Pharmaceutical compounds

Abstract

The invention provides the use of a compound for the manufacture of a medicament for the treatment of pain, wherein the compound is a compound of the formula (VI): ##STR00001##
or a salt, solvate, tautomer or N-oxide thereof;
wherein the bicyclic group: ##STR00002##
is selected from the structures C1, C5 and C6: ##STR00003##
wherein n, R.sup.1, R.sup.2a, R.sup.3, R.sup.4a, R.sup.8 and R.sup.10 are as defined in the claims. The invention also provides the use of a compound of the formula (VI) for the manufacture of a medicament for the prophylaxis or treatment of a fungal, protozoal, viral or parasitic disease state or condition (other than a disease state or condition due to Plasmodium falciparum) or for use in the prophylaxis or treatment of Ewing's sarcoma, atherosclerosis or lupus erythematosus.

Claims

1. A method for treating a pain in a subject in need thereof, which comprises administrating to said subject a therapeutically effective amount of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]methanone or a salt thereof wherein said pain is nociceptive pain.

2. The method according to claim 1, wherein the salt is a L-lactate salt of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]methanone.

Description

EXAMPLES

(1) The invention will now be illustrated, but not limited, by reference to the specific embodiments described in the following examples.

(2) In the examples, the following abbreviations may be used.

(3) AcOH acetic acid

(4) BOC tert-butyloxycarbonyl

(5) Bn benzyl

(6) CDI 1,1-carbonyldiimidazole

(7) DMAW90 Solvent mixture: DCM: MeOH, AcOH, H.sub.2O (90:18:3:2)

(8) DMAW120 Solvent mixture: DCM: MeOH, AcOH, H.sub.2O (120:18:3:2)

(9) DMAW240 Solvent mixture: DCM: MeOH, AcOH, H.sub.2O (240:20:3:2)

(10) DCM dichloromethane

(11) DMF dimethylformamide

(12) DMSO dimethyl sulphoxide

(13) EDC 1-ethyl-3-(3′-dimethylaminopropyl)-carbodiimide

(14) Et.sub.3N triethylamine

(15) EtOAc ethyl acetate

(16) Et.sub.2O diethyl ether

(17) h hour(s)

(18) HOAt 1-hydroxyazabenzotriazole

(19) HOBt 1-hydroxybenzotriazole

(20) MeCN acetonitrile

(21) MeOH methanol

(22) min. minutes

(23) P.E. petroleum ether

(24) r.t. room temperature

(25) SiO.sub.2 silica

(26) TBTU N,N,N′,N′-tetramethyl-O-(benzotriazol-1-yl)uronium tetrafluoroborate

(27) THF tetrahydrofuran

(28) Proton magnetic resonance (.sup.1H NMR) spectra were recorded on a Bruker AV400 instrument operating at 400.13 MHz, in DMSO-dB or MeOH-d.sub.4 (as indicated) at 27° C., unless otherwise stated and are reported as follows: chemical shift δ/ppm (number of protons, multiplicity where s=singlet, d=doublet, t=triplet, q=quartet, m=multiplet, br=broad). The residual protic solvent was used as the internal reference.

(29) In the examples, the compounds prepared were characterised by liquid chromatography and mass spectroscopy using the system and operating conditions set out below. Where atoms with different isotopes are present and a single mass quoted, the mass quoted for the compound is the monoisotopic mass (i.e. .sup.35Cl; .sup.79Br etc.). Different systems were used, as described below, and these were equipped with, and were set up to run under, closely similar operating conditions. The operating conditions used are also described below.

(30) System Description:

(31) System 1 (Analytical System):

(32) HPLC System: Waters 2795

(33) Mass Spec Detector: Micromass Platform LC

(34) PDA Detector: Waters 2996 PDA

(35) System 2 (Preparative and Analytical System):

(36) HPLC System: Waters Fractionlynx system

(37) Mass Spec Detector: Waters ZQ

(38) PDA Detector: Waters 2996 PDA

(39) System 3 (Preparative and Analytical System):

(40) HPLC System: Agilent 1100 system

(41) Mass Spec Detector: LC/MSD

(42) UV Detector: Agilent MWD

(43) Operating Conditions:

(44) Acidic Analytical Conditions:

(45) Eluent A: H.sub.2O (0.1% Formic Acid)

(46) Eluent B: CH.sub.3CN (0.1% Formic Acid)

(47) Gradient: 5-95% eluent B over 3.5 minutes (over 15 minutes w/column 2)

(48) Flow: 0.8 ml/min

(49) Column 1: Phenomenex Synergi 4μ MAX-RP 80A, 2.0×50 mm

(50) Column 2: Phenomenex Synergi 4μ MAX-RP 80A, 2.0×150 mm

(51) Basic Analytical Conditions:

(52) Eluent A: H.sub.2O (10 mM NH.sub.4HCO.sub.3 buffer adjusted to pH=9.2 with NH.sub.4OH)

(53) Eluent B: CH.sub.3CN

(54) Gradient: 5-95% eluent B over 3.5 minutes

(55) Flow: 0.8 ml/min

(56) Column: Phenomenex Gemini 5μ 2.0×50 mm

(57) MS conditions (Waters Systems):

(58) Capillary voltage: 3.6 kV (3.40 kV on ES negative)

(59) Cone voltage: 25 V

(60) Source Temperature: 120° C.

(61) Scan Range: 125-800 amu

(62) Ionisation Mode: ElectroSpray Positive, Negative or Positive & Negative

(63) MS Conditions (Agilent Systems):

(64) Capillary voltage: 4000 V (3500 V on ES Negative)

(65) Fragmentor/Gain: 150/1

(66) Drying gas Temp/flow: 350° C./13.0 Lmin.sup.−1

(67) Nebuliser pressure: 50 psig

(68) Scan Range: 125-800 amu

(69) Ionisation Mode: ElectroSpray Positive or Negative

(70) The starting materials for each of the Examples are commercially available unless otherwise specified.

A. General Synthetic Methods

(71) In the following general methods, the volumes stated may vary according to the scale of the reaction, as will be apparent to the skilled person.

(72) Method A1

(73) Amide Coupling (Acid Chloride Method)

(74) A mixture of a carboxylic acid (1 equivalent) and thionyl chloride (1.5 equivalents) in benzene (or toluene) was stirred and held at reflux for 2 hours. Excess amine was added dropwise to the hot solution and the mixture stirred at room temperature for 15 minutes. Alternatively, the acid chloride could be isolated by evaporation and then re-dissolved in a 9:1 mixture of dichloromethane: triethylamine and the amine then added and the mixture stirred under nitrogen at room temperature for 1-18 hours. In either case, the mixture was diluted with ethyl acetate and extracted successively with water, saturated aqueous sodium bicarbonate and 2M hydrochloric acid. The organic layer was reduced to dryness in vacuo and the pure products were obtained either by trituration with ethyl acetate or by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether) or in a few cases by preparative HPLC/MS.

(75) Method A2

(76) Amide Coupling (EDC, HOBt Method)

(77) A stirred solution of the acid (1 equivalent) in dichloromethane (10 ml) was treated successively with N-ethyl-N′-(3-dimethylaminopropyl)carbodiimide hydrochloride (1.2 equivalents), 1-hydroxybenzotriazole (1.2 equivalents) and the amine (1.5 equivalents) and the mixture was stirred at room temperature overnight. The mixture was washed successively with 2M hydrochloric acid and 2M sodium hydroxide, the organic layer was separated and the solvent removed in vacuo to afford the products. The products were either obtained pure or were purified by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether or methanol in ethyl acetate as appropriate).

(78) Method A3

(79) Anisole or benzyl ether dealkylation (BBr.sub.3 Method)

(80) A stirred solution of the anisole or benzyl ether (1 equivalent) in dichloromethane at 0° C. was treated dropwise with a 1M solution of boron tribromide in dichloromethane (1.5 equivalents per group to be deprotected) and the mixture was stirred for 2 hours. The reaction was quenched by the addition of water and saturated aqueous sodium bicarbonate, the organic layer was separated and the solvent was removed in vacuo. The pure products were obtained either by trituration with diethyl ether or ethyl acetate or by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether).

(81) Method A4

(82) Amide Coupling (EDC, HOAt Method)

(83) A stirred solution of the acid (1 equivalent) in dimethylformamide (5 ml) was treated successively with N-ethyl-N′-(3-dimethylaminopropyl)carbodiimide hydrochloride (1.2 equivalents), 1-hydroxy-7-aza-benzotriazole (1.2 equivalents) and the amine (1.5 equivalents) and the mixture was stirred at room temperature overnight. DMF was evaporated and crude dissolved in EtOAc and was washed successively with saturated sodium bicarbonate, the organic layer was separated and the solvent removed in vacuo. The products were either obtained pure or were purified by column chromatography on silica (eluting with mixtures of ethyl acetate in petroleum ether or methanol in ethyl acetate as appropriate).

(84) Method A5

(85) Hydrogenation

(86) A stirred solution of protected derivative (1 equivalent) and a catalytic amount of 10% palladium on carbon (typically 30-50 mg) in ethanol (5-10 ml), methanol (5-10 ml) or methanol/DCM (3 ml/3 ml) was stirred at room temperature under an atmosphere of hydrogen for 2-16 hours. The catalyst was removed by filtration, washed with methanol (5 ml) and the solvent removed in vacuo to afford the products. Some required purification by flash chromatography, eluting typically with ether.

(87) Method A6

(88) Suzuki Coupling

(89) The aryl bromide (1 equivalent, typically 0.5 mmol), boronic acid or potassium vinyl trifluoroborate derivative (1.2 equivalents) and caesium carbonate (3 equivalents) were dissolved in THF (10 ml) under nitrogen. 1,1′-Bis(diphenylphosphino)ferrocene] palladium(II) dichloride (0.1 equivalent) was added and then water (1 ml). The mixture begins to darken until black. The mixture was then heated at reflux under nitrogen until the reaction is complete (845 hrs). The mixture was cooled, diluted with DCM and magnesium sulphate added. The mixture was filtered and the solvent evaporated. The resulting residues were purified by flash chromatography in pet. ether/ether mixtures, and generally gave product in good yield (˜60-80%).

(90) Method A7

(91) Resorcinol Mono-O-Methylation

(92) Dimethyl sulphate (1 equivalent) was added to a stirred solution of the resorcinol (1 equivalent) and potassium carbonate (2.2 equivalents) in acetonitrile (10 ml per mmol of substrate) and the mixture was stirred at room temperature for 16 hours. The solvent was removed in vacuo, the residue partitioned between dichloromethane and water, the organic layer separated and the solvent removed in vacuo. The pure products were obtained either after column chromatography on silica (eluting with mixtures of petroleum ether and ethyl acetate) or by preparative HPLC/MS.

(93) Method A8

(94) Electrophilic Aromatic Fluorination

(95) 1-(Chloromethyl)-4-fluoro-1,4-diazoniabicyclo[2.2.2]octane bis(tetrafluoroborate) (1 equivalent) was added to a solution of the substrate (1 equivalent) in acetonitrile (15 ml per mmol of substrate) and the mixture was stirred at room temperature for 16 hours. The solvent was removed in vacuo and the residue was partitioned between ethyl acetate and water. The organic layer was separated and reduced to dryness in vacuo. The pure products were obtained either after column chromatography on silica (eluting with mixtures of petroleum ether and ethyl acetate) or by preparative HPLC/MS.

B. Synthesis of Carboxylic Acid Intermediates

Preparation B1

4-Hydroxy-3-isopropylbenzoic acid

(96) ##STR00127##

(97) Carbon tetrachloride (28 ml, 0.26 mol) and copper powder (1.0 g) were added to a stirred solution of 2-isopropylphenol (27.2 g, 0.2 mol) in 50% aqueous sodium hydroxide (120 ml) and the mixture was held at reflux for 16 hours. Upon cooling the mixture was acidified to pH 2 or below by the addition of concentrated hydrochloric acid and was extracted with ethyl acetate. The organic layer was extracted with a saturated aqueous solution of sodium bicarbonate and the aqueous layer acidified to pH 2 or below by the very careful addition of concentrated hydrochloric acid. The solution was extracted with ethyl acetate, the organic layer was washed with water, separated and the solvent removed in vacuo to afford 4-hydroxy-3-isopropylbenzoic acid (12.5 g, 35%) as a bright red solid that was used without further purification. .sup.1H NMR (DMSO-d.sub.6) 12.36 (1H, br s), 10.13 (1H, br s), 7.73 (1H, d), 7.63 (1H, dd), 6.85 (1H, d), 3.22 (1H, m), 1.19 (6H, d). MS: [M−H].sup.+ 179.

(98) Alternatively, if required, the crude product may be purified using a three step procedure involving di-benzylation [according to the conditions outlined below in Preparation B5 for the synthesis of methyl 5-acetyl-2,4-bis-benzyloxybenzoate (BnBr, K.sub.2CO.sub.3, MeCN, reflux)], column chromatography on silica to remove highly coloured impurities (eluting with 3-5% ethyl acetate in petroleum ether) and catalytic hydrogenation [according to Method A5 outlined above (10% Pd/C, EtOH, H.sub.2)] to afford 4-hydroxy-3-isopropylbenzoic acid as a colourless solid.

Preparation B2

5-Ethyl-2-methoxybenzoic acid

(99) ##STR00128##

(100) n-Butyl lithium (2.5M in hexanes, 38.5 ml, 100.0 mmol) was added dropwise under a nitrogen atmosphere to a stirred solution of 4-ethylanisole (11.7 g, 86.0 mmol) and N,N,N′,N′-tetramethylethylenediamine (10 ml, 88.0 mmol) in anhydrous diethyl ether (100 ml) and the mixture was stirred and held at 30° C. for 16 hours. The mixture was cooled and poured slowly in to a mixture of excess solid carbon dioxide in anhydrous diethyl ether. Upon warming to room temperature the mixture was made basic by the addition of 2M sodium hydroxide, the aqueous layer was separated and acidified to pH 2 or below by the addition of concentrated hydrochloric acid. The mixture was extracted with diethyl ether, the organic layer separated and the solvent removed in vacuo to afford 5-ethyl-2-methoxybenzoic acid (5.7 g, 37%) as a pale yellow oil. .sup.1H NMR (DMSO-d.sub.6) 12.50 (1H, br s), 7.48 (1H, d), 7.33 (1H, dd), 7.03 (1H, d), 2.56 (2H, q), 1.17 (3H, q). MS: [M+H].sup.+ 181.

Preparation B3

2,4-Bis-benzyloxy-5-chloro-benzoic acid

(101) ##STR00129##

(102) 1-(2,4-Bis-benzyloxy-5-chloro-phenyl)-ethanone [prepared as per WO 2004/0500087] (1.10 g, 3.0 mmol) was added to a stirred solution of sodium hydroxide (1.20 g, 30.0 mmol) in water (10 ml) and dioxane (10 ml). Bromine (1.44 g, 9.0 mmol) was added dropwise and the mixture stirred at room temperature for 3 hours. The dioxane was removed by evaporation in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid. The mixture was extracted with ethyl acetate, the organic layer separated and the solvent removed in vacuo to afford 2,4-bis-benzyloxy-5-chloro-benzoic acid (900 mg, 81%) as a pale yellow solid. .sup.1H NMR (DMSO-d.sub.6) 12.58 (1H, br s), 7.77 (1H, s), 7.55-7.30 (10H, m), 7.11 (1H, s), 5.31 (2H, s), 5.27 (2H, s). MS: [M+H].sup.+ 369.

