the pure product: yellow solid. (Yield: 65%).

(413) APCI-MS: (M+H).sup.+=463

(414) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.44-8.34 (m, 2H), 8.29 (d, J=1.9 Hz, 1H), 8.19 (d, J=1.8 Hz, 1H), 8.13-7.82 (m, 5H), 7.66 (dd, J=8.9, 1.9 Hz, 1H), 7.37 (d, J=8.8 Hz, 1H), 3.97 (s, 3H), 3.76 (dd, J=13.0, 6.4 Hz, 1H), 3.53-3.42 (m, 2H), 1.34 (d, J=6.6 Hz, 3H).

Example 71

(Z)-3-(2-(1-(2-aminoacetyl)-5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile hydrochloride NC

(415) ##STR00104##

(416) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (150 mg), 2-(tert-butoxycarbonylamino)acetic acid (87 mg), PyBOP (310 mg), Triethylamine (0.11 ml), Reaction time 3 hours at RT. Then HCl 37% (0165 ml), DMF (2 ml) 2 hours at reflux. Aspect of the product: yellow solid. (Yield: 50%).

(417) APCI-MS: (M+H).sup.+=435

(418) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.60 (s, 2H), 8.38 (d, J=8.4 Hz, 2H), 8.31 (d, J=1.9 Hz, 1H), 8.22 (d, J=1.8 Hz, 1H), 8.06-7.97 (m, 2H), 7.70 (dd, J=8.9, 1.9 Hz, 1H), 7.39 (d, J=8.8 Hz, 1H), 4.68 (s, 2H), 3.98 (s, 3H).

Example 72

(Z)-3-(2-(5-bromo-1-(2-(piperazin-1-yl)acetyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile hydrochloride NC

(419) ##STR00105##

(420) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (150 mg), 2-(4-(tert-butoxycarbonyl)piperazin-1-yl)acetic acid (121 mg), PyBOP (310 mg), Triethylamine (011 ml), reaction time 3 hours at RT. Then HCl 37% (0.165 ml), DMF (2 ml) 2 hours at reflux. Aspect of the product: yellow solid. (Yield: 46%).

(421) APCI-MS: (M+H).sup.+=504

(422) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 9.36 (s, 1H), 8.44-8.33 (m, 2H), 8.31 (d, J=1.8 Hz, 1H), 8.21 (d, J=1.8 Hz, 1H), 8.03 (dd, J=8.7, 2.1 Hz, 1H), 7.96 (s, 1H), 7.68 (dd, J=8.9, 1.8 Hz, 1H), 7.39 (d, J=8.8 Hz, 1H), 4.62 (s, 2H), 3.98 (s, 3H), 3.26 (d, J=17.6 Hz, 8H).

Example 73

(Z)-3-(2-(5-bromo-1-(2-(2-(2-methoxyethoxy)ethoxy)acetyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(423) ##STR00106##

(424) (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (150 mg), 2-(2-(2-methoxyethoxy)ethoxy)acetic acid (88 mg), PyBOP (310 mg), Triethylamine (0.11 ml), DMF (2 ml). Reaction time 18 hours at RT. Poured in water and washed with acetonitrile. Aspect of the product: yellow solid. (Yield: 33%).

(425) APCI-MS: (M+H).sup.+=538

(426) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.39 (d, J=8.9 Hz, 1H), 8.33-8.23 (m, 2H), 8.18 (d, J=1.7 Hz, 1H), 8.01 (dd, J=8.7, 2.0 Hz, 1H), 7.94 (s, 1H), 7.65 (dd, J=8.9, 1.8 Hz, 1H), 7.38 (d, J=8.8 Hz, 1H), 4.93 (s, 2H), 3.98 (s, 3H), 3.81-3.68 (m, 2H), 3.60 (d, J=5.0 Hz, 2H), 3.55-3.48 (m, 2H), 3.39 (d, J=5.3 Hz, 2H), 3.19 (s, 3H).

Example 74

(S,Z)-3-(2-(1-(2-amino-3-hydroxypropanoyl)-5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile hydrochloride

(427) ##STR00107##

(428) (S,Z)-tert-butyl 1-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-3-hydroxy-1-oxopropan-2-ylcarbamate (220 mg), HCl 37% (110 l), EtOH (2 ml). Reaction time 1 h at reflux. Aspect of the product: yellow solid. (Yield: 61%).