Preparation B4

3-(1,2-Dimethyl-allyl)-4-hydroxy-benzoic acid

(103) ##STR00130##

(104) Ethyl 4-hydroxybenzoate (1.66 g, 10.0 mmol) and anhydrous potassium carbonate (2.07 g, 15.0 mmol) in acetonitrile (30 ml) was treated with 3-methyl-2-butenyl chloride (1.35 ml, 12.0 mmol) and the mixture was stirred and held at reflux for 3 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between dichloromethane and water. The organics were separated and the solvent removed in vacuo to afford ethyl 4-(3-methyl-but-2-enyloxy)-benzoate (2.23 g, 95%) as a pale yellow liquid which was used without further purification. .sup.1H NMR (DMSO-d.sub.6) 7.89 (2H, d), 7.04 (2H, d), 5.44 (1H, t), 4.62 (2H, d), 4.28 (2H, q), 1.77 (3H, s), 1.73 (3H, s), 1.31 (3H, t). MS: [M+H].sup.+ 235.

(105) Ethyl 4-(3-methyl-but-2-enyloxy)-benzoate (2.23 g, 9.53 mmol) was dissolved in anisole (8 ml) and the mixture stirred and held at reflux for 4 days. The solvent was removed in vacuo and the residue subjected to column chromatography on silica. Elution with 20% ethyl acetate in petroleum ether afforded ethyl 3-(1,2-dimethyl-allyl)-4-hydroxy-benzoate (600 mg, 27%) as a Colourless solid. .sup.1H NMR (DMSO-d.sub.6) 10.32 (1H, br s), 7.67 (1H, dd), 7.62 (1H, s), 6.90 (1H, d), 4.90 (1H, s), 4.85 (1H, s), 4.25 (2H, q), 3.75 (1H, q), 1.61 (3H, s), 1.30 (3H, t), 1.26 (3H, d). MS: [M+H].sup.+ 235.

(106) Ethyl 3-(1,2-dimethyl-allyl)-4-hydroxy-benzoate (600 mg, 2.56 mmol) was dissolved in methanol (20 ml), a solution of potassium hydroxide (560 mg, 10.0 mmol) in water (10 ml) was added and the mixture was stirred and held at reflux for 16 hours. Upon cooling the methanol was removed in vacuo and the solution acidified to pH 2 or below by the addition of 2M hydrochloric acid. The solution was extracted with dichloromethane, the organic layer was separated and the solvent was removed in vacuo to afford 3-(1,2-dimethyl-allyl)-4-hydroxy-benzoic acid (270 mg, 51%) as a colourless gum. .sup.1H NMR (DMSO-d.sub.6) 12.38 (1H, br s), 10.22 (1H, br s), 7.63 (2H, m), 6.88 (1H, d), 4.90 (1H, s), 4.87 (1H, s), 3.75 (1H, q), 1.60 (3H, s), 1.28 (3H, d). MS: [M−H].sup.+ 205.

Preparation B5

2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid

(107) ##STR00131##

(108) Acetic anhydride (3.06 g, 30.0 mmol) was added to methyl 2,4-dihydroxybenzoate (5.04 g, 30.0 mmol) in boron trifluoride diethyl etherate (7.6 ml) and the mixture was stirred and held at reflux for 3 hours and then allowed to cool to room temperature. Water (80 ml) was added and the mixture stirred at room temperature for 30 minutes. The resulting yellow solid was removed by filtration and sucked as dry as possible under vacuum. The solid was dissolved in dichloromethane and was washed with water, the organic layer was separated and the solvent removed in vacuo to afford methyl 5-acetyl-2,4-dihydroxybenzoate as a bright yellow solid (2.62 g, 42%) which was used without further purification. .sup.1H NMR (DMSO-d.sub.6) 12.58 (1H, s), 11.22 (1H, s), 8.33 (1H, s), 6.45 (1H, s), 3.90 (3H, s), 2.62 (3H, s). MS: [M+H].sup.+ 211.

(109) Methyl 5-acetyl-2,4-dihydroxybenzoate (2.62 g, 12.48 mmol) was dissolved in acetonitrile (40 ml), anhydrous potassium carbonate (4.93 g, 35.7 mmol) was added and the stirred mixture was treated with benzyl bromide (5.09 g, 29.75 mmol) and held at reflux for 3 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between water and dichloromethane. The organic layer was separated and the solvent removed in vacuo to afford methyl 5-acetyl-2,4-bis-benzyloxybenzoate (3.48 g, 71%) as a colourless solid which was dried at 50° C. in a vacuum oven and used without further purification. .sup.1H NMR (DMSO-d.sub.6) 8.21 (1H, s), 7.55 (4H, m), 7.43 (4H, m), 7.37 (2H, m), 7.04 (1H, s), 5.38 (4H, s), 3.79 (3H, s), 2.48 (3H, s). MS: [M+H].sup.+ 391.

(110) A stirred suspension of methyltriphenylphosphonium bromide (1.96 g, 5.5 mmol) in anhydrous tetrahydrofuran (20 ml) at 0° C. under a nitrogen atmosphere was treated dropwise with n-butyl lithium (1.6 M in hexanes, 3.5 ml, 5.5 mmol) and the resulting bright yellow solution was stirred at 0° C. for 30 minutes. A solution of methyl 5-acetyl-2,4-bis-benzyloxybenzoate (1.95 g, 5.00 mmol) in anhydrous tetrahydrofuran (20 ml) was added dropwise and the resulting mixture was allowed to warm to room temperature and was stirred for 16 hours. Methanol (10 ml) was added and the solvent was removed in vacuo. The residues were partitioned between dichloromethane and water, the organic layer was separated and the solvent removed in vacuo to afford a brown gum that was purified by column chromatography on silica. Elution with 7% ethyl acetate in petroleum ether afforded methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate as a colourless solid (700 mg, 36%). .sup.1H NMR (DMSO-d.sub.6) 7.59 (1H, s), 7.52 (2H, d), 7.64-7.32 (8H, m), 6.97 (1H, s), 5.28 (2H, s), 5.22 (2H, s), 5.09 (1H, s), 5.04 (1H, s), 3.76 (3H, s), 2.02 (3H, s). MS: [M+H].sup.+ 389.

(111) Methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate (700 mg, 1.80 mmol) was dissolved in methanol (20 ml), a solution of potassium hydroxide (286 mg, 5.1 mmol) in water (4 ml) was added and the mixture was stirred and held at reflux for 3 hours. Upon cooling the solvent was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid. The mixture was extracted with dichloromethane, the organic layer was separated and the solvent removed in vacuo to afford 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (600 mg, 89%) as a colourless solid. .sup.1H NMR (DMSO-d.sub.6) 7.52 (2H, d), 7.47-7.29 (9H, m), 6.82 (1H, s), 5.20 (2H, s), 5.17 (2H, s), 5.06 (1H, s), 5.04 (1H, s), 2.03 (3H, s). MS: [M+H].sup.+ 375.

Preparation B6

2,4-Bis-benzyloxy-5-bromo-benzoic acid

(112) ##STR00132##

(113) 2,4-dihydroxy-5-bromobenzoic acid (5.16 g, 22.15 mmol) was dissolved in DMF (40 ml) and potassium carbonate (12.2 g) and benzyl bromide (8 ml) were sequentially added. The mixture was stirred at room temperature for 18 hours under nitrogen. An aqueous solution of potassium hydroxide (2 g) in water (25 ml) was then added, followed by methanol (50 ml) and the mixture heated to reflux with vigorous stirring for 24 hours. The mixture was then allowed to cool, was poured into 1N HCl (250 ml) and was then extracted with ether and then DCM. The combined organic layers were dried over magnesium sulphate and the solvent evaporated in vacuo. The resulting solid material was washed with P.E. and then Et.sub.2O (3×50 ml) to yield pure product (5.2 g, 56%). .sup.1H NMR (MeOH-d.sub.4) 8.06 (1H, s), 7.51-7.30 (10H, m), 6.85 (1H, s), 5.22 (2H, s), 5.20 (2H, s). MS: [M+H].sup.+ 413.

Preparation B7

Synthesis of (Z)-4-benzyloxy-3-(1-methyl-propenyl)-benzoic acid

(114) ##STR00133##

(115) Methyl 3-bromo-4-hydroxybenzoate [prepared as per Tetrahedron, 2003, 59, 9173] (3.47 g, 15.0 mmol) was dissolved in acetonitrile (50 ml), anhydrous potassium carbonate (3.11 g, 22.5 mmol) was added and the stirred mixture was treated with benzyl bromide (3.08 g, 18.0 mmol) and held at reflux for 5 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between water and dichloromethane. The organic layer was separated, the solvent removed in vacuo and the residue subjected to column chromatography on silica. Elution with 10% ethyl acetate in petroleum ether afforded methyl 4-benzyloxy-3-bromobenzoate (3.6 g, 75%) as a colourless solid. .sup.1H NMR (DMSO-d.sub.6) 8.12 (1H, d), 7.96 (1H, dd), 7.51 (2H, m), 7.43 (2H, t), 7.35 (2H, m), 5.32 (2H, s), 3.84 (3H, s).

(116) Methyl 4-benzyloxy-3-bromobenzoate (1.61 g, 5.0 mmol), caesium carbonate (4.89 g, 15.0 mmol), (E)-2-buten-2-yl boronic acid (600 mg, 6.0 mmol) and [1,1′-bis(diphenylphosphino)ferrocenyl]palladium (II) chloride (204 mg, 0.25 mmol) were dissolved in anhydrous tetrahydrofuran (100 ml), water (10 ml) was added and the mixture was stirred and held at reflux under an atmosphere of nitrogen for 16 hours. Upon cooling the solvent was removed in vacuo and the mixture partitioned between dichloromethane and water. The organic layer was separated, the solvent removed in vacuo and the residue subjected to column chromatography on silica. Elution with 5% ethyl acetate in petroleum ether afforded methyl (Z)-4-benzyloxy-3-(1-methyl-propenyl)-benzoate (600 mg, 41%) as a colourless solid. .sup.1H NMR (DMSO-d.sub.6) 7.88 (1H, dd), 7.59 (1H, d), 7.40 (4H, m), 7.34 (1H, m), 7.23 (1H, d), 5.57 (1H, q), 5.21 (2H, s), 3.82 (3H, s), 1.94 (3H, s), 1.38 (3H, d).

(117) Methyl (Z)-4-benzyloxy-3-(1-methyl-propenyl)-benzoate (592 mg, 2.0 mmol) was dissolved in methanol (20 ml), a solution of potassium hydroxide (336 mg, 6.0 mmol) in water (7 ml) was added and the mixture was stirred and held at reflux for 3 hours. Upon cooling the solvent was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid. The mixture was extracted with dichloromethane, the organic layer was separated and the solvent removed in vacuo to afford (Z)-4-benzyloxy-3-(1-methyl-propenyl)-benzoic acid (460 mg, 82%) as a colourless solid. .sup.1H NMR (DMSO-d.sub.6) 7.85 (1H, dd), 7.57 (1H, d), 7.40 (4H, m), 7.34 (1H, m), 7.18 (1H, d), 5.57 (1H, q), 5.21 (2H, s), 1.96 (3H, s), 1.40 (3H, d). MS: [M+H].sup.+ 283.

Preparation B8

Synthesis of 2,4-bis-benzyloxy-5-tert-butyl-benzoic acid

(118) ##STR00134##

(119) 1-(2,4-Bis-benzyloxy-5-tert-butyl-phenyl)-ethanone [prepared as per WO 2004/072051] (2.02 g, 5.2 mmol) was dissolved in 1,4-dioxane (30 ml), a solution of sodium hydroxide (2.08 g, 52.0 mmol) in water (30 ml) was added and the mixture was stirred and treated dropwise with bromine (0.8 ml, 15.6 mmol). The resulting mixture was stirred at room temperature for 16 hours. The 1,4-dioxane was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid. The mixture was extracted with ethyl acetate, the organic layer was separated, the solvent removed in vacuo and the residue subjected to column chromatography on silica. Elution with 30% ethyl acetate in petroleum ether afforded 2,4-bis-benzyloxy-5-tert-butyl-benzoic acid (1.6 g, 79%) as a pale yellow oil. .sup.1H NMR (DMSO-d) 12.18 (1H, br s), 7.69 (1H, s), 7.52 (4H, t), 7.45-7.33 (6H, m), 6.93 (1H, s), 5.24 (2H, s), 5.23 (2H, s), 1.32 (9H, s). MS: [M+H].sup.+ 391.

Preparation B9

Synthesis of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid

Alternative Synthesis

(120) ##STR00135##

Step 1: Synthesis of 2,4-bis-benzyloxy-5-bromo-benzoic acid benzyl ester

(121) ##STR00136##

(122) To a 10 L jacketed vessel, fitted with a flange lid containing stirrer, thermometer and dropping funnel, was charged acetone (2.5 L) followed by 5-bromo-2,4-dihydroxybenzoic acid (100 g, 0.43 mol) and potassium carbonate (356 g, 2.58 mol). To the stirring mixture at ambient was added benzyl bromide (185 mL, 1.55 mol) at a rate of −20 ml/min. The mixture was heated at 60° C. for 18 h and then taken to 45° C. Water (1.5 L) was added and the mixture stirred for 30 min. The mixture was extracted with EtOAc (2×1 L) and the combined organic portions reduced in vacuo. To the residue was added Et.sub.2O (200 mL) and petroleum ether (1 L), the mixture stirred for 30 min and the solid formed collected by filtration and dried in vacuo to give the title compound (197.2 g) as a white solid.

Step 2: Synthesis of Potassium Isopropenyl Trifluoroborate

(123) ##STR00137##

(124) To a solution of 2-bromopropene (20 mL, 225 mmol) in an hydrous THF (250 mL) stirring under a N.sub.2 atmosphere at −78° C. was added over 30 mins n-BuLi (2.5M in hexanes) (100 mL, 250 mmol) and the mixture stirred for 30 mins. To the mixture at −78° C. was slowly added triethyl borate (58 mL, 340 mmol) at a rate to ensure that the temperature of the reaction mixture did not exceed −65° C. The resulting solution was then stirred at −78° C. for 30 mins, allowed to slowly warm to ambient and stirred for a further 90 mins. Potassium hydrogen fluoride (105 g, 1.35 mol) was added to the mixture followed by water (250 mL). The mixture was stirred at ambient for 14 h and then reduced to dryness.

(125) The procedure was repeated as above and following reduction to dryness the two residues were combined for further work-up.

(126) To the combined residues was added acetone (800 mL), the mixture stirred for 1 h and then filtered. The solid collected was washed with acetone (200 mL) and the combined filtrates reduced in vacuo to give a solid. This solid was triturated with Et.sub.2O (250 mL) and then dried in vacuo to give the title compound (28.2 g) as a white solid.

Step 3: Synthesis of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid benzyl ester

(127) ##STR00138##

(128) To a mixture of 2,4-bis-benzyloxy-5-bromo-benzoic acid benzyl ester (42.9 g, 85.7 mmol), potassium isopropenyl trifluoroborate (14.0 g, 95.2 mmol) and caesium carbonate (83.8 g, 257.1 mmol) in THF (800 mL) was added Pd(PPh.sub.3).sub.4 (2.0 g) followed by water (150 mL). The mixture was heated at reflux for 72 h then allowed to cool to ambient. The mixture was reduced in vacuo to remove THF and then partitioned between water (500 mL) and EtOAc (300 mL). The organic portion was washed with brine, dried (MgSO.sub.4), filtered and reduced in vacuo to give the title compound (40.9 g) as a brown oil.

Step 3A

Alternative Synthesis of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid benzyl ester

(129) ##STR00139##

(130) A mixture of 2,4-bis-benzyloxy-5-bromo-benzoic acid benzyl ester (10.0 g, 20 mmol), potassium isopropenyl trifluoroborate (4.0 g, 27.2 mmol) and n-butylamine (6.0 mL, 60 mmol) in 2-propanol/water (2:1, 200 mL) was purged with N.sub.2 for 5 minutes. To this mixture was added [1,1′-bis(diphenyl-phosphino)ferrocene]dichloropalladium(II) (816 mg, 1.09 mmol) and the mixture was heated at reflux for 20 h. The mixture was allowed to cool to ambient then diluted with water (400 mL) and extracted with EtOAc (2×300 mL). The combined organic extracts were washed with 1M aqueous HCl, brine, dried (MgSO.sub.4), filtered through a plug of Celite and the filtrate reduced in vacuo to give the title compound (11.1 g) as a brown gum.

Step 4: Synthesis of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid

(131) ##STR00140##

(132) To a solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid benzyl ester (40.8 g, 87.9 mmol) in THF-MeOH-water (3:1:1, 300 mL total) was added lithium hydroxide (8.42 g, 352 mmol). The mixture was heated at 50° C. for 16 h, allowed to cool to ambient and then diluted with water (300 mL). The mixture was taken to pH˜1 using conc. HCl (˜30 mL) and then extracted with EtOAc (2×200 mL). The combined organic extracts were washed with brine, dried (MgSO.sub.4), filtered and reduced in vacuo. The solid residue was taken up in P.E-MeOH (9:1, 300 mL total), the slurry stirred for 1 h at ambient and the solid collected by filtration. The solid was dried in vacuo to give the title compound (26.8 g) as an off-white solid. .sup.1H NMR (400 MHz, DMSO-d.sub.6) 12.30 (s, 1H), 7.61 (s, 1H), 7.53 (d, J=7.0 Hz, 2H), 7.47-7.31 (m, 8H), 6.94 (s, 1H), 5.23 (d, J=14.0 Hz, 4H), 5.08 (d, J=9.0 Hz, 2H), 2.04 (s, 3H).

Preparation B10

2,4-Bis-benzyloxy-5-isopropyl-benzoic acid

Step 1

Preparation of 1-(2-4-Bis-benzyloxy-phenyl)-ethanone

(133) ##STR00141##
Material Inputs:

(134) TABLE-US-00003 S. No. Item Quantity Eq. 1. 1,3 Dihydroxy acetophenone 50 g 1 2. Benzyl bromide 97 ml 3 3. Acetonitrile 750 ml 15 times 4. Potassium carbonate 115 g 3

(135) 1,3 Dihydroxy acetophenone (50 g) was placed in a 2 L single neck RB flask equipped with a reflux condenser and a guard tube. Acetonitrile (750 ml), potassium carbonate (115 g) and benzyl bromide (97 ml) were added and the mixture was heated at reflux (90° C.) for 16 hours. On completion, the acetonitrile was removed under reduced pressure. Water (200 ml) was added to the reaction mixture which was then extracted with ethyl acetate (500 ml). The organic layer was separated and dried over sodium sulphate. The solvent was removed under reduced pressure to give a residue which was washed n-hexane (600 ml) to give the product.

(136) TABLE-US-00004 Quantity of the product obtained: 105.1 g Yield 96.24% Nature Solid Colour Brown

Step 2

Preparation of 24-Bis-benzyloxy-1-isopropenylbenzene

(137) ##STR00142##
Material Inputs:

(138) TABLE-US-00005 S. No. Item Quantity Eq. 1. Compound of Step 1 20 g 1 2. n-BuLi (1.6 M) 92.6 ml 2.3 3. Methyl-triphenylphosphonium iodide 53.4 g 2.2 4. THF 200 ml 10 times

(139) Methyl-triphenylphosphonium iodide (53.4 g) and THF (100 ml) were introduced into a 1 L 3-neck RB flask equipped with an addition funnel and an inlet for nitrogen atmosphere and the mixture was cooled to 0° C. n-BuLi (92.6 ml) was added dropwise to the reaction mixture over a period of 15 min at 0° C. The reaction mixture was stirred for 10 min at 0° C. and further stirred at RT for 30 min. 1-(2-4-Bis-benzyloxy-phenyl)-ethanone (20 g) in THF (100 ml) was added dropwise to the reaction mixture over a period of 10 min at 0° C. and the reaction mixture was stirred overnight at RT. The progress of the reaction was monitored by TLC (10% EtOAc/n-hexane, product R.sub.f˜0.9). On completion, methanol (˜100 ml) was added to the reaction mixture and the solvent was removed under reduced pressure to give a residue. n-Hexane (1 L) was added to the residue which was refluxed (75° C.) for 30 min. before filtering the mixture was through a Celite bed and washing the bed with n-hexane (500 ml). The solvent was removed under reduced pressure to give a residue, which was further purified by column chromatography (SiO.sub.2 2% EtOAc/n-hexane).

(140) TABLE-US-00006 Quantity of the product obtained: 12.5 g Yield 63.13% Nature Liquid. Colour Colorless

Step 3

4-Isopropyl-benzene-1,3-diol

(141) ##STR00143##
Material Inputs:

(142) TABLE-US-00007 S. No. Item Quantity Eq. 1. 2-4-Bis-benzyloxy-1- 12.5 g 1 isopropenylbenzene 2. Ethanol 125 ml 10 times 3. 20% Palladium hydroxide 2 g

(143) To a mixture of 2-4-bis-benzyloxy-1-isopropenylbenzene (12.5 g) in ethanol (125 ml) in a 500 ml hydrogenation flask was added 20% palladium hydroxide (2 g). The reaction mixture was hydrogenated at 80 psi for 36 h. The progress of the reaction was monitored by TLC (10% EtOAc/n-hexane, product R.sub.f˜0.1). On completion, the reaction mixture was filtered through a bed of Celite and the bed was washed with ethanol (300 ml). The solvent was removed under reduced pressure to give a crude product, which was used as such for the next step.

(144) TABLE-US-00008 Quantity of the product obtained 5.8 g (crude) Nature Solid. Colour Colourless.

Step 4

1-(2,4-Dihydroxy-5-isopropyl-phenyl)-ethanone

(145) ##STR00144##
Material Inputs:

(146) TABLE-US-00009 S. No. Item Quantity Eq. 1. 4-Isopropyl-benzene-1,3-diol 5.8 g 1 2. Boron trifluoride etherate 28.7 ml 6 3. Acetic acid 4.55 ml 2

(147) 4-isopropyl-benzene-1,3-diol (5.8 g) and boron trifluoride etherate (28.7 ml) were introduced into a 250 ml single neck RB flask equipped with a reflux condenser and an inlet for nitrogen atmosphere stirred at RT for 10 min. Acetic acid (4.55 ml) was added to the reaction mixture and stirred at 90° C. for 16 h. On completion, 10% sodium acetate (300 ml) was added to the reaction mixture which was stirred at RT for 4 hours. The reaction mixture was then extracted with ethyl acetate (300 ml) and washed with saturated sodium bicarbonate (100 ml) and the organic layer was dried over sodium sulphate. The reaction was monitored by TLC (10% EtOAc/n-hexane, product R.sub.f˜0.5). The solvent was removed under reduced pressure to give a residue, which was further purified by column chromatography (SiO.sub.2, 10% EtOAc/n-hexane).

(148) TABLE-US-00010 Quantity of the product obtained 3.2 g Yield 43.24% Nature Solid. Colour Colourless

Step 5

1-(2,4-Bis-benzyloxy-5-isopropyl-phenyl)-ethanone

(149) ##STR00145##
Material Inputs:

(150) TABLE-US-00011 S. No. Item Quantity Eq. 1. 1-(2,4-Dihydroxy-5-isopropyl-phenyl)- 3.2 g 1 ethanone 2. Benzyl bromide 5.89 ml 3 3. Potassium carbonate 6.82 g 3 4. Acetonitrile 60 ml 20 times

(151) To a mixture of 1-(2,4-dihydroxy-5-isopropyl-phenyl)-ethanone (3.2 g), acetonitrile (60 ml) and potassium carbonate (10.6 g) in a a 250 ml single neck RB flask equipped with a reflux condenser and a guard tube was added benzyl bromide (9.1 ml). The reaction mixture was refluxed (90° C.) for 16 h. The progress of the reaction was monitored by TLC (10% EtOAc/n-hexane, product R.sub.f˜0.5). On completion, acetonitrile was removed under reduced pressure. Water (100 ml) was added to the residue obtained and the resulting mixture was extracted with ethyl acetate (200 ml). The organic layer was dried over sodium sulphate. The solvent was removed under reduced pressure to give a residue to which n-hexane (150 ml) was added to give the product.

(152) TABLE-US-00012 Quantity of the product obtained 5.1 g Yield 83.6% Nature Solid. Colour Colourless

Step 6

2,4-Bis-benzyloxy-5-isopropyl-benzoic acid

(153) ##STR00146##
Material Inputs:

(154) TABLE-US-00013 S. No. Item Quantity 1. 1-(2,4-Bis-benzyloxy-5-isopropyl- 7 g phenyl)-ethanone 2. Sodium hypobromide 13 g in water 100 ml 3. Dioxane 100 ml
Procedure:

(155) A mixture of a mixture of 1-(2,4-Bis-benzyloxy-5-isopropyl-phenyl)-ethanone (7 g) in dioxane (100 ml) in a 500 ml single neck RB flask equipped with a guard tube was cooled to 10° C. and sodium hypobromide [13 g in water (100 ml)] was added. The reaction mixture was stirred overnight at RT. The progress of the reaction was monitored by TLC (30% EtOAc/n-hexane, product R.sub.f˜0.5). On completion, sodium bisulphite (7 g) was added to the reaction mixture which was cooled to 0° C. The reaction mixture was then acidified with HCl (˜10 ml) to pH˜2, extracted with ethyl acetate (100 ml) and washed with water (25 ml). The organic layer was dried over sodium sulphate, and the solvent was removed under reduced pressure to give a residue, which was further purified by column chromatography (SiO.sub.2, 10% EtOAc/n-hexane).

(156) TABLE-US-00014 Quantity of the product obtained 3.4 g Yield 48.3% Nature Solid. Colour Colourless.

C. Synthesis of Isoindoline Intermediates

Preparation C1

Synthesis of 4,7-difluoroisoindoline

(157) ##STR00147##

(158) A mixture of 1,4-difluoro-2,3-dimethylbenzene (4.26 g, 30.0 mmol), N-bromosuccinimide (10.68 g, 60.0 mmol) and dibenzoyl peroxide (75 wt % in water, 120 mg) in carbon tetrachloride (50 ml) was stirred and held at reflux for 16 hours. Upon cooling to room temperature the mixture was filtered, the solids washed with carbon tetrachloride (10 ml), the organic extracts combined and the solvent removed in vacuo to afford 2,3-bis-bromomethyl-1,4-difluorobenzene (9.0 g, 100%) as a pale yellow liquid that solidified upon standing. .sup.1H NMR (DMSO-d.sub.6) 7.36 (2H, dd), 4.78 (4H, s).

(159) A solution of 4-toluenesulphonamide (2.44 g, 14.28 mmol) in N,N-dimethylformamide (10 ml) was added dropwise to a vigorously stirred suspension of sodium hydride (1.2 g, 60 wt % in mineral oil, 30.0 mmol) in anhydrous N,N-dimethylformamide (60 ml). The mixture was stirred at room temperature for 1 hour, at 110° C. for 1 hour and was then cooled to 60° C. and a solution of 2,3-bis-bromomethyl-1,4-difluorobenzene (4.28 g, 14.28 mmol) in N,N-dimethylformamide (30 ml) was added dropwise. The mixture was stirred at 60° C. for 1 hour and then at room temperature for 16 hours. The solvent was removed in vacuo and the residue partitioned between dichloromethane and 1M hydrochloric acid. The organic layer was separated, washed with 5% aqueous potassium carbonate solution, the organics were separated and the solvent removed in vacuo. The residue was rinsed with diethyl ether, filtered and the solids sucked dry under reduced pressure to afford 4,7-difluoro-2-(toluene-4-sulfonyl)isoindoline (2.46 g, 56%) as a pale tan solid. .sup.1H NMR (DMSO-d.sub.6) 7.82 (2H, d), 7.43 (2H, d), 7.15 (2H, dd), 4.66 (4H, s), 2.36 (3H, s). MS: [M+H].sup.+ 310.

(160) A mixture of 4,7-difluoro-2-(toluene-4-sulfonyl)isoindoline (2.36 g, 7.64 mmol), phenol (2.36 g, 25.11 mmol), 48% hydrogen bromide in water (20 ml) and propionic acid (4 ml) was stirred and held at reflux for 6 hours. Upon cooling to room temperature water (50 ml) was added and the mixture extracted with diethyl ether (2×100 ml). The aqueous layer was basified by the addition of 2M sodium hydroxide and was extracted with diethyl ether (3×100 ml). The combined extracts were evaporated to dryness in vacuo to afford 4,7-difluoroisoindoline (586 mg, 50%) as a brown oil that solidified upon standing. .sup.1H NMR (DMSO-d.sub.6) 7.06 (2H, dd), 4.12 (4H, s). MS: [M+H].sup.+ 156.

Preparation C2

Synthesis of 5-hydroxyisoindoline hydrobromide

(161) ##STR00148##

(162) A solution of dimethyl 4-methoxyphthalate (36.75 g, 0.16 mol) in methanol (100 ml) was treated with a solution of potassium hydroxide (28.0 g, 0.5 mol) in water (50 ml) and the mixture was stirred and held at reflux for 4 hours. Upon cooling to room temperature the methanol was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 5M hydrochloric acid. The solid material was filtered off, washed with water and sucked dry under reduced pressure overnight to afford 4-methoxyphthalic acid (31.8 g, 99%) as an off white solid. .sup.1H NMR (DMSO-d.sub.6) 12.90 (2H, br s), 7.74 (1H, d), 7.12-7.05 (2H, m), 3.84 (3H, s). MS: [M+H].sup.+ 197.

(163) Acetic anhydride (40 ml) was added to a mixture of 4-methoxyphthalic acid (30.8 g, 0.16 mol) in anhydrous tetrahydrofuran (150 ml) and the mixture was stirred and held at reflux for 4 hours. Upon cooling to room temperature the solvent was removed in vacuo to afford 4-methoxyphthalic anhydride (27.8 g, 99%) as an off white solid. .sup.1H NMR (DMSO-d.sub.6) 8.02 (1H, d), 7.59 (1H, d), 7.49 (1H, dd), 3.97 (3H, s). MS: [M+H].sup.+ 179.

(164) A mixture of 4-methoxyphthalic anhydride (27.8 g, 0.16 mol) and formamide (175 ml) was stirred and held at 210° C. for 5 hours and was then allowed to cool to room temperature overnight. The solid material was filtered off, washed sequentially with water (100 ml), 50% aqueous acetone (50 ml) and diethyl ether (200 ml) and sucked dry under reduced pressure to afford 4-methoxyphthalimide (21.3 g, 77%) as a pale yellow solid. .sup.1H NMR (DMSO-d.sub.6) 11.15 (1H, br s), 7.74 (1H, d), 7.33-7.28 (2H, m), 3.92 (3H, s).

(165) A stirred solution of 4-methoxyphthalimide (21.3 g, 0.12 mol) in anhydrous tetrahydrofuran (425 ml) at 0° C. was treated dropwise with a solution of borane in tetrahydrofuran (1M, 340 ml, 0.34 mol) and the resulting mixture was stirred and held at reflux for 16 hours. The mixture was cooled to 0° C., methanol (150 ml) was added dropwise followed by 5M hydrochloric acid (150 ml) and the mixture was stirred and held at reflux for 3 hours. Upon cooling to room temperature the organic solvent was removed in vacuo, the mixture was diluted with water (750 ml) and was extracted with dichloromethane (3×750 ml). The aqueous layer was basified to pH 12 or above by the addition of 5M sodium hydroxide, extracted with dichloromethane (3×750 ml) and the combined extracts were evaporated to dryness in vacuo to afford 5-methoxyisoindoline (8.34 g, 47%) as a brown oil. .sup.1H NMR (DMSO-d.sub.6) 7.13 (1H, d), 6.84 (1H, d), 6.74 (1H, dd), 4.05 (2H, s), 4.01 (2H, s), 3.73 (3H, s). MS: [M+H].sup.+ 150.