(429) APCI-MS: (M+H).sup.+=465

(430) .sup.1H NMR (300 MHz, DMSO-d.sub.6) ppm 8.68 (d, J=19.5 Hz, 3H), 8.38 (d, J=8.9 Hz, 1H), 8.31 (d, J=1.9 Hz, 1H), 8.20 (d, J=1.8 Hz, 1H), 8.01 (dd, J=9.6, 2.9 Hz, 2H), 7.70 (dd, J=8.9, 1.9 Hz, 1H), 7.38 (d, J=8.8 Hz, 1H), 5.69 (s, 1H), 5.24 (s, 1H), 3.96 (d, J=5.0 Hz, 5H).

Example 75

(Z)-3-(2-(5-bromo-1-(5-oxopyrrolidine-2-carbonyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile

(431) ##STR00108##

(432) To a mixture of (Z)-3-(2-(5-bromo-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (100 mg) in DMF (2 ml) was added 5-oxopyrrolidine-2-carboxylic acid (37 mg), BOP (128 mg), TEA (581). The mixture was stirred at RT for 3 hours, poured in water and filtered. Aspect of the pure product: yellow solid. (Yield: 60%).

(433) ESI+MS: (M+H).sup.+=490

(434) .sup.1H NMR (300 MHz, MeOD-d.sub.6) ppm 8.44 (s, 1H), 8.22 (s, 1H), 8.08 (s, 1H), 7.91 (dd, J=8.8, 2.0 Hz, 1H), 7.95 (s, 1H), 7.65 (dd, J=14.0, 2.1 Hz, 2H), 7.71 (s, 1H), 7.40-7.31 (m, 2H), 4.21 (t, J=6.0 Hz, 1H), 3.98 (s, 3H), 2.73-2.13 (m, 4H).

Example 76

(R,Z)-3-(2-(5-bromo-1-(2,6-diaminohexanoyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile dihydrochloride

(435) ##STR00109##

(436) (R,Z)-tert-butyl-6-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-6-oxohexane-1,5-diyl dicarbamate (320 mg), HCl 37% (164 l), EtOH (3 ml). Reaction time 1 hour at reflux. Aspect of the pure product: pale yellow solid. (Yield: 67%).

(437) APCI-MS: (M+H).sup.+=506

(438) .sup.1H NMR (300 MHz, CD.sub.3OD) ppm 8.68 (s, 3H), 8.40 (d, J=8.9 Hz, 1H), 8.32 (d, J=1.7 Hz, 1H), 8.22 (d, J=1.8 Hz, 1H), 8.07-8.00 (m, 2H), 7.91 (s, 2H), 7.71 (dd, J=8.9, 1.8 Hz, 1H), 7.40 (d, J=8.8 Hz, 1H), 5.19 (s, 1H), 3.99 (s, 3H), 2.72 (s, 2H), 1.96 (s, 2H), 1.46 (m, 4H).

Example 77

(Z)-diethyl (2-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-2-oxoethyl)phosphonate

(439) ##STR00110##

(440) To a solution of (Z)-2-(5-chloro-1H-indol-3-yl)-3-(4-methoxypyridin-3-yl)acrylonitrile (0.382 g, 1 eq) in THF (6 mL) was added NaH 60% in mineral oil (0.048 g, 1.2 eq). The resulting solution was stirred 30 min at room temperature and a solution of diethyl (2-chloro-2-oxoethyl)phosphonate (0.216 g, 1 eq) in THF (3 mL). The mixture was stirred overnight, quenched with saturated NH.sub.4Cl, extracted with AcOEt, dried over Na.sub.2SO.sub.4, filtrated and concentrated. The residue was purified by silicagel chromatography CH.sub.2Cl.sub.2/MeOH (100:0 to 90:10) to give the title compound as yellow solid (36%).

(441) ESI-MS: (M+H).sup.+=556

(442) .sup.1H NMR (300 MHz, CD.sub.3OD) ppm 8.28 (s, 1H), 8.07 (s, 1H), 7.81 (dd, J=8.5 Hz, 1.8 Hz, 1H), 7.43 (d, J=8.9 Hz, 1H), 7.36 (d, J=8.9 Hz, 1H), 7.29 (d, J=8.5 Hz, 1H), 4.2 (q, J=7.0 Hz, 4H), 4.06 (s, 3H), 3.10 (d, J=20.0 Hz, 2H), 1.36 (t, J=7.0 Hz, 6H).

Example 78

(Z)-2-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-2-oxoethyl diphosphate, tetrabutylammonium salt

(443) ##STR00111##

(444) To a solution of (Z)-3-(2-(5-bromo-1-(2-bromoacetyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (0.036 g, 1 eq) in AcCN (1 mL) was added Tributylammonium pyrophosphate (0.092 g, 1.3 eq). The reaction mixture was stirred 48 h at room temperature and concentrated to give the title compound as a yellow solid (60%).

(445) ESI-MS: (MH).sup.+=590

(446) .sup.1H NMR (300 MHz, CD.sub.3OD) ppm 8.24 (d, J=1.9 Hz, 1H), 8.00 (s, 1H), 7.75 (dd, J=8.7 Hz, 2.1 Hz, 1H), 7.71-7.67 (m, 2H), 7.50 (d, J=8.7 Hz, 1H), 7.35 (s, 1H), 7.31 (dd, J=8.7 Hz, 1.7 Hz, 1H), 7.25 (d, J=8.7 Hz, 1H), 4.97 (d, J=20 Hz, 2H), 4.03 (s, 3H), 4.06 (s, 3H), 3.27-3.22 (m, 24H), 1.72-1.62 (m, 24H), 1.49-1.37 (m, 24H), 1.03 (t, J=7.2 Hz, 36H).

Example 79

(Z)-3-(5-bromo-3-(1-cyano-2-(5-cyano-2-methoxyphenyl)vinyl)-1H-indol-1-yl)-3-oxopropyl dihydrogen phosphate

(447) ##STR00112##

(448) To a solution of (Z)-3-(2-(5-bromo-1-(3-hydroxypropanoyl)-1H-indol-3-yl)-2-cyanovinyl)-4-methoxybenzonitrile (0.047 g, 1 eq) in THF (3.1 mL) and AcCN (3.6 mL) was added DIPEA (90 L). The reaction mixture was cooled to 0 C. and POCl.sub.3 (0.078 mL, 8 eq) was added dropwise. The resulting solution was stirred 3 h at 0 C. and 1 M KH.sub.2PO.sub.4 (PH=4) (5 mL) was added dropwise. The mixture was stirred overnight and concentrated. The residue was purified by C18 flash chromatography H.sub.2O/MeOH (100:0 to 0:100) to give the title compound as yellow solid (40%).

(449) .sup.1H NMR (300 MHz, CD.sub.3OD) ppm 8.30 (s, 1H), 8.04 (s, 1H), 8.00-7.97 (m, 2H), 7.83-7.80 (m, 2H), 7.40 (dd, J=9.0 Hz, 1.5 Hz, 1H), 7.29 (d, J=8.7 Hz, 1H), 4.06 (s, 3H), 3.73 (m, 2H), 3.23 (m, 2H).

(450) Evaluation of Inhibitory Effects on the Microtubule Stimulated ATPase Activity of the MKlp2 Motor Domain.

(451) Material and Methods

(452) MKlp2 ATPase activity was measured by monitoring real time free phosphate generation using the Kinesin ELIPA Assay Kit. The assay is based upon an absorbance shift (330 nm-360 nm) that occurs when 2-amino-6-mercapto-7-methylpurine ribonucleoside (MESG) is catalytically converted to 2-amino-6-mercapto-7-methylpurine in the presence of inorganic phosphate (Pi). One molecule of Pi will yield one molecule of 2-amino-6-mercapto-7-methylpurine in an essentially irreversible reaction. Hence, the absorbance at 360 nm is directly proportional to the amount of Pi generated in the kinesin ATPase reaction.

(453) Human recombinant MKlp2 motor domain.sub.1-519, His tagged (Cytoskeleton, Cat. #MP05), plus porcine brain microtubules (Cytoskeleton, cat. #MT002) were used.

(454) All experiments were performed at 22 C.

(455) Condition 1, compound preparation.

(456) The compounds were dissolved in DMSO at 30 the maximum concentration to be tested. Each compound had a seven point dose-response evaluation, with final concentrations 100, 33, 11, 3.7, 1.2, 0.4, 0.13 M. DMSO solutions were pipetted directly into each well.

(457) Condition 2, reaction's motor mix.