(166) 5-Methoxyisoindoline (8.34 g, 55.97 mmol) in 48% aqueous hydrobromic acid (100 ml) was stirred and held at reflux for 16 hours. Upon cooling to room temperature the solvent was removed in vacuo to afford 5-hydroxyisoindoline hydrobromide (11.32 g, 93%) as a tan solid. .sup.1H NMR (DMSO-d.sub.6) 9.63 (1H, br s), 9.32 (2H, br s), 7.18 (1H, d), 6.79 (1H, d), 6.76 (1H, dd), 4.42 (2H, t), 4.38 (2H, t). MS: [M+H].sup.+ 136.

Preparation C3

Synthesis of 5-chloro-2,3-dihydro-1H-isoindole

(167) ##STR00149##

(168) A mixture of 3,4-dimethylchlorobenzene (10 g, 71.1 mmol), N-bromosuccinimide (25 g, 142.2 mmol), and benzoyl peroxide (0.147 g, 0.6 mmol), was refluxed in 80 ml of carbon tetrachloride for 18 hours. After cooling, the insoluble material was filtered off and washed with a small amount of carbon tetrachloride. The filtrate and the washings were combined and concentrated under reduced pressure to obtain 20 g of 1,2-bis-bromomethyl-4-chloro-benzene as a pale yellow oil product as a major component.

(169) To a suspension of 60% sodium hydride (3.0 g, 0.125 mmol) in mineral oil in 80 ml of anhydrous DMF (100 ml) was dropwise added a solution of para-toluene sulphonamide (5.6 g, 32.60 mmol) in 30 ml of DMF over 1 hour with vigorous stirring at room temperature. After the addition, the mixture was stirred for 1 hour at room temperature and another 1 hour heating at 90° C. To this mixture was added dropwise a solution of 1,2-bis-bromomethyl-4-chloro-benzene (4 g, 14.18 mmol) in 20 ml of anhydrous DMF at 60° C. and then stirred overnight at room temperature. The resultant mixture was poured onto ice and the resulting precipitate was collected by filtration. The precipitate was washed with 1N hydrochloric acid, 5% sodium carbonate and brine then dried (MgSO.sub.4), filtered and evaporated to give 2.8 g of 5-Chloro-2-(toluene-4-sulphonyl)-2,3-dihydro-1H-isoindole as a pale yellow solid. MS: [M+H].sup.+ 308

(170) 1.0 g of 2-(p-toluensulphonyl)-5-chloroisoindoline and 1.0 g of phenol were added to a mixture of 8 ml of 48% hydrobromic acid and 1.4 ml of propionic acid, and then mixture was heated at reflux for 6 hours. The resultant reaction mixture was diluted with 10 ml of water and extracted twice with 50 ml of ethyl acetate. The water layer was basified with aqueous sodium hydroxide solution and extracted with ethyl acetate three times. The extract was concentrated and the crude product was diluted with 4N HCl/dioxane and stirred for 15 minutes before evaporating the HCl and then re-evaporating with toluene three times to give 0.3 g of 5-chloro-2,3-dihydro-1H-isoindole hydrochloride as a black solid. MS: [M+H].sup.+153-15

Preparation C4

Synthesis of 5-chloro-6-methoxy-2,3-dihydro-1H-isoindole

(171) ##STR00150##

(172) A mixture of 1-chloro-2-methoxy-4,5-dimethyl-benzene (3 g, 17.6 mmol), N-bromosuccinimide (6.3 g, 35.3 mmol), and benzoyl peroxide (0.100 g, 0.41 mmol) in carbon tetrachloride (40 ml) was heated at reflux for 18 hours. After cooling, the insoluble material was removed by filtration, washed with a small amount of carbon tetrachloride and the filtrate evaporated to give 1,2-bis-bromomethyl-4-chloro-5-methoxy-benzene as an oil product as a major component. MS: [M+H].sup.+ 329

(173) A solution of 4-methoxybenzylamine (2.4 g, 17.6 mmol) in acetone (110 ml) was added dropwise to a mixture of 1,2-bis-bromomethyl-4-chloro-5-methoxy-benzene (assumed theoretical, 17.6 mmol) and Na.sub.2CO.sub.3 (12 g, 114 mmol) in acetone/water (10 ml:12.5 ml) then stirred at room temperature for 2 hours and concentrated in vacuo. The crude material was dissolved in ethyl acetate and extracted with 2N HCl. The aqueous layer was neutralized with sodium carbonate, extracted with ethyl acetate (×2), dried (MgSO.sub.4) and evaporated under vacuum to give 5-chloro-6-methoxy-2-(4-methoxy-benzyl)-2,3-dihydro-1H-isoindole (0.8 g, 2.6 mmol) as a brown gum. MS: [M+H].sup.+ 304

(174) A solution of 5-chloro-6-methoxy-2-(4-methoxy-benzyl)-2,3-dihydro-1H-isoindole (600 mg) and anisole (0.3 ml) in trifluoroacetic acid (6 ml) was heated at 180° C. (50 W) for 40 minutes in a CEM discover microwave synthesiser. The reaction mixture was evaporated and re-evaporated with toluene. The crude material was partitioned between DCM and water, the aqueous layer washed with DCM (×3) then evaporated and re-evaporated with toluene to give 5-chloro-6-methoxy-2,3-dihydro-1H-isoindole (256 mg) as green crystals. MS: [M+H].sup.+ 184

Preparation C5

Synthesis of 2,3-dihydro-1H-isoindol-5-ylamine trifluoroacetate

(175) ##STR00151##

(176) A solution of 4-nitro-o-xylene (15.1 g; 0.1 mol) in carbon tetrachloride (150 ml) was treated with N-bromosuccinimide (36 g; 0.2 mol) followed by benzoyl peroxide (1 g) then heated at reflux overnight. The reaction was allowed to cool to ambient temperature, filtered and the filtrate evaporated to give 32 g of crude 1,2-bis-bromomethyl-4-nitro-benzene as a mobile oil. The crude product was dissolved in benzene (200 ml) then treated dropwise over 30 minutes with a solution of 2,4-dimethoxybenzylamine (15 ml) and triethylamine (27.85 ml) in benzene (100 ml) then heated at 80° C. for 3 hours. The reaction was cooled, washed with water followed by saturated sodium bicarbonate. The organics were extracted with 2M HCl (2×150 ml) then combined aqueous basified with 2M NaOH and extracted with EtOAc (×2). The combined EtOAc layer was dried (MgSO.sub.4), evaporated then purified by flash column chromatography eluting with EtOAc/P.E. (1:3-1, 2-1:1). Product containing fraction were combined and evaporated to give 10.15 g of 2-(2,4-dimethoxy-benzyl)-5-nitro-2,3-dihydro-1H-isoindole as a brown solid. .sup.1H NMR (DMSO-d.sub.6) 8.12 (2H, m), 7.50 (1H, d), 7.25 (1H, d), 6.55 (1H, d), 6.52 (1H, dd), 3.93 (4H, s), 3.80 (3H, s), 3.78 (2H, s), 3.75 (3H, s).

(177) 2-(2,4-dimethoxy-benzyl)-5-nitro-2,3-dihydro-1H-isoindole (13 g) in TFA (18 ml) was treated with anisole (6 ml) then heated in a CEM microwave synthesiser at 120° C. (30 Watts) for 20 minutes (carried out batch wise, 6 times). The reaction mixture was evaporated in vacuo and the residue partitioned between DCM and water. The water layer was separated, washed with DCM (×3) then evaporated and re-evaporated with toluene/MeOH (×3) to give 9.8 g of 5-nitro-2,3-dihydro-1H-isoindole trifluoroacetic acid salt as a beige solid. .sup.1H NMR (DMSO-d.sub.6) 9.85 (2H, br s), 8.32 (1H, d), 8.25 (1H, dd), 7.70 (1H, d), 4.68 (2H, s), 4.65 (2H, s).

(178) A mixture of 5-nitro-2,3-dihydro-1H-isoindole trifluoroacetic acid salt (9.8 g) and 10% palladium on carbon (1 g) in methanol (75 ml) was hydrogenated at room temperature and pressure for 16 hours. The reaction was filtered through Celite™, the filtrate evaporated and re-evaporated with toluene to give 8.76 g of 2,3-dihydro-1H-isoindol-5-ylamine mono trifluoroacetic acid salt as a dark brown solid. .sup.1H NMR (DMSO-d.sub.6) 9.45 (2H, br s), 7.05 (1H, d), 6.60 (2H, m), 5.35 (2H, br s), 4.40 (2H, s), 4.30 (2H, s).

Preparation C6

Synthesis of 5-morpholin-4-ylmethyl-2,3-dihydro-1H-isoindole ditrifluoroacetate

(179) ##STR00152##

(180) Steps 1 and 2 were carried out in a manner analogous to that described in Preparation C5 using methyl 3,4-dimethylbenzoate as the starting material.

(181) A mixture of 2-(2,4-dimethoxybenzyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid methyl ester (4.65 g; 14.2 mmol) and lithium hydroxide monohydrate (660 mg; 1.1 equiv.) in 4:1:1 THF-MeOH—H.sub.2O (60 ml) was stirred at room temperature overnight. A further 170 mg of base were added and stirring continued for 7 hours. The reaction was evaporated then re-evaporated with MeOH/toluene (×2). A mixture of the crude 2-(2,4-dimethoxy-benzyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid lithium salt (1.5 g; 4.7 mmol), morpholine (820 μl; 2 equiv.), EDAC (1.1 g; 1.2 equiv.) and HOBt (760 mg; 1.2 equiv.) in DMF (25 ml) was stirred at room temperature overnight then evaporated in vacuo. The residue was partitioned between EtOAc and saturated NaHCO.sub.3, the EtOAc layer was separated, washed with brine, dried (MgSO.sub.4) and evaporated. Purification by flash column chromatography (2% then 5% MeOH/DCM as eluant) gave 1.1 g of [2-(2,4-dimethoxybenzyl)-2,3-dihydro-1H-isoindol-5-yl]-morpholin-4-yl-methanone as a red/brown gum. .sup.1H NMR (DMSO-d.sub.6) 7.30-7.18 (4H, m), 6.56 (1H, d), 6.52 (1H, dd), 3.85 (4H, s), 3.78 (5H, m), 3.73 (3H, s).

(182) A solution of [2-(2,4-dimethoxybenzyl)-2,3-dihydro-1H-isoindol-5-yl]-morpholin-4-yl-methanone (1.05 g; 2.75 mmol) in dry THF (20 ml) under a nitrogen atmosphere was treated with 1M lithium aluminium hydride solution then stirred at room temperature overnight. The reaction was quenched by the cautious addition of saturated sodium sulphate solution, then diluted with EtOAc (40 ml), filtered through Celite™ and evaporated. Purification by flash column chromatography (2% then 5% MeOH/DCM as eluant) gave 340 mg of 2-(2,4-dimethoxybenzyl)-5-morpholin-4-ylmethyl-2,3-dihydro-1H-isoindole as a pale brown gum.

(183) A mixture of 2-(2,4-dimethoxybenzyl)-5-morpholin-4-ylmethyl-2,3-dihydro-1H-isoindole (340 mg) and anisole (350 μl) in trifluoroacetic acid (1.5 ml) was heated at 13° C. in a CEM microwave synthesiser for 1 hour then evaporated and re-evaporated with toluene. The residue was partitioned between DCM and water. The water layer was separated, washed with DCM (×3) then evaporated and re-evaporated with toluene/MeOH (×3) to give 422 mg of 5-morpholin-4-ylmethyl-2,3-dihydro-1H-isoindole ditrifluoroacetate as a brown gum. .sup.1H NMR (DMSO-d.sub.6) 10.30 (1H, br s), 9.60 (2H, br s), 7.55-7.45 (3H, m), 4.45 (4H, s), 4.45-4.30 (2H, m), 4.20-3.88 (2H, m), 3.70-3.55 (2H, m), 3.30-3.00 (4H, m).

Preparation C7

Synthesis of ethyl-2,3-dihydro-1H-isoindole-5-carboxylate trifluoroacetate

(184) ##STR00153##

(185) A solution of 2-(2,4-dimethoxy-benzyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid methyl ester (215 mg) and anisole (200 μl) in 1 ml of TFA was heated at 140° C. for 30 minutes in a CEM discover microwave synthesiser. The reaction was partitioned between water and DCM, the water layer was separated, washed with DCM then evaporated and re-evaporated with toluene/MeOH (×2) to give 105 mg of the title compound. .sup.1H NMR (DMSO-d.sub.6) 9.70 (2H, br s), 8.02 (1H, s), 8.98 (1H, d), 7.57 (1H, d), 4.60 (2H, s), 4.56 (2H, s), 3.89 (3H, s).

Preparation C8

4-Hydroxy-2-(4-methoxy-benzyl)-isoindole-1,3-dione

(186) ##STR00154##

(187) A mixture of 3-hydroxyphthalic anhydride (543 mg, 3.31 mmol), 4-methoxybenzylamine (0.43 mL, 3.31 mmol) and acetic acid (3 mL) was heated at 100° C. for 4 hours. The mixture was allowed to cool and diluted with water (20 mL). The white solid was collected by filtration, washed well with water and dried to give the title compound (760 mg, 81%). .sup.1H NMR (DMSO-d.sub.6) 11.03 (1H, s), 7.61 (1H, dd), 7.28 (1H, d), 7.23-7.19 (3H, m), 6.89-6.86 (2H, m), 4.63 (2H, s), 3.71 (3H, s). MS: [M−H.sup.+] 282.

Preparation C9

4-Hydroxy-2-(2,4-dimethoxy-benzyl)-isoindole-1,3-dione

(188) ##STR00155##

(189) A mixture of 3-hydroxyphthalic anhydride (1.24 g, 7.6 mmol), 2,4-dimethoxybenzylamine (1.14 mL, 7.6 mmol) and acetic acid (5 mL) was heated at 80° C. for 24 hours. The mixture was allowed to cool and diluted with water (20 mL). The white solid was collected by filtration, washed well with water and dried to give the title compound (1.73 g, 73%). .sup.1H NMR (DMSO-d.sub.6) 11.00 (1H, s), 7.62 (1H, dd), 7.29 (1H, d), 7.21 (1H, d), 6.90 (1H, d), 6.56 (1H, d), 6.43 (1H, dd), 4.59 (2H, s), 3.79 (3H, s), 3.72 (3H, s). MS: [M−H.sup.+] 314.

Preparation C10

2-(4-Methoxy-benzyl)-4-[2-(2-methoxy-ethoxy)-ethoxy]-isoindole-1,3-dione

(190) ##STR00156##

(191) 1-(2-Bromo-ethoxy)-2-methoxy-ethane (107 mg, 0.58 mmol) was added to a suspension of 4-hydroxy-2-(4-methoxy-benzyl)-isoindole-1,3-dione (150 mg, 0.53 mmol) and potassium carbonate (200 mg, 1.4 mmol) in DMF (2 mL). After 3.5 hours, a catalytic amount of potassium iodide was added. After a further 17 hours, the mixture was warmed to 60° C. After 3 hours, an additional amount of 1-(2-bromo-ethoxy)-2-methoxy-ethane (20 mg, 0.11 mmol) was added and the mixture maintained at 60° C. for a further 20 hours. The mixture was concentrated in vacuo then the residue was taken up in ethyl acetate and washed with potassium carbonate solution and brine. The organic phase was dried (MgSO.sub.4) and concentrated to give the title compound as a yellow oil (149 mg, 73%). .sup.1H NMR (methanol-d.sub.4) 7.71 (1H, t), 7.43-7.40 (2H, m), 7.31-7.27 (2H, m), 6.87-6.83 (2H, m), 4.71 (2H, s), 4.37-4.34 (2H, m), 3.92-3.89 (2H, m), 3.77-3.74 (5H, m), 3.55-3.53 (2H, m), 3.33 (3H, s). MS: [M+H].sup.+386.

Preparation C11

2-(2,4-Dimethoxy-benzyl)-4-(2-dimethylamino-ethoxy)-isoindole-1,3-dione

(192) ##STR00157##

(193) A mixture of 2-(2,4-dimethoxy-benzyl)-4-hydroxy-isoindole-1,3-dione (317 mg, 1.01 mmol), 2-dimethylaminoethyl chloride hydrochloride (160 mg, 1.11 mmol) and potassium carbonate (350 mg, 2.5 mmol) in DMF (4 mL) was heated at 60° C. for 18 hours. The mixture was concentrated in vacuo, taken up in ethyl acetate and extracted twice with 1N hydrochloric acid. The aqueous extracts were made basic with solid potassium carbonate and extracted with ethyl acetate (×2). The combined organic extracts were washed with brine, dried (MgSO.sub.4) and concentrated to give the title compound (236 mg, 61%) as an off-white solid. .sup.1H NMR (methanol-d.sub.4) 7.73 (1H, t), 7.44-7.40 (2H, m), 7.02 (1H, d), 6.51 (1H, d), 6.42 (1H, dd), 4.72 (2H, s), 4.33 (2H, t), 3.80 (3H, s), 3.76 (3H, s), 2.87 (2H, t), 2.40 (6H, s). MS: [M+H].sup.+ 385.

Preparation C12

2-(2,4-Dimethoxy-benzyl)-4-(3-morpholin-4-yl-propoxy)-isoindole-1,3-dione

(194) ##STR00158##

(195) A mixture of 2-(2,4-dimethoxy-benzyl)-4-hydroxy-isoindole-1,3-dione (313 mg, 1.00 mmol), 4-(3-chloropropyl)morpholine (160 mg, 1.11 mmol) and potassium carbonate (350 mg, 2.5 mmol) in DMF (5 mL) was heated at 60° C. for 18 hours. The mixture was diluted with ethyl acetate and extracted twice with 1N hydrochloric acid. The aqueous extracts were made basic with solid potassium carbonate and extracted with ethyl acetate. The organic extract was washed with brine, dried (MgSO.sub.4) and concentrated to give a yellow solid which was recrystallised from methanol/petrol then ethyl acetate/chloroform/petrol to give the title compound (298 mg, 68%) as an off-white solid. .sup.1H NMR (methanol-d.sub.4) 7.72 (1H, t), 7.41 (1H, d), 7.39 (1H, d), 7.02 (1H, d), 6.51 (1H, d), 6.43 (1H, dd), 4.72 (2H, s), 4.27 (2H, t), 3.81 (3H, s), 3.76 (3H, s), 3.68 (4H, t), 2.61 (2H, t), 2.50 (4H, m), 2.05 (2H, qn). MS: [M+H].sup.+ 441.

Preparation C13

2-(4-Methoxy-benzyl)-4-[2-(2-methoxy-ethoxy)-ethoxy]-2,3-dihydro-1H-isoindole

(196) ##STR00159##

(197) 2-(4-Methoxy-benzyl)-4-[2-(2-methoxy-ethoxy)-ethoxy]-isoindole-1,3-dione (149 mg, 0.38 mmol) was treated with a 1M solution of lithium aluminium hydride in THF (5 mL, 5 mmol). The mixture was maintained at r.t. for 4 hours, 60° C. for 1 hour, then r.t. for a further 18 hours. The mixture was then cooled in ice and quenched by the dropwise addition of water (0.2 mL), 2N sodium hydroxide solution (0.4 mL) and water (0.4 mL). Magnesium sulfate was added, followed by ethyl acetate and then the mixture was stirred at r.t. for 15 minutes. The solids were removed by filtration, being well washed with ethyl acetate. Concentration of the filtrate gave a residue which was absorbed onto an SCX cartridge and washed with 5% methanol/dichloromethane then eluted with 10% 1M ammonia in methanol/dichloromethane to afford the title compound (134 mg, 97%). .sup.1H NMR (methanol-d.sub.4) 7.43-7.39 (2H, m), 7.27 (1H, t), 6.99-6.96 (2H, m), 6.90 (1H, d), 6.88 (1H, d), 4.33 (2H, s), 4.28 (2H, s), 4.23 (2H, s), 4.18-4.15 (2H, m), 3.85-3.79 (5H, m), 3.67-3.64 (2H, m), 3.54-3.51 (2H, m), 3.33 (3H, s). MS: [M+H].sup.+ 358.

Preparation C14

2-(2,4-Dimethoxy-benzyl)-4-(2-dimethylamino-ethoxy)-2,3-dihydro-1H-isoindole

(198) ##STR00160##

(199) 2-(2,4-Dimethoxy-benzyl)-4-(2-dimethylamino-ethoxy)-isoindole-1,3-dione (201 mg, 0.52 mmol) was treated with a 1M solution of lithium aluminium hydride in THF (5 mL, 5 mmol). After 7.5 hours at r.t. a further portion of lithium aluminium hydride solution (5 mL, 5 mmol) was added and the mixture maintained for further 18 hours. The mixture was then cooled in ice and quenched by the dropwise addition of water (0.4 mL), 2N sodium hydroxide solution (0.8 mL) and water (0.8 mL). Magnesium sulfate was added, followed by ethyl acetate and then the mixture was stirred at r.t. for 1 hour. The solids were removed by filtration, being well washed with ethyl acetate. Concentration of the filtrate gave the title compound (192 mg, 103%) as a brown oil which was carried forward without further purification. .sup.1H NMR (methanol-d.sub.4) 7.24 (1H, d), 7.16 (1H, t), 6.82-6.78 (2H, m), 6.55 (1H, d), 6.51 (1H, dd), 4.12 (2H, t), 3.92 (4H, s), 3.86 (2H, s), 3.82 (3H, s), 3.80 (3H, s), 2.76 (2H, t), 2.33 (6H, s). MS: [M+H].sup.+ 357.

Preparation C15

2-(2,4-Dimethoxy-benzyl)-4-(3-morpholin-4-yl-propoxy)-2,3-dihydro-1H-isoindole

(200) ##STR00161##

(201) 2-(2,4-Dimethoxy-benzyl)-4-(3-morpholin-4-yl-propoxy)-isoindole-1,3-dione (298 mg, 0.68 mmol) was treated with a 1M solution of lithium aluminium hydride in THF (5 mL, 5 mmol) and maintained at r.t. for 21 hours. The mixture was heated to 75° C. for 1 hour then cooled in ice and quenched by the dropwise addition of water (0.2 mL), 2N sodium hydroxide solution (0.4 mL) and water (0.4 mL). Magnesium sulfate was added, followed by ethyl acetate and then the mixture was stirred at r.t. for 1 hour. The solids were removed by filtration, being well washed with ethyl acetate. Concentration of the filtrate gave a crude product which was purified by flash chromatography on silica, eluting with 5% methanol in DCM. This afforded the title compound (233 mg, 83%) as a red oil. .sup.1H NMR (methanol-d.sub.4) 7.24 (1H, d), 7.15 (1H, t), 6.80 (1H, d), 6.78 (1H, d), 6.56 (1H, d), 6.52 (1H, dd), 4.05 (2H, t), 3.94 (2H, s), 3.88 (2H, s), 3.87 (2H, s), 3.83 (3H, s), 3.80 (3H, s), 3.70-3.68 (4H, m), 2.54-2.50 (2H, m), 2.49-2.47 (4H, m), 2.00-1.93 (2H, m). MS: [M+H].sup.+ 413.

Preparation C16

4-[2-(2-Methoxy-ethoxy)-ethoxy]-2,3-dihydro-1H-isoindole

(202) ##STR00162##

(203) A solution of 2-(4-methoxy-benzyl)-4-[2-(2-methoxy-ethoxy)-ethoxy]-2,3-dihydro-1H-isoindole (45 mg, 0.13 mmol) in 1,2-dichloroethane (2 mL) was treated with O-chloroethyl chloroformate (0.1 mL, 0.93 mmol). After 17 hours, methanol (5 mL) was added and the mixture stirred for 3 hours. The solvents were removed in vacuo to afford the title compound as a greenish-black solid, which was used without further purification. .sup.1H NMR (methanol-d.sub.4) 7.36 (1H, t), 6.98 (2H, d), 4.60 (2H, s), 4.57 (2H, s), 4.23-4.21 (2H, m), 3.85-3.83 (2H, m), 3.69-3.67 (2H, m), 3.57-3.54 (2H, m), 3.36 (3H, s). MS: [M+H].sup.+ 238.

Preparation C17

[2-(2-(2,3-Dihydro-1H-isoindol-4-yloxy)-ethyl]-dimethyl-amine

(204) ##STR00163##

(205) A solution of 2-(2,4-dimethoxy-benzyl)-4-(2-dimethylamino-ethoxy)-2,3-dihydro-1H-isoindole (170 mg, 0.48 mmol) in trifluoroacetic acid (0.5 mL) and anisole (0.5 mL) was heated at 150° C. under microwave irradiation for 10 minutes. The mixture was diluted with ethyl acetate and extracted twice with water. The combined aqueous extracts were concentrated to give the title compound as a purple oil (240 mg, including residual TFA and/or water). .sup.1H NMR (methanol-d.sub.4) 7.42 (1H, t), 7.07 (1H, d), 7.04 (1H, d), 4.64 (4H, br.s), 4.47-4.44 (2H, m), 3.65-3.63 (2H, m), 3.01 (6H, s). MS: [M+H].sup.+ 207.

Preparation C18

4-(3-Morpholin-4-yl-propoxy)-2,3-dihydro-1H-isoindole

(206) ##STR00164##

(207) A solution of 2-(2,4-dimethoxy-benzyl)-4-(3-morpholin-4-yl-propoxy)-2,3-dihydro-1H-isoindole (233 mg, 0.56 mmol) in trifluoroacetic acid (1.0 mL) and anisole (0.5 mL) was heated at 150° C. under microwave irradiation for 10 minutes. The mixture was diluted with diethyl ether and extracted twice with water. The combined aqueous extracts were concentrated to give an oil which was dissolved in methanol and concentrated in vacuo to afford the title compound as a brown oil (348 mg, including residual TFA and/or water). .sup.1H NMR (methanol-d.sub.4) 7.40 (1H, t), 7.03 (1H, d), 6.99 (1H, d), 4.63 (2H, s), 4.59 (2H, s), 4.21 (2H, t), 4.14-4.04 (2H, m), 3.85-3.73 (2H, m), 3.61-3.52 (2H, m), 3.41-3.36 (2H, m), 3.25-3.13 (2H, m), 2.32-2.25 (2H, m). MS: [M+H].sup.+ 263.

Preparation C19

Synthesis of 4-bromo-2,3-dihydro-1H-isoindole trifluoroacetate

(208) ##STR00165##

(209) Prepared in a manner analogous to 5-nitro-2,3-dihydro-1H-isoindole (described in preparation C5). .sup.1H NMR (DMSO-d.sub.6) 9.73 (2H, br s), 7.60 (1H, d), 7.45 (1H, d), 7.35 (1H, t), 4.65 (2H, s), 4.55 (2H, s).

Preparation C20

Synthesis of 5-bromo-2,3-dihydro-1H-isoindole

(210) ##STR00166##

(211) A mixture of 4-bromophthalic anhydride (25 g) in formamide (75 ml) was heated at 200° C. for 16 hours then allowed to cool to room temperature. The reaction mixture was diluted with water (200 ml), filtered, the filter cake was washed with water then diethyl ether and sucked dry to give 20.85 g of light mustard solid.

(212) 280 ml of 1M Borane-THF complex was added dropwise to a stirred solution of 4-bromophthalimide (20.85 g; 92.2 mmol) in anhydrous THF (200 ml) at 0° C. then heated at reflux overnight. The reaction was cooled to 0° C. then treated cautiously with methanol (100 ml) followed by 2M HCl (100 ml) then heated at reflux for 3 hours. The reaction mixture was cooled and the organics evaporated. The aqueous was diluted with water (100 ml) the extracted with DCM (×3). The aqueous was basified with 2M NaOH then extracted with DCM (×3). The combined DCM extracts were dried (MgSO.sub.4), filtered and evaporated to give 6.99 g of 5-bromo-2,3-dihydro-1H-isoindole as a dark brown gummy solid. .sup.1H NMR (DMSO-d.sub.6) 7.45 (1H, s), 7.36 (1H, d), 7.20 (1H, d), 4.05 (4H, s).

Preparation C21

Synthesis of 2,3-dihydro-1H-isoindole-5-carboxylic acid methyl ester trifluoroacetate

(213) ##STR00167##

(214) 2-(2,4-Dimethoxybenzyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid methyl ester (preparation C6, step 2 product) was deprotected in a manner analogous to 5-nitro-2,3-dihydro-1H-isoindole (described in preparation C5) to give the title compound. .sup.1H NMR (DMSO-d.sub.6) 9.70 (2H, br s), 8.00 (1H, s), 7.95 (1H, d), 7.57 (1H, d), 4.60 (4H, s), 2.88 (3H, s).

D. Synthesis of Benzylated Resorcinol Intermediates

Preparation D1

Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-[5-(2-methoxy-ethoxy)-1,3-dihydro-isoindol-2-yl]-methanone

(215) ##STR00168##

(216) (2,4-Bis-benzyloxy-5-isopropyl-phenyl)-(5-hydroxy-1,3-dihydro-isoindol-2-yl)-methanone (A2 from 2,4-bis-benzyloxy-5-isopropyl-benzoic acid (Preparation B10) and 5-hydroxyisoindoline) (100 mg, 0.2 mmol), 1-chloro-2-methoxy-ethane (23.6 mg, 0.25 mmol) and K.sub.2CO.sub.3 (34.5 mg, 0.25 mmol) in DMF (4 ml) were combined and stirred for 2 hours at room temperature. A further 0.25 mmol of 1-chloro-2-methoxy-ethane and K.sub.2CO.sub.3 was added then heated at 90° C. for 16 hours. Reaction cooled to room temperature and diluted with EtOAc then filtered. The filtrate was reduced in vacuo then purified by flash column chromatography, eluting with 100% petroleum ether to 100% ethyl acetate to afford 115 mg of the title compound as a colourless gel. MS: [M+H].sup.+ 552

Preparation D2

Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-[5-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]-methanone

(217) ##STR00169##

(218) A mixture of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-hydroxy-1,3-dihydro-isoindol-2-yl)-methanone (100 mg, 0.2 mmol), 4-(3-chloropropyl)morpholine (82 mg, 0.5 mmol) and K.sub.2CO.sub.3 (104 mg, 0.75 mmol) in DMF (5 ml) was heated at 90° C. for 16 hours. The reaction mixture was diluted with EtOAc and filtered. The filtrate was reduced in vacuo and purified by flash column chromatography, eluting with 0-100% P.E./EtOAc then 0-10% MeOH/EtOAc to give the title compound as a colourless gel (90.1 mg). MS: [M+H].sup.+ 621.

Preparation D3

Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-ethoxy)-1,3-dihydro-isoindol-2-yl]-methanone

(219) ##STR00170##

(220) A mixture of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-hydroxy-1,3-dihydro-isoindol-2-yl)-methanone (100 mg, 0.2 mmol), 2-dimethylaminoethylchloride.HCl (72 mg, 0.5 mmol) and K.sub.2CO.sub.3 (173 mg, 1.25 mmol) in DMF (5 ml) was heated at 90° C. for 16 hours. Dilute reaction mixture with EtOAc and filtered. The reaction mixture was diluted with EtOAc and filtered. The filtrate was reduced in vacuo and purified by flash column chromatography, eluting 100% DCM then 90% DMAW 90 to give the title compound as an off white gel (79 mg). MS: [M+H].sup.+ 565

Preparation D4

Synthesis of 2,4-bis-benzyloxy-5-isopropyl-benzoyl chloride

(221) ##STR00171##

(222) 2,4-Bis-benzyloxy-5-isopropyl-benzoic acid (Preparation B10) (0.2 g, 0.53 mmol) was dissolved in DCM (10 ml) and treated with oxalyl chloride (1.5 g, 12 mmol) and a catalytic amount of DMF. The reaction mixture was stirred at room temperature for 14 hours and the solvent was then removed in vacuo. The crude material was dissolved in toluene and evaporated. Crude 2,4-bis-benzyloxy-5-isopropyl-benzoyl chloride was obtained as an oil (200 mg).