(458) The following were mixed sequentially in the specified order at RT to obtain the motor mix.

(459) 20 mL: 15 mM Pipes-NaOH pH 7.0, 10 mM MgCl.sub.2, 30 M Tx (Buffer 1).

(460) 10 mL: 5MSEG (ELIPA 1 reagent, Cat. #BK051).

(461) 10 mL: 2.5 mg/mL porcine brain microtubules (310 mg Cat. #MT002-XL resuspended in 12 mL of Buffer 1).

(462) 0.25 mL: 500 g/mL MKlp2 protein.

(463) 0.5 mL: 100PNP (ELIPA 2 reagent, cat. #BK051).

(464) Condition 3, reaction initiation.

(465) The motor mix was pipetted into each well to obtain 80% of total volume. The reaction was initiated by adding a 20% of total volume of 5 mM ATP into each well.

(466) The reactions were measured in a SpectraMax M2 (Molecular Devices) set in kinetic mode and 360 nm absorbance wavelength. The start protocol was 5 second rapid circular mixing, 21 readings, 30 seconds apart.

(467) IC50 values were determined as the concentration to inhibit 50% of the MKlp2 ATPase activity.

(468) Evaluation of Cytotoxicity Effects on Human Cancer Cells

(469) Material and Methods

(470) The effects of the compounds of the invention on the viability of human cancer cells were studied on various human cancer cell lines of differing tissue origins (A549, NCI-H460: lung cancer; MDA-MB-231: breast cancer; HCT-116, HT-29: colon cancer; MIA-PaCa-2: pancreatic cancer; K562: leukaemia). All cell lines were obtained from ATCC or ECACC.

(471) Cells were cultured in the culture media described below, under a 37 C., 5% CO.sub.2 humidified atmosphere, according to a standard operating procedure.

(472) Organ Cell Line Culture Medium

(473) A549: RPMI 1640+10% FBS+2 mM sodium pyruvate

(474) NCI-H460: RPMI 1640+10% FBS+10 mM HEPES+1 mM Sodium pyruvate+2.5 g/l glucose

(475) HCT-116: Mc Coy's 5a+10% FBS+0.5 mM Ultraglutamine

(476) HT-29: Mc Coy's 5a+10% FBS+0.5 mM Ultraglutamine

(477) MDA-MB-231: Ham's F12+10% FBS

(478) K562: RPMI 1640+10% FBS+2 mM ultraglutamine

(479) MIA-PaCa-2: DMEM+10% FBS

(480) On D0, the cells were plated in 90 l in 96 wells plates at densities ranging from 500 to 5,000 cells per well.

(481) On D1, the cells were treated as described below: the compounds of invention were diluted in DMSO in order to obtain a concentration of 5 mM. This solution was serially diluted in PBS+10% FBS in order to obtain the concentrations of 500.000, 166.667, 55.555, 18.518, 6.173, 2.058, 0.686, 0.229, 0.076 and 0.025 M. The addition of 10 l in each well allowed the testing concentrations of 50.000, 16.6667, 5.5555, 1.8518, 0.6173, 0.2058, 0.0686, 0.0229, 0.0076 and 0.0025 M.

(482) Following addition of the test substance the cells were protected from light. The solvents (DMSO and specific control solvent were added at the maximal concentration: 10 l/well (3 wells/condition)).

(483) On D4, the Cell Proliferation Reagent WST-1 was added to each well (10 l/well), according to a standard operating procedure. The cells were then incubated for 30 min to 4 h at 37 C.-5% CO.sub.2. After these incubations, the 96-well plates were shaken thoroughly for 1 min with Multiskan EX apparatus (Thermo Labsystems, France). The absorbence was measured at 450 nm, the reference wavelength being 620 nm. The analysis of the results was performed with the Ascent software 2.6 (Thermo Labsystems, France), Microsoft Excel 2003 and GraphPad Prism 4.03 softwares to give the concentration of the compounds that induces the death of 50% of the cells (IC50).

(484) The results for some compounds considered in above-cited examples in term of inhibition of microtubule stimulated ATPase activity of MKlp2 are illustrated in Tables 3 and 6 hereafter.

(485) The results for some compounds considered in above-cited examples in term of cytotoxicity on K562 cells are illustrated in Table 4 and 6 hereafter.