Preparation D5

Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-morpholin-4-yl-1,3-dihydro-isoindol-2-yl)-methanone

(223) ##STR00172##

(224) A solution of 2,4-bis-benzyloxy-5-isopropyl-benzoic acid (505 mg; 1.3 mmol) (Preparation B5), 5-nitroisoindoline, trifluoroacetate (360 mg; 1 equiv.), EDAC (300 mg; 1.2 equiv.), HOBt (210 mg; 1.2 equiv.) and NEt.sub.3 (270 μl; 1.5 equiv.) in DMF (10 ml) was stirred at room temperature overnight then evaporated in vacuo. The residue was partitioned between EtOAc and 2M HCl, the EtOAc layer was separated, washed with saturated NaHCO.sub.3, dried (MgSO.sub.4) and evaporated. Purification by flash column chromatography (1:4 then 1:2 then 1:1 EtOAc/P.E. as eluant) gave 460 mg of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-nitro-1,3-dihydro-isoindol-2-yl)methanone. MS: [M+H].sup.+ 523.

(225) A solution of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-nitro-1,3-dihydro-isoindol-2-yl)methanone (460 mg; 0.88 mmol) in ethanol (25 ml) was treated with tin (II) chloride dihydrate (1 g; 5 equiv.) then heated at reflux overnight then evaporated in vacuo. The residue was partitioned between EtOAc and saturated NaHCO.sub.3, the EtOAc layer was separated, dried (MgSO.sub.4) and evaporated to give 380 mg of (5-amino-1,3-dihydro-isoindol-2-yl)-(2,4-bis-benzyloxy-5-isopropyl-phenyl)-methanone.

(226) A mixture of (5-amino-1,3-dihydro-isoindol-2-yl)-(2,4-bis-benzyloxy-5-isopropyl-phenyl)-methanone (100 mg; 0.2 mmol), bis(2-chloroethyl)ether (30 μl; 1.1 equiv.), Hunig's base (125 μl; 3.5 equiv.) and tetrabutylammonium iodide (10 mg) in NMP (1 ml) was heated in a CEM microwave synthesiser at 150° C. for 30 minutes. A further 30 μl of Hunigs base and 125 μl of bis(2-chloroethyl)ether were added and heating repeated for the same time. The reaction mixture was partitioned between EtOAc and saturated NH.sub.4Cl solution, the EtOAc layer was separated, washed with more saturated NH.sub.4Cl solution, then brine, dried (MgSO.sub.4) and evaporated. Purification by flash column chromatography (1:2 then 1:1 then 2:1 EtOAc/P.E. as eluant) gave 60 mg of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-morpholin-4-yl-1,3-dihydro-isoindol-2-yl)-methanone. MS: [M+H].sup.+ 563.

Preparation D6

Synthesis of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid

(227) ##STR00173##

(228) A solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid methyl ester (390 mg) in methanol (10 ml) and 2M NaOH (10 ml) was heated at 50° C. for 48 hours then evaporated. The residue was acidified with 2M HCl, the solid collected by filtration, washed with water and sucked dry to give 255 mg of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid as a white solid. [M+H].sup.+ 520.

Examples

(229) By following the methods described above, the compounds set out in the Table below were prepared.

(230) TABLE-US-00015 Example Number Compound Chemical Name Method NMR Data MS 1 (5-Chloro-2- hydroxy-phenyl)- (1,3-dihydro- isoindol-2-yl)- methanone A1. From 5- chloro-2- hydroxy- benzoic acid and isoindoline .sup.1H NMR (MeOH-d.sub.4) 7.20-7.42 (6 H, m), 6.92 (1 H, d), 4.94 (2 H, s), 4.74 (2 H, s) MS: [M + H].sup.+ 274 2 (3-tert-Butyl-4- hydroxy-phenyl)- (2,3-dihydro- indol-1-yl)- methanone A2. From 3- tert-butyl-4- hydroxy- benzoic acid and indoline .sup.1H NMR (DMSO-d.sub.6) 7.56 (2 H, br m), 7.40 (1 H, s), 7.33 (1 H, d), 7.26 (1 H, d), 7.13 (1 H, t), 6.98 (1 H, t), 6.85 (1 H, d), 4.07 (2 H, t), 3.08 (2 H, t), 1.38 (9 H, s) MS: [M + H].sup.+ 296 3 (3-tert-Butyl-4- hydroxy-phenyl)- (3,4-dihydro-2H- quinolin-1-yl)- methanone A2. From 3- tert-butyl-4- hydroxy- benzoic acid and 1,2,3,4- tetrahydro- quinoline .sup.1NMR (DMSO-d.sub.6) 11.05 (1 H, br s), 8.17 (1 H, d), 8.04 (2 H, m), 7.88 (1 H, d), 7.67 (1 H, t), 7.54 (1 H, t), 7.09 (1 H, d), 3.39 (1 H, m), 3.28 (1 H, m), 1.40 (9 H, s), 1.07 (3 H, m), 0.84 (1 H, m) MS: [M + H].sup.+ 310 4 (3,4-Dihydro-1H- isoquinolin-2-yl)- (4-hydroxy-3- isopropyl- phenyl)- methanone A2. From 3- isopropyl-4- hydroxy- benzoic acid and 1,2,3,4- tetrahydro- isoquinoline .sup.1H NMR (DMSO-d.sub.6) 9.77 (1 H, br s), 7.24 (1 H, d), 7.17 (4 H, s), 7.18 (1 H, dd), 6.84 (1 H, d), 4.68 (2 H, s), 3.70 (2 H, br s), 3.23 (1 H, m), 2.87 (2 H, m), 1.18 (6 H, d) MS: [M + H].sup.+ 296 5 (1,3-Dihydro- isoindol-2-yl)- (2,4-dihydroxy-5- isopropyl- phenyl)- methanone A2 & A5. From 2,4-Bis- benzyloxy-5- isopropenyl- benzoic acid (B9) and isoindoline .sup.1H NMR (DMSO-d.sub.6) 10.03 (1 H, s), 9.63 (1 H, s), 7.29 (4 H, br m), 7.03 (1 H, s), 6.40 (1 H, s), 4.77 (4 H, br s), 3.09 (1 H, m), 1.14 (6 H, d) MS: [M + H].sup.+ 298 6 (3-tert-Butyl-4- hydroxy-phenyl)- (1,4-dioxa-8-aza- spiro[4.5]dec-8- yl)-methanone A4. From 3- tert-butyl-4- hydroxy- benzoic acid and 4- piperidine ethylene ketal .sup.1H NMR (DMSO-d.sub.6) 9.82 (1 H, s), 7.22 (1 H, s), 7.13 (1 H, dd), 6.82 (1 H, d), 3.91 (4 H, s), 3.52 (4 H, br m), 1.63 (4 H, bm), 1.37 (9 H, s) MS: [M + H].sup.+ 320 7 0 (3-tert-Butyl-4- hydroxy-phenyl)- (1,3-dihydro- isoindol-2-yl)- methanone A4. From 3- tert-butyl-4- hydroxy- benzoic acid and isoindoline .sup.1H NMR (DMSO-d.sub.6) 9.82 (1 H, s), 7.41 (1 H, s), 7.38 (2 H, dd), 7.29 (3H, br m), 6.82 (1 H, d), 4.82 (4 H, br m), 1.37 (9 H, s) MS: [M + H].sup.+ 296 8 (3-tert-Butyl-4- hydroxy-phenyl)- pyrrolo[3,2- b]pyridin-1-yl- methanone A4. From 3- tert-butyl-4- hydroxy- benzoic acid and 1H-pyrrolo [3,2-b]pyridine .sup.1H NMR (DMSO-d.sub.6) 8.57 (1 H, dd), 8.43 (1 H, d), 7.89 (1 H, dd), 7.63 (1 H, s), 7.56 (1 H, dd), 7.35 (1 H, m), 7.09 (1 H, d), 6.84 (1 H, dd), 1.37 (9 H, s) MS: [M + H].sup.+ 295 9 8-(3-tert-Butyl-4- hydroxy- benzoyl)-2- methyl-2,8-diaza- spiro[4.5]decan- 1-one A4. From 3- tert-butyl-4- hydroxy- benzoic acid and 4-spiro-[3- (N-methyl-2- pyrrolidinone] piperidine hydrochloride .sup.1H NMR (DMSO-d.sub.6) 9.82 (1 H, s), 7.22 (1 H, s), 7.13 (1 H, dd), 6.82 (1 H, d), 3.98 (2 H, br m), 3.34 (2 H, s), 3.13 (2 H, m), 2.71 (3 H, s), 1.92 (2 H, t), 1.60 (2 H, m), 1.43 (2 H, m), 1.37 (9 H, s) MS: [M + H].sup.+ 345 10 (1,3-Dihydro- isoindol-2-yl)-(4- hydroxy-3- isopropyl- phenyl)- methanone A4. From 3- isopropyl-4- hydroxy- benzoic acid and isoindoline .sup.1H NMR (DMSO-d.sub.6) 9.82 (1 H, s), 7.4 (2 H, s), 7.38 (1 H, dd), 7.30 (3 H, m), 6.82 (1 H, d), 4.82 (4 H, dd), 3.23 (1 H, m), 1.23 (6 H, s) MS: [M + H].sup.+ 282 11 (3-tert-Butyl-4- hydroxy-phenyl)- (3,4-dihydro-1H- isoquinolin-2-yl)- methanone A4. From 3- tert-butyl-4- hydroxy- benzoic acid and 1,2,3,4- tetrahydro isoquinoline .sup.1H NMR (DMSO-d.sub.6) 7.22 (1 H, s), 7.13 (5 H, m), 6.82 (1 H, d), 4.70 (2 H, s), 3.75 (2 H, br s), 2.85 (2 H, t), 1.37 (9 H, s) MS: [M + H].sup.+ 310 12 (1,3-Dihydro- isoindol-2-yl)-(5- ethyl-2,4-di- hydroxy-phenyl)- methanone A2, A6 and A5. From 2,4-Bis- benzyloxy-5- bromo-benzoic acid, isoindoline and potassium vinyl trifluoroborate .sup.1H NMR (MeOH-d.sub.4) 7.30 (4 H, s), 7.15 (1 H, s), 6.38 (1 H, s), 4.91 (4 H, s), 2.58 (2 H, q), 1.18 (3 H, t) MS: [M + H].sup.+ 284 13 (5-Cyclopropyl- 2,4-dihydroxy- phenyl)-(1,3- dihydro-isoindol- 2-yl)-methanone A2, A6 and A5. From 2,4-Bis- benzyloxy-5- bromo-benzoic acid, isoindoline and cyclopropane boronic acid .sup.1H NMR (DMSO-d.sub.6) 7.40-7.23 (4 H, m), 6.73 (1 H, s), 6.40 (1 H, s), 4.75 (4 H, br s), 1.92 (1 H, m), 0.78 (2 H, m), 0.53 (2 H, m) MS: [M + H].sup.+ 296 14 (5-sec-Butyl-2,4- dihydroxy- phenyl)-(1,3- dihydro-isoindol- 2-yl)-methanone A2, A6 and A5. From 2,4-Bis- benzyloxy-5- bromo-benzoic acid, isoindoline and 2-buten-2-yl boronic acid .sup.1H NMR (MeOH-d.sub.4) 7.30 (4 H, s), 7.15 (1 H, s), 6.39 (1 H, s), 4.92 (4 H, s), 3.00 (1 H, q), 1.63 (2 H, m), 1.18 (3 H, t), 0.88 (3 H, t) MS: [M + H].sup.+ 312 15 (1,3-Dihydro- isoindol-2-yl)-(3- ethoxy-4- hydroxyphenyl)- methanone Method A4. From 3-ethoxy- 4-hydroxy- benzoic acid and isoindoline .sup.1H NMR (DMSO-d.sub.6) 7.45 (1 H, br s), 7.30 (3 H, d), 7.18 (1 H, d), 7.08 (1 H, dd), 6.85 (1 H, d), 4.85 (4 H, s), 4.10 (2 H, q), 1.38 (3 H, t) MS: [M + H].sup.+ 284 16 (1,3-Dihydro- isoindol-2-yl)- (2,4-dihydroxy- phenyl) methanone A2 and A5. From 2,4-Bis- benzyloxy-5- bromo-benzoic acid and isoindoline .sup.1H NMR (MeOH-d.sub.4) 7.30 (5 H, m), 7.15 (1 H, s), 6.42 (1 H, s), 6.38 (1 H, s), 4.93 (4 H, s) MS: [M + H].sup.+ 256

Example 17

Synthesis of (5-chloro-1,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone

(231) ##STR00190##

(232) A solution of 2,4-bis-benzyloxy-5-isopropyl-benzoic acid (Preparation B10) (0.451 g, 1.2 mmol), EDC (0.276 mg, 1.44 mmol), HOAt (0.196 mg, 1.44 mmol), triethylamine (0.5 ml, 3.6 mmol) and 5-chloro-2,3-dihydro-1H-isoindole (0.187 g, 1.2 mmol) (Preparation C3) in DMF (5 ml) was stirred at room temperature for 16 hours, then evaporated under vacuum. The crude material was dissolved in ethyl acetate and extracted twice with saturated NaHCO.sub.3, organics washed with water three times, then evaporated under vacuum to give 0.5 g of 2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-chloro-1,3-dihydro-isoindol-2-yl)-methanone. MS: [M+H].sup.+ 512

(233) Boron trichloride (1M in DCM) was added dropwise to a solution of 2,4-bis-benzyloxy-5-isopropyl-phenyl)-(5-chloro-1,3-dihydro-isoindol-2-yl)-methanone (0.5 g, 0.97 mmol) in dry DCM (10 ml) at 0° C. under nitrogen, then stirred for at 0° C. for 1 hour, warmed to room temperature and stirred for a further 3 hours. The reaction was quenched with ice, partitioned between DCM and water. The DCM layer was dried (MgSO.sub.4), evaporated under vacuum, then purified by flash silica column chromatography eluting with 80% P.E.: EtOAc to give 0.1 g of (5-chloro-1,3-dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone as a white solid. MS: [M+H].sup.+ 332. .sup.1H NMR (DMSO-d.sub.6) 10.0 (1H, s) 9.60 (1H, s), 7.45 (1H, br s), 7.33 (2H, br s), 7.0 (1H, s), 6.4 (1H, s), 4.80 (4H, br s), 3.10 (1H, m), 1.15 (6H, d).

Example 18

Synthesis of [5-(3-amino-propoxy)-1,3-dihydro-isoindol-2-yl]-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone hydrochloride

(234) ##STR00191##

(235) A solution of {3-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindol-5-yloxy]-propyl}carbamic acid tert-butyl ester (Example 46) (1 g) in EtOAc (10 ml) was treated with a saturated solution of HCl in EtOAC (20 ml) then stirred at room temperature for 2 hours. The reaction mixture was evaporated and re-evaporated with ethanol (×3). The title compound was isolated as a cream foam (840 mg). .sup.1H NMR (DMSO-d6) 10.05 (1H, br s), 9.60 (1H, s), 7.88 (3H, br s), 7.30-7.18 (1H, m), 7.05 (1H, s), 7.00-6.85 (2H, m), 6.42 (1H, s), 4.75 (2H, br s) 4.70 (2H, br S), 4.05 (2H, t), 3.10 (1H, m), 3.00-2.95 (2H, m), 2.00 (2H, tt), 1.15 (6H, d). MS: [M+H].sup.+ 371.

Example 19

(5-Bromo-2,4-dihydroxy-phenyl)-(1,3-dihydro-isoindol-2-yl)-methanone

(236) ##STR00192##

(237) A solution of 5-bromo-2,4-dihydroxy-benzoic acid (520 mg, 2.33 mmol) in DMF (5 mL) was treated with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (471 mg, 2.45 mmol) then HOBt (362 mg, 2.68 mmol). After 25 min, 2,3-dihydro-1H-isoindole (0.5 mL, 2.63 mmol) was added then the mixture was stirred at r.t. for 18 h. The solvent was removed in vacuo then the residue was taken up in ethyl acetate and washed with 1N hydrochloric acid, saturated sodium bicarbonate solution and brine then dried (MgSO.sub.4) and concentrated. The residue was triturated with methanol to afford the title compound as a grey solid (328 mg, 44%). .sup.1H NMR (DMSO-d.sub.6) 10.45 (1H, s), 10.32 (1H, s), 7.36 (1H, br.s), 7.35 (1H, s), 7.28 (3H, br.s), 6.59 (1H, s), 4.77 (2H, br.s), 4.71 (2H, br.s). MS: [M+H].sup.+ 332/334.

Example 20

(1,3-Dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-trifluoromethyl-phenyl)-methanone

20A. (2,4-Bis-benzyloxy-5-bromo-phenyl)-(1,3-dihydro-isoindol-2-yl)-methanone

(238) ##STR00193##

(239) According to general method A2, 2,4-bis-benzyloxy-5-bromo-benzoic acid (1.02 g, 2.47 mmol) gave a residue which was purified by flash chromatography on silica (ethyl acetate/petrol gradient, 0-20%) to afford the title compound as a white crystalline solid (501 mg, 39%). .sup.1H NMR (methanol-d.sub.4) 7.52 (1H, s), 7.49-7.46 (2H, m), 7.42-7.37 (2H, m), 7.34 (t, 2H), 7.30-7.24 (4H, m), 7.23-7.20 (3H, m), 7.16 (1H, d), 6.94 (1H, s), 5.24 (2H, s), 5.16 (2H, s), 4.86 (2H, s), 4.60 (2H, s). MS: [M+H].sup.+ 514/516.

20B. (2,4-Bis-benzyloxy-5-trifluoromethyl-phenyl)-(1,3-dihydro-isoindol-2-yl)-methanone

(240) ##STR00194##

(241) A mixture of (2,4-bis-benzyloxy-5-bromo-phenyl)-(1,3-dihydro-isoindol-2-yl)-methanone (491 mg, 0.95 mmol), sodium trifluoroacetate (649 mg, 4.8 mmol) and copper (I) iodide (364 mg, 1.91 mmol) were dried under vacuum (0.04 mbar) for 6 hours. The flask was flushed with nitrogen, DMF (5 mL) was added and the mixture heated at 150° C. for 17 hours. After cooling to r.t., the mixture was diluted with DCM (100 mL) and filtered through Celite, rinsing with DCM. The filtrate was concentrated to dryness and the residue was partially purified by flash chromatography on silica (ethyl acetate/petrol gradient, 0-20%). The purest fraction was recrystallised from methanol to afford the title compound as a white solid (140 mg, 29%). .sup.1H NMR (methanol-d.sub.4) 7.60 (1H, s), 7.48-7.44 (2H, m), 7.40 (2H, t), 7.37-7.21 (m, 9H), 7.17 (1H, d), 7.02 (1H, s), 5.29 (2H, s), 5.24 (2H, s), 4.88 (2H, s), 4.62 (2H, s). MS: [M+H].sup.+ 504.

20C. (1,3-Dihydro-isoindol-2-yl)-(2,4-dihydroxy-5-trifluoromethyl-phenyl)-methanone

(242) ##STR00195##

(243) A solution of (2,4-bis-benzyloxy-5-trifluoromethyl-phenyl)-(1,3-dihydro-isoindol-2-yl)-methanone (140 mg, 0.28 mmol) in methanol (5 mL) was hydrogenated at atmospheric pressure over 10% palladium on charcoal (34 mg) for 4 hours. A further portion of catalyst was added (31 mg) and hydrogenation continued for a further 1.5 hours. The mixture was filtered through Celite, eluting with methanol, then the filtrate was concentrated in vacuo to afford the title compound as a white solid (91 mg, quant.). .sup.1H NMR (DMSO-d.sub.6) 10.79 (1H, s), 10.70 (1H, s), 7.40-7.35 (2H, m), 7.31-7.35 (3H, m), 6.61 (1H, s), 4.79 (2H, br.s), 4.68 (2H, br.s). MS: [M+H].sup.+ 324.

Example 21

(2,4-Dihydroxy-5-isopropyl-phenyl)-{4-[2-(2-methoxy-ethoxy)-ethoxy]-1,3-dihydro-isoindol-2-yl}methanone

(244) ##STR00196##

(245) A solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (96 mg, 0.26 mmol) and DMF (1 drop, cat.) in DCM (3 mL) was cooled in ice then treated with oxalyl chloride (112 □L, 1.28 mmol). After 2 hours the mixture was concentrated in vacuo then azeotroped with toluene. The resulting acid chloride was dissolved in DCM (4 mL) and added to a solution of 4-[2-(2-methoxy-ethoxy)-ethoxy]-2,3-dihydro-1H-isoindole (0.26 mmol, assuming a quantitative yield from the preceding step (debenzylation procedure C16)) and triethylamine (0.20 mL, 1.4 mmol) in DCM (1 mL). After 2 hours the mixture was diluted with ethyl acetate and washed with 1N hydrochloric acid, brine, sodium bicarbonate solution and brine. The organic phase was dried (MgSO.sub.4) and concentrated to give a black residue. This was partially purified by flash chromatography on silica (ethyl acetate/petrol gradient, 20-33%) to afford an impure sample of the intermediate (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-{4-[2-(2-methoxy-ethoxy)-ethoxy]-1,3-dihydro-isoindol-2-yl}-methanone.

(246) A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-{4-[2-(2-methoxy-ethoxy)-ethoxy]-1,3-dihydro-isoindol-2-yl}-methanone in methanol (5 mL) was hydrogenated at atmospheric pressure over 10% palladium on charcoal (12 mg) for 3 hours. A further portion of catalyst (12 mg) was added and hydrogenation continued for a further 7 hours. The mixture was filtered through Celite, eluting with methanol, then the filtrate was concentrated in vacuo to give a residue which was purified by preparative HPLC (basic method). This afforded the title compound as a white solid (17 mg, 16% over two steps). .sup.1H NMR (methanol-d.sub.4) 7.25 (1H, t), 7.17 (1H, s), 6.95-6.82 (2H, m), 6.37 (1H, s), 4.89 (2H, br.s), 4.83 (overlaps with H.sub.2O, br.s), 4.16 (2H, br.s), 3.82 (2H, br.s), 3.66 (2H, br.s), 3.52 (2H, br.s), 3.39-3.28 (overlaps with MeOH, m), 3.20 (1H, sept), 1.21 (6H, d). MS: [M+H].sup.+ 416.

Example 22

(2,4-Dihydroxy-5-isopropyl-phenyl)-[4-(2-dimethylamino-ethoxy)-1,3-dihydro-isoindol-2-yl]-methanone

(247) ##STR00197##

(248) A solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (189 mg, 0.50 mmol) and DMF (1 drop, cat.) in DCM (5 mL) was cooled in ice then treated with oxalyl chloride (1120 L, 1.28 mmol). After 2 hours the mixture was concentrated in vacuo then azeotroped with toluene. The resulting acid chloride was dissolved in DCM (5 mL) and added to a solution of [2-(2,3-dihydro-1H-isoindol-4-yloxy)-ethyl]-dimethyl-amine (0.48 mmol, assuming a quantitative yield from the preceding step (C17)) and triethylamine (0.50 mL, 3.6 mmol) in DCM (3 mL). After 16 hours the mixture was diluted with ethyl acetate and washed with saturated potassium carbonate solution and brine. The organic phase was dried (MgSO.sub.4) and concentrated to give a residue which was partially purified by flash chromatography on silica (methanol/DCM gradient, 5-10% followed by 10% 2M methanolic ammonia/DCM) to afford an impure sample of the intermediate (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(2-dimethylamino-ethoxy)-1,3-dihydro-isoindol-2-yl]-methanone.

(249) A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(2-dimethylamino-ethoxy)-1,3-dihydro-isoindol-2-yl]-methanone in methanol (5 mL) was hydrogenated at atmospheric pressure over 10% palladium on charcoal (40 mg) for 22 hours. The mixture was filtered through Celite, eluting with methanol, then the filtrate was concentrated in vacuo to give a residue which was purified by preparative HPLC (acidic method). This afforded the formate salt of the title compound as a white solid (9 mg, 5% over two steps). .sup.1H NMR (methanol-d.sub.4) 8.52 (0.7H, s), 7.29 (1H, t), 7.17 (1H, s), 6.98-6.86 (2H, m including 6.90 (1H, d)), 6.37 (1H, s), 4.89 (2H, br.s), 4.87 (2H, br.s), 4.28 (2H, br.s), 3.29-3.5 (3H, m including 3.20 (1H, sept)), 2.81-2.51 (6H, br.d), 1.21 (6H, d). MS: [M+H].sup.+ 385.

Example 23

(2,4-Dihydroxy-5-isopropyl-Phenyl)-[4-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]methanone

(250) ##STR00198##

(251) A solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (210 mg, 0.56 mmol) and diisopropylethylamine (0.25 mL, 1.4 mmol) in DCM (5 mL) was treated with bromo-tris-pyrrolidino-phosphonium hexafluorophosphate (PyBrOP) (287 mg, 0.62 mmol). After 1 hour a solution of 4-(3-morpholin-4-yl-propoxy)-2,3-dihydro-1H-isoindole (0.56 mmol, assuming a quantitative yield from the preceding step(C18)) in DCM (5 mL) was added. After 4 hours the mixture was diluted with ethyl acetate and washed with water, 1N sodium hydroxide solution and brine. The organic phase was dried (MgSO.sub.4) and concentrated to give a residue which was absorbed onto an SCX column. This was washed with 10% methanol/DCM then the product was eluted with 25% 2M methanolic ammonia/DCM) to afford an impure sample of the intermediate (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]-methanone.

(252) A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]-methanone in methanol (5 mL) was hydrogenated at atmospheric pressure over 10% palladium on charcoal (45 mg) for 4 hours. The mixture was filtered through Celite, eluting with methanol, then the filtrate was concentrated in vacuo to give a residue which was purified by preparative HPLC (basic method). This afforded the title compound as a white solid (16 mg, 6% over two steps). .sup.1H NMR (methanol-d.sub.4) 7.24 (1H, t), 7.18 (1H, s), 6.89 (1H, d), 6.84 (1H, d), 6.37 (1H, s), 4.87 (2H, br.s), 4.78 (2H, br.s), 4.11-4.04 (2H, m), 3.72-3.66 (4H, m), 3.21 (1H, sept), 2.60-2.42 (6H, m), 2.05-1.92 (2H, m), 1.21 (6H, d). MS: [M+H].sup.+ 441.

Examples 24 to 47

(253) By following the methods described above, the compounds of Examples 24 to 47 were prepared.