(486) The results for some compounds considered in above-cited examples in term of cytotoxicity on other human cancer cells are illustrated in Table 5 hereafter.

(487) TABLE-US-00003 TABLE 3 MKlp2 X R1 R1 R2 R3 IC50 (M) W02010/150211-Example 1 N H H H H 3.8 W02010/150211-Example 4 N OCH.sub.3 H H H 4.2 W02010/150211-Example 22 N OCH.sub.2CH.sub.3 H H H 5.2 W02010/150211-Example 23 N OCH(CH.sub.3).sub.2 H H H 5.2 W02010/150211-Example 24 N Cl H H H 1.1 W02010/150211-Example 28 N OCH.sub.3 H H F 2.4 W02010/150211-Example 31 N OCH.sub.3 H CH.sub.3 H 2.3 W02010/150211-Example 47 N OCH.sub.3 H Cl H 1.6 W02010/150211-Example 37 N Br H H H 1.5 W02010/150211-Example 26 N H OCH.sub.3 H H 26.5 W02010/150211-Example 52 N OCH.sub.3 H H OCH.sub.3 12.6 W02010/150211-Example 30 CCN OCH.sub.3 H H H 4.4 Example 2 N OCH.sub.3 H OCH.sub.2OH.sub.3 H 0.3 Example 3 N Cl H Cl H 0.8 Example 4 N Br H Cl H 0.7 Example 5 N Br H OCH.sub.3 H 0.05 Example 5b N Br H OCH.sub.3 H 0.61 Example 6 N Cl H N(CH.sub.3).sub.2 H 0.5 Example 7 N Cl H N(CH.sub.3).sub.2 H 0.8 Example 8 N Br H N(CH3).sub.2 H 0.9 Example 9 N Cl H OCH.sub.3 H 0.1 Example 10 N Cl H OC.sub.6H.sub.5 H 0.3 Example 11 N Br H OC.sub.6H.sub.5 H 0.5 Example 12 N OCH.sub.3 H OCH.sub.3 H 0.3 Example 13 N Br H OCH.sub.2CH.sub.3 H 0.1 Example 14 N Br H OCH(CH.sub.3).sub.2 H 0.2 Example 15 N Br H SCH.sub.3 H 0.2 Example 16 N Br H SCH.sub.2CH.sub.3 H 0.3 Example 17 N Br H (C.sub.6H.sub.4)3-Br H 0.3 Example 18 N Cl H (C.sub.6H.sub.4)3-Br H 0.3 Example 19 N Br H SC.sub.6H.sub.5 H 0.3 Example 20 N Br H SCH.sub.2C.sub.6H.sub.5 H 0.2 Example 21 N Br H SC.sub.6H.sub.5-3,4-(OCH.sub.3).sub.2 H 0.4 Example 22 N Br H OC.sub.6H.sub.54-F H 0.09 Example 23 N Cl H OC.sub.6H.sub.54-F H 0.09 Example 24 N Br H N(CH.sub.2CH.sub.3).sub.2 H 1.1 Example 25 N Br H C.sub.6H.sub.54-CF.sub.3 H 0.4 Example 26 N Cl H C.sub.6H.sub.54-CF.sub.3 H 0.5 Example 27 N Cl H SC.sub.6H.sub.54-F H 0.3 Example 28 N Br H SC.sub.6H.sub.54-F H 0.09 Example 29 N Cl H C.sub.4H.sub.3O H 0.03 Example 30 N Cl H SC.sub.5H.sub.4N H 0.3 Example 31 N Br H SC.sub.5H.sub.4N H 0.4 Example 32 N Br H C.sub.2H.sub.2N.sub.3 H 0.06 Example 33 N Cl H C.sub.2H.sub.2N.sub.3 H 0.1 Example 34 N Br H C.sub.4H.sub.3O H 0.07 Example 34b N Br H C.sub.4H.sub.3O H 0.96 Example 35 N Cl H OCH.sub.3 H <0.07 Example 36 N Br H S(CH.sub.2).sub.2N(CH.sub.3).sub.2 H 0.3 Example 37 CCN Br H OC.sub.6H.sub.54-F H 0.9 Example 38 CCN Br H OCH.sub.3 H 0.2 Example 39 CCN Br H N(CH.sub.3).sub.2 H 0.2 Example 40 CCN Cl H OCH.sub.3 H 0.2 Example 41 CCN Cl H N(CH.sub.3).sub.2 H 0.4 Example 42 CCN Cl H SCH.sub.2CH.sub.3 H 0.6 Example 45 N H Br OCH.sub.3 H 0.2 Example 46 N H F OCH.sub.3 H 0.12 Example 47 N H Cl OCH.sub.3 H 0.2 Example 48 N.sup.+O.sup. Br H C.sub.4H.sub.3O H 0.73 Example 49 N.sup.+O.sup. Cl H OCH.sub.3 H 0.56 Example 54 CCN H OCH.sub.3 OCH.sub.3 H 1.4 Example 59: Prodrug with Ra = N Br H OCH.sub.3 H 0.57 COCH.sub.2CO.sub.2CH.sub.3 Example 62: Prodrug with Ra = N Br H OCH.sub.3 H 0.35 CO.sub.2(CH.sub.2).sub.2-piperazinyl-CH.sub.3

(488) TABLE-US-00004 TABLE 4 K562 X R1 R1 R2 R3 IC50 (M) W02010/150211-Example 1 N H H H H 21.4 Example 3 N Cl H Cl H 3.6 Example 4 N Br H Cl H 1.4 Example 5 N Br H OCH.sub.3 H 0.8 Example 5b N Br H OCH.sub.3 H 3.92 Example 34b N Br H C.sub.4H.sub.3O H 14.26 Example 8 N Br H N(CH.sub.3).sub.2 H 7.5 Example 9 N Cl H OCH.sub.3 H 1.4 Example 12 N OCH.sub.3 H OCH.sub.3 H 0.9 Example 15 N Br H SCH.sub.3 H 2.9 Example 24 N Br H N(CH.sub.2CH.sub.3).sub.2 H 3.1 Example 29 N Cl H C.sub.4H.sub.3O H 4.7 Example 34 N Br H C.sub.4H.sub.3O H 2.2 Example 38 CCN Br H OCH.sub.3 H 0.03 Example 39 CCN Br H N(CH.sub.3).sub.2 H 0.6 Example 48 N.sup.+O.sup. Br H C.sub.4H.sub.3O H 6.71 Example 49 N.sup.+O.sup. Cl H OCH.sub.3 H 9.23 Example 50 N Cl H OH H 14.72 Example 51 CCN Br H OH H 0.5 Example 52 CCN Cl H OCF.sub.3 H 0.85 Example 53 CCN Br H OCF.sub.3 H 0.38 Example 54 CCN H OCH.sub.3 OCH.sub.3 H 0.02

(489) TABLE-US-00005 TABLE 5 X R1 R1 R2 R3 IC50 (M) Example 5 N Br H O H HCT-116: 0.7 CH.sub.3 MDA-MB-231: 1.7 MIA-PaCa-2: 0.7 NCIH460: 0.9 Example 38 CCN Br H O H HCT-116: 0.06 CH.sub.3 MDA-MB-231: 0.07 MIA-PaCa-2: 0.04 NCIH460: 0.04 Example 39 CCN Br H N H A549: 0.5 (CH.sub.3).sub.2 HT-29: 0.3 MDA-MB-231: 0.3 MIA-PaCa-2: 0.2

(490) TABLE-US-00006 TABLE 6 MKlp2 K562 X R1 R1 R2 R3 Z/E Ra IC50 (M) IC50 (M) Example 55 N Br H OCH.sub.3 H Z COCH.sub.3 13.7 0.55 Example 56 CCN Br H OCH.sub.3 H Z COCH.sub.3 16.7 0.25 Example 57 N Br H OCH.sub.3 H Z COC(CH.sub.3).sub.3 20 0.2 Example 58 CCN Br H OCH.sub.3 H Z COC(CH.sub.3).sub.3 7.5 0.06 Example 61 N Br H OCH.sub.3 H Z COCH.sub.2N(CH.sub.3).sub.2 0.4 0.33 Example 63 CCN Br H OCH.sub.3 H Z COCH.sub.2N(CH.sub.3).sub.2 10.6 0.04 Example 64 CCN Br H OCH.sub.3 H Z CO.sub.2C(CH.sub.3).sub.3 19.6 0.24 Example 65 CCN Br H OCH.sub.3 H Z COCH.sub.2CH(NHBoc)Cbz 10.1 0.04 Example 66 CCN Br H OCH.sub.3 H Z CO(CH.sub.2).sub.2CH(Boc)-NHBoc 5.4 0.03 Example 67 CCN Br H OCH.sub.3 H Z COCH.sub.2CH(NH.sub.2)Cbz 3.1 0.04 Example 68 CCN Br H OCH.sub.3 H Z COCH.sub.2-piperazinyl-CH.sub.3 >50 0.03 Example 69 CCN Br H OCH.sub.3 H Z COCH.sub.2-piperazinyl-CH.sub.3HCl >50 0.03 Example 70 CCN Br H OCH.sub.3 H Z COCH.sub.2CH(CH.sub.3)NH.sub.2 >50 0.01 Example 71 CCN Br H OCH.sub.3 H Z COCH.sub.2NH.sub.2HCl 1.9 0.01 Example 72 CCN Br H OCH.sub.3 H Z COCH.sub.2-piperazinylHCl >50 0.01 Example 73 CCN Br H OCH.sub.3 H Z COCH.sub.2O(CH.sub.2).sub.2O(CH.sub.2).sub.2OCH.sub.3 >50 0.03 Example 74 CCN Br H OCH.sub.3 H Z COCH(NH.sub.2)CH.sub.2OH 2.1 0.01 Example 75 CCN Br H OCH.sub.3 H Z CO-oxopyrrolidine 4.6 0.02 Exemple 76 CCN Br H OCH.sub.3 H Z COCH(NH.sub.2)(CH.sub.2)4NH.sub.2 1.3 0.04 Exemple 79 CCN Br H OCH.sub.3 H Z CO(CH.sub.2).sub.2PO.sub.4H.sub.2 >50 0.4
Stability Studies in Mouse or Human Plasma

(491) The study was to evaluate the stability of the disclosed compounds after incubation in mouse or human plasma and to measure the metabolites formed. For the disclosed compound, a stock solution was prepared at 200 M in DMSO. This solution was then 100-fold diluted in 1 ml of mouse or human plasma in order to obtain the required concentration of 2 M. One aliquot of 100 l was taken (T0) and the remaining solution was incubated at 37 C. in water bath for 60 min, 120 min and 240 min.

(492) At the end of each incubation time 100 l of plasma was taken, 100 l of acetonitrile containing 0.1% of formic acid were added to each aliquot in order to stop the enzymatic reaction and to precipitate the proteins. Samples was vortexed/mixed and centrifuged 5 min at 16434.6 g (=14000 rpm) (4 C.). After centrifugation, the clear supernatant (at least 150 l) was transferred into 1.2 ml HPLC glass vials and sealed. Samples were placed into the refrigerated autosampler and 20 l were injected into a HPLC-MS/MS.

(493) The results are expressed with the percentage of test substance remaining by comparing area under specific chromatographic peak of test samples after incubation with area under specific chromatographic peak at T0 (Tables 7 and 8 and FIGS. 1 and 2).

(494) TABLE-US-00007 TABLE 7 Remaining compound 63 (%) Incubation (mean SEM, n = 2) time (min) Mouse plasma Human plasma 0 100.0 0.0 100.0 0.0 60 9.0 0.8 49.2 5.1 120 10.7 8.7 27.2 5.4 240 4.9 0.5 21.2 11.2 Half-life time: ~33 min ~60 min

(495) TABLE-US-00008 TABLE 8 Compound 38 formed from compound 63 (arbitrary units) Incubation (mean SEM, n = 2) time (min) Mouse plasma Human plasma 0 0.6 0.1 0.5 0.0 60 3.2 0.1 2.5 0.4 120 3.7 0.4 2.6 0.1 240 2.9 0.2 3.4 0.8
In-Vivo Evaluation of Anti-Tumor Activity of Compound 38 in Nude Mice Bearing Subcutaneous Human Colon Carcinoma HCT-116 Xenografts
Protocol:

(496) The effects of one compound of the invention were studied on the tumor growth of human cancer cells in nude mice. The human colon carcinoma HCT-116 cell line was obtained from ATCC. The induction in nude mice was realized by subcutaneous injection in the right flank of each mouse of 1010.sup.6 HCT-116 cells in 200 l serum-free medium. When the tumor volume reached 130 mm.sup.3, mice were randomized in to 3 groups (10 mice/group).

(497) Mice of group 1 were treated by intraperitoneal injection of vehicle (solutol HS15 at 38% in NaCl 0.9%) according to the treatment schedule 1Q2Dx3 for 1 week (from D0 to D7) and then 1Q1Dx21 (from D8 to D27).

(498) Mice of group 2 were treated by intraperitoneal injection of cisplatin (diluted in NaCl 0.9%) at 4 mg/kg according to the treatment schedule 1Q3Dx3.

(499) Mice of group 3 were treated by intraperitoneal injection of compound 38 (diluted in solutol HS15 at 38% in NaCl 0.9%) at 37.5 mg/kg according to the treatment schedule 1Q2Dx3 for 1 week (from D0 to D7) and then 1Q1Dx21 (from D8 to D27).

(500) The body weight and tumor volume of mice were recorded twice a week until the end of the experiment. The results are illustrated in Table 9 and FIG. 3.

(501) TABLE-US-00009 TABLE 9 Mean body weight change (MBWC) of mice of each group. At the beginning of the treatment, the mean body weight (MBW) was 21 g. MBWC MBWC MBWC MBWC MBWC MBWC MBWC MBWC Treatment D0-D4 D0-D7 D0-D10 D0-D14 D0-D17 D0-D21 D0-D24 D0-D28 vehicle 0.48 g +0.21 g 0.07 g 0.18 g +0.68 g +0.62 g +1.04 g +1.48 g Cisplatin 1.1 g 2.54 g 3.33 g 2.22 g 1.26 g 0.38 g 0.02 g 0.1 g 4 mg/kg Cpd 38 1.04 g 0.76 g 1.08 g 1.57 g 1.58 g 1.43 g 1.83 g 2.54 g 37.5 mg/kg
Results:

(502) Antitumor activity was observed in HCT-116 xenograft bearing nude mice, treated with cisplatin at 4 mg/kg, validating the sensitivity of the tumor model to antitumora antitumor agents (FIG. 3).

(503) Antitumor activity was observed in HCT-116 xenograft bearing nude mice and treated with compound 38 at 37.5 mg/kg (FIG. 3).

(504) Compound 38 treatment was well tolerated in nude mice bearing HCT-116 xenograft (table 9).

(505) A moderate loss of body weight was observed during treatment (Table 9 ranging from 4% to 12%).

(506) In-Vivo Evaluation of Anti-Tumor Activity of Compound 38 in Nude Mice Bearing Subcutaneous Human Large Cell Lung Carcinoma NCI-H460 Xenografts

(507) Protocol:

(508) The effects of one compound of the invention were studied on the tumor growth of human cancer cells in nude mice. The human large lung carcinoma NCI-H460 cell line was obtained from ATCC. The induction in nude mice was realized by subcutaneous injection in the right flank of each mouse with 510.sup.6 NCI-H460 cells in 200 l serum-free medium. When the tumor volume reached 80 mm.sup.3, mice were randomized in 2 groups (10 mice/group).

(509) Mice of group 1 were treated by intraperitoneal injection of vehicle (solutol HS15 at 38% in NaCl 0.9%) according to the treatment schedule 1Q1Dx17.

(510) Mice of group 2 were treated by intraperitoneal injection of compound 38 (diluted in solutol HS15 at 38% in NaCl 0.9%) at 37.5 mg/kg according to the treatment schedule 1Q1Dx17.

(511) The tumor volume was recorded twice a week until the end of the experiment.

(512) TABLE-US-00010 TABLE 10 Mean body weight change (MBWC) of mice of each group. At the beginning of the treatment, the mean body weight (MBW) was 21 g. MBWC MBWC MBWC MBWC MBWC MBWC Treatment D0-D3 D0-D4 D0-D7 D0-D10 D0-D14 D0-D17 vehicle +0.33 g +0.16 g +1.3 g +1.54 g +2.48 g +3.44 g Cpd 38 0.96 g 1.76 g 1.82 g 2.12 g 1.83 g 1.55 g 37.5 mg/kg
Results:

(513) Antitumor activity was observed on NCI-H460 xenograft bearing nude mice, treated with compound 38 at 37.5 mg/kg (FIG. 4).

(514) Compound 38 treatment was well tolerated in nude mice bearing NCI-H460 xenograft (table 10).

(515) A moderate loss of body weight was observed during treatment (Table 9 ranging from 5% to 10%).