(254) TABLE-US-00016 Exam- ple Number Compound Chemical Name Method N.M.R. Data M.S. 24 (3-sec-Butyl-4- hydroxy-phenyl)- (1,3-dihydro- isoindol-2-yl)- methanone A2 and A5. From (Z)-4- benzyloxy-3-(1- methyl- propenyl)- benzoic acid and isoindoline .sup.1H NMR (DMSO-d.sub.6) 9.73 (1 H, br s), 7.37 (1 H, d), 7.32 (1 H, dd), 7.30 (4 H, br s), 6.86 (1 H, d), 4.87 (2 H, s), 4.82 (2 H, s), 3.03 (1 H, m), 1.63 (1 H, m), 1.57 (1 H, m), 1.19 (3 H, d), 0.82 (3 H, t) MS: [M + H].sup.+ 296 25 00 (5-tert-Butyl-2,4- dihydroxy- phenyl)-(1,3- dihydro-isoindol- 2-yl)-methanone A2 and A5. From 2,4-bis- benzyloxy-5- tert-butyl- benzoic acid and isoindoline .sup.1H NMR (DMSO-d.sub.6) 7.34 (2 H, m), 7.29 (2 H, m), 7.10 (1 H, s), 6.33 (1 H, s), 4.83 (4 H, s), 1.35 (9 H, s) MS: [M + H].sup.+ 312 26 01 (5-Chloro-2,4- dihydroxy- phenyl)-(1,3- dihydro-isoindol- 2-yl)-methanone A2 and A3. From 2,4-bis- benzyloxy-5- chloro-benzoic acid and isoindoline .sup.1H NMR (DMSO-d.sub.6) 10.42 (1 H, s), 10.33 (1 H, s), 7.38 (2 H, m), 7.30 (2 H, m), 7.24 (1 H, s), 6.60 (1 H, s), 4.78 (2 H, br s), 4.72 (2 H, br s) MS: [M + H].sup.+ 290 27 02 (1,3-Dihydro- isoindol-2-yl)-(2- hydroxy-5- isopropyl-4- methoxy-phenyl)- methanone A2, A5 & A7. From 2,4-bis- benzyloxy-5- isopropenyl- benzoic acid and isoindoline H NMR (DMSO-d.sub.6) 10.21 (1 H, br s), 7.33 (2 H, br s), 7.28 (2 H, br s), 7.13 (1 H, s), 6.50 (1 H, s), 4.80 (4 H, br s), 3.79 (3 H, s), 3.15 (1 H, m), 1.14 (6 H, d) MS: [M + H].sup.+ 312 28 03 (4,7-difluoro-1,3- dihydro-isoindol- 2-yl)-(2,4- dihydroxy-5- isopropyl- phenyl)- methanone A2 and A5. From 2,4-bis- benzyloxy-5- isopropenyl- benzoic acid and 4,7- difluoro- isoindoline H NMR (DMSO-d.sub.6) 9.97 (1 H, br s), 9.66 (1 H, br s), 7.22 (2 H, dd), 7.03 (1 H, s), 6.42 (1 H, s), 4.84 (4 H, br s), 3.10 (1 H, m), 1.13 (6 H, d) MS: [M + H].sup.+ 334 29 04 (2,4-dihydroxy-5- isopropyl- phenyl)-(5-fluoro- 1,3-dihydro- isoindol-2-yl)- methanone A2 and A5. From 2,4-bis- benzyloxy-5- isopropenyl- benzoic acid and 5-fluoro- isoindoline [Ref. U.S. Pat. No. 5,026,856] H NMR (DMSO-d.sub.6) 10.02 (1 H, br s), 9.58 (1 H, s), 7.37 (1 H, br m), 7.20 (1 H, br m), 7.12 (1 H, td), 7.04 (1 H, s), 6.41 (1 H, s), 4.78 (2 H, br s), 4.75 (2 H, br s), 3.11 (1 H, m), 1.16 (6 H, d) MS: [M + H].sup.+ 316 30 05 (1,3-dihydro- isoindol-2-yl)-(3- fluoro-2,4- dihydroxy-5- isopropyl- phenyl)- methanone A8. From (1,3- dihydro- isoindol-2-yl)- (2,4-dihydroxy- 5-isopropyl- phenyl)- methanone H NMR (DMSO-d.sub.6) 12.23 (1 H, br s), 7.39 (1 H, m), 7.35-7.25 (3 H, m), 6.84 (1 H, d), 5.53 (1 H, s), 4.74 (2 H, s), 4.59 (2 H, s), 2.52 (1 H, m), 1.11 (3 H, d), 0.84 (3 H, d); .sup.19F NMR (DMSO- d6) 19.3 MS: [M + H].sup.+ 316 31 06 (1,3-dihydro- isoindol-2-yl)-(2- fluoro-4,6- dihydroxy-3- isopropyl- phenyl)- methanone A8. From (1,3- dihydro- isoindol-2-yl)- (2,4-dihydroxy- 5-isopropyl- phenyl)- methanone H NMR (DMSO-d.sub.6) 12.03 (1 H, br s), 7.40-7.35 (2 H, m), 7.33- 7.28 (2 H, m), 6.53 (1 H, br d), 5.53 (1 H, s), 5.07 (1 H, brd), 4.98 (1 H, br d), 4.79 (2 H, s), 2.90 (1 H m), 1.03 (6 H, m); .sup.19F NMR (DMSO-d.sub.6) 24.9 MS: [M + H].sup.+ 316 32 07 (2,4-dihydroxy-5- isopropyl- phenyl)-(4-fluoro- 1,3-dihydro- isoindol-2-yl)- methanone hydrochloride From 2,4-bis- benzyloxy-5- isopropyl- benzoic acid (B10) and 4- fluoro-2,3- dihydro-1H- isoindole H NMR (DMSO-d.sub.6) 7.35 (2 H, m), 7.20 (1 H, m), 7.1 (1 H, t), 7.0 (1 H, s), 6.4 (1 H, s), 4.80 (4 H, br s), 1.20 (6 H, s) MS: [M + H].sup.+ 316 33 08 (5-chloro-6- methoxy-1,3- dihydro-iso-indol- 2-yl)-(2,4- dihydroxy-5- isopropyl- phenyl)- methanone A2 and A5. From 2,4-bis- benzyloxy-5- isopropyl- benzoic acid (B10) and 5- chloro-6- methoxy-2,3- dihydro-1H- isoindole .sup.1H NMR (Me- d.sub.3-OD) 7.32 (1 H, s), 7.17 (1 H, s), 7.05 (1 H, s), 6.37 (1 H, s), 4.89 (2 H, s), 3.89 (3 H, s), 3.36 (3 H, m), 1.23 (6 H, d) MS: [M + H].sup.+ 362 34 09 (2,4-dihydroxy-5- isopropyl- phenyl)-[5-(2- methoxy-ethoxy)- 1,3-dihydro- isoindol-2-yl]- methanone A5. From (2,4- bis-benzyloxy- 5-isopropyl- phenyl)-[5-(2- methoxy- ethoxy)-1,3- dihydro- isoindol-2-yl]- methanone H NMR (DMSO-d.sub.6) 10.02 (1 H, s), 9.60 (1 H, s), 7.22 (1 H, brs), 7.03 (1 H, s), 6.90 (1 H, br s), 6.85 (1 H, d), 6.4 (1 H, s), 4.74 (4 H, br d), 4.08 (2 H, br s), 3.65 (2 H, t), 3.18-3.03 (1 H, m), 1.15 (6 H, s), 3.30 (3 H, s) MS: [M + H].sup.+ 372 35 0 (2,4-dihydroxy-5- isopropyl- phenyl)-[5-(3- morpholin-4-yl- propoxy)-1,3- dihydro-isoindol- 2-yl]-methanone A5. From (2,4- bis-benzyloxy- 5-isopropyl- phenyl)-[5-(3- morpholin-4-yl- propoxy)-1,3- dihydro- isoindol-2-yl]- methanone H NMR (DMSO-d.sub.6) 10.02 (1 H, s), 9.60 (1 H, s), 7.22 (1 H, br s), 7.03 (1 H, s), 6.90 (1 H, br s), 6.85 (1 H, d), 6.4 (1 H, s), 4.74 (4 H, br d), 4.08 (2 H, br s), 3.55 (4 H, brs), 3.18- 3.03 (1 H, m), 2.40 (2 H, s), 2.38 (4 H, br s), 1.85 (2 H, t), 1.15 (6 H, s) MS: [M + H].sup.+ 441 36 (2,4-dihydroxy-5- isopropyl- phenyl)-[5-(2- dimethylamino- ethoxy)-1,3- dihydro-isoindol- 2-yl]-methanone A5. From (2,4- bis-benzyloxy- 5-isopropyl- phenyl)-[5-(2- dimethylamino- ethoxy)-1,3- dihydro- isoindol-2-yl]- methanone H NMR (DMSO-d.sub.6) 10.02 (1 H, s), 9.60 (1 H, s), 7.22 (1 H, br s), 7.03 (1 H, s), 6.90 (1 H, br s), 6.85 (1 H, d), 6.40 (1 H, s), 4.74 (4 H, br d), 4.08 (2 H, br s), 3.18-3.03 (1 H, m), 2.71 (2 H, br s), 2.30 (6 H, s), 1.15 (6 H, s) MS: [M + H].sup.+ 385 37 (2,4-dihydroxy-5- isopropyl- phenyl)-(2-oxa- 5-aza- bicyclo[2.2.1]hept- 5-yl)-methanone A2 and A5. From (2,4- bis-benzyloxy- 5-isopropenyl- benzoic acid (Preparation B5) and 2-oxa- 5-aza- bicyclo[2.2.1]- heptane H NMR (DMSO-d.sub.6) 9.64 (1 H, s), 7.02 (1 H, s), 6.31 (1 H, s), 4.65 (2 H, s), 3.78 (2 H, dd), 3.31 (2 H, s), 3.07 (1 H, m), 1.77 (2 H, m), 1.10 (6 H, m) MS: [M + H].sup.+ 278 38 (3,4-dihydro-1H- isoquinolin-2-yl)- (2,4-dihydroxy-5- isopropyl- phenyl)- methanone A2 and A5. From (2,4- bis-benzyloxy- 5-isopropenyl- benzoic acid (Preparation B5) and 1,2,3,4- tetrahydro- isoquinoline .sup.1H NMR (Me- d.sub.3-OD) 7.19 (1 H, s), 7.14- 7.09 (1 H, br s), 7.02 (1 H, s), 6.37 (1 H, s), 4.75 (2 H, s), 3.80 (2 H, t), 3.24-3.15 (1 H, m), 2.95 (2 H, t), 1.19 (6 H, d) MS: [M + H].sup.+ 312 39 (5-amino-1,3- dihydro-isoindol- 2-yl)-(2,4- dihydroxy-5- isopropyl- phenyl)- methanone A2 & A5. From 2,4-bis- benzyloxy-5- isopropenyl- benzoic acid (Preparation B5) and 5-nitro- isoindoline. TFA (C5 but omitting hydrogenation step) H NMR (DMSO-d.sub.6) 7.05 (1 H, s), 6.95- 6.85 (1 H, m), 6.60-6.50 (2 H, m), 6.25 (1 H, s), 4.6-4.5 (4 H, m), 3.10 (1 H, h), 1.10 (6 H, d) MS: [M + H] + 313 40 (2,4-dihydroxy-5- isopropyl- phenyl)-(5- methoxy-1,3- dihydro-isoindol- 2-yl)-methanone A2 and A5. From 2,4-bis- benzyloxy-5- isopropenyl- benzoic acid (Preparation B5) and 5- methoxy- isoindoline. H NMR (DMSO-d.sub.6) 10.05 (1 H, s), 9.60 (1 H, s), 7.30-7.15 (1 H, m), 7.05 (1 H, s), 7.00-6.85 (1 H, m), 6.82 (1 H, d), 6.40 (1 H, s), 4.75 (2 H, s) 4.70 (2 H, s), 3.75 (3 H, s), 3.10 (1 H, m), 1.13 (6 H, d) MS: [M + H] + 328 41 (2,4-dihydroxy-5- isopropyl- phenyl)-(5- morpholin-4-yl- 1,3-dihydro- isoindol-2-yl)- methanone A5 from (2,4- bis-benzyloxy- 5-isopropyl- phenyl)-(5- morpholin-4-yl- 1,3-dihydro- isoindol-2-yl)- methanone (D5). H NMR (DMSO-d.sub.6) 9.60 (1 H, br s), 7.30- 7.15 (1 H, m), 7.05 (1 H, s), 7.00-6.90 (2 H, m), 6.40 (1 H, s), 4.75 (2 H, s) 4.70 (2 H, s), 3.75 (4 H, m), 3.15-3.05 (5 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 383 42 (2,4-dihydroxy-5- isopropyl- phenyl)-[5-(4- methyl-piperazin- 1-yl)-1,3-dihydro- isoindol-2-yl]- methanone As for Example 41 but using bis(2- chloroethyl)- methylamine hydrochloride in step 2. H NMR (DMSO-d.sub.6) 7.30-7.15 (1 H, m), 7.05 (1 H, s), 6.95-6.85 (2 H, m), 6.40 (1 H, s), 4.70 (2 H, br s) 4.65 (2 H, brs), 3.15- 3.05 (5 H, m), 2.45 (4 H, m), 2.20 (4 H, s), 1.85 (3 H, s), 1.15 (6 H, d) MS: [M + H].sup.+ 396 43 2-(2,4-dihydroxy- 5-isopropyl- benzoyl)-2,3- dihydro-1H- isoindole-5- carboxylic acid methyl ester A2 and A5. From 2,4- bis-benzyloxy- 5-isopropenyl- benzoic acid (Preparation B5) and 2,3- dihydro-1H- isoindole-5- carboxylic acid methyl ester, TFA (Preparation C21) H NMR (DMSO-d.sub.6) 10.05 (1 H, br s), 9.60 (1 H, s), 8.00-7.92 (1 H, m), 7.90 (1 H, s), 7.55-7.42 (1 H, m), 7.05 (1 H, d), 6.40 (1 H, s), 4.85 (4 H, br s) 3.85 (3 H, s), 3.10 (1 H, m), 1.13 (6 H, d) MS: [M + H].sup.+ 356 44 2-(2,4-dihydroxy- 5-isopropyl- benzoyl)-2,3- dihydro-1H- isoindole-5- carboxylic acid A5, from 2-(2,4- bis-benzyloxy- 5-isopropyl- benzoyl)-2,3- dihydro-1H- isoindole-5- carboxylic acid. H NMR (DMSO-d.sub.6) 12.90 (1 H, br s), 10.05 (1 H, br s), 9.60 (1 H, s), 8.00-7.92 (1 H, m), 7.90 (1 H, d), 7.55- 7.40 (1 H, m), 7.05 (1 H, d), 6.45 (1 H, s), 4.85 (4 H, br s) 3.10 (1 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 342 45 0 (2,4-dihydroxy-5- isopropyl- phenyl)-(5- morpholin-4- ylmethyl-1,3- dihydro-isoindol- 2-yl)-methanone hydrochloride A2 and A5. From 2,4- bis-benzyloxy- 5-isopropenyl- benzoic acid (Preparation B5) and 5- morpholin-4- ylmethyl-2,3- dihydro-1H- isoindole ditrifluoro- acetate (C6). H NMR (DMSO-d.sub.6) 11.03 (1 H, br s), 10.05 (1 H, br s), 9.78 (1 H, br s), 7.60-7.38 (3 H, m), 7.05 (1 H, s), 6.45 (1 H, s), 4.80 (4 H, m), 4.33 (2 H, d), 3.95- 3.85 (2 H, m), 3.32-3.22 (2 H, m), 3.28-3.00 (5 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 397 46 {3-[2-(2,4- dihydroxy-5- isopropyl- benzoyl)-2,3- dihydro-1H- isoindol-5-yloxy]- propyl}-carbamic acid tert-butyl ester As for Example 34, A2 (from benzyloxy-5- isopropyl- benzoic acid (Preparation B5) and 5- hydroxy- isoindoline), alkylation using 3-(BOC- amino)propyl bromide, then A5. H NMR (DMSO-d.sub.6) 10.05 (1 H, br s), 9.60 (1 H, s), 7.30-7.15 (1 H, m), 7.05(1 H, s), 6.98-6.80 (3 H, m), 6.40 (1 H, s), 4.75 (2 H, br s) 4.70 (2 H, br s), 3.95 (2 H, s), 3.15-3.05 (3 H, m), 1.80 (2 H, tt), 1.37 (9 H, s), 1.15 (6 H, d) MS: [M + H].sup.+ 471 47 (2,4-dihydroxy-5- isopropyl- phenyl)-(5- methyl-1,3- dihydro-isoindol- 2-yl)-methanone A2 and A5. From 2,4- bis-benzyloxy- 5-isopropenyl- benzoic acid (Preparation B5) and 5- morpholin-4- ylmethyl-2,3- dihydro-1H- isoindole ditrifluoro- acetate (C6). Biproduct from Example 45. H NMR (DMSO-d.sub.6) 10.05 (1 H, s), 9.60 (1 H, s), 7.25-7.08 (3 H, m), 7.05 (1 H, s), 6.40 (1 H, s), 4.75 (4 H, m), 3.10 (1 H, m), 2.30 (1 H, s), 1.15 (6 H, d) MS: [M + H].sup.+ 312

Example 48

Synthesis of (2,4-dihydroxy-5-isopropyl-Phenyl-[5-(2-isopropylamino-ethoxy)-1,3-dihydro-isoindol-2-yl]-methanone

(255) ##STR00223##

(256) To a suspension of [5-(3-amino-ethoxy)-1,3-dihydro-isoindol-2-yl]-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone hydrochloride (Example 57) (250 mg, 0.702 mmoles) in 1,2-dichloroethane (10 ml) was added acetone (62 μl, 0.842 mmoles), sodium triacetoxyborohydride (178 mg, 0.842 mmoles) and acetic acid (48 μl, 0.842 mmoles) and then heated at 60° C. for 24 hours. To the reaction mixture was added further acetone (52 μl, 0.702 mmoles), sodium triacetoxyborohydride (149 mg, 0.702 mmoles) and acetic acid (40 μl, 0.702 mmoles) and heated at 60° C. for a further 2 hours. The reaction mixture was then filtered and the mother liquor purified by flash chromatography [Biotage SP4: 25M, flow rate 25 ml/min, gradient 20% to 100% DMAW 90 in DCM) to give (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(2-isopropylamino-ethoxy)-1,3-dihydro-isoindol-2-yl]-methanone as a light brown viscous oil (140 mg, 50%). .sup.1H NMR (DMSO-d6) 10.05 (1H, br s); 9.60 (1H, br s); 7.23 (1H, br s); 7.05 (1H, s); 6.93 (1H, br s); 6.85 (1H, br d); 6.40 (1H, s); 4.70 (4H, br m); 4.00 (2H, t); 3.10 (1H, m); 2.90 (2H, t); 2.80 (1H, m); 1.15 (6H, d); 1.00 (6H, d). MS: [M+H].sup.+ 399.

Example 49

Synthesis of N-{2-[2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindol-5-yloxy]-ethyl}-2-morpholin-4-yl-acetamide

(257) ##STR00224##

(258) To a solution of [5-(3-amino-ethoxy)-1,3-dihydro-isoindol-2-yl]-(2,4-dihydroxy-5-isopropyl-phenyl)-methanone hydrochloride (100 mg, 0.255 mmoles) in DMF (10 ml) was added EDC (59 mg, 0.306 mmoles), HOBt (41 mg, 0.306 mmoles), morpholin-4-yl-acetic acid (37 mg, 0.255 mmoles) and triethylamine (43p1, 0.306 mmoles) and stirred at ambient temperature for one hour. To the reaction mixture was added further EDC (20 mg, 0.104 mmoles), HOBt (14 mg, 0.104 mmoles), morpholin-4-yl-acetic acid (12 mg, 0.083 mmoles) and triethylamine (14 μl, 0.100 mmoles) and stirred at ambient temperature for a further 2 hours. Solvent removed in vacuo. The residue was purified by flash chromatography [Biotage SP4: 25S, flow rate 25 ml/min, gradient 20% DMAW 90 in DCM to 100% DMAW 90] and then by preparative HPLC to give N-[2-[2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindol-5-yloxy]-ethyl]-2-morpholin-4-yl-acetamide as a colourless viscous oil (40 mg, 33%). .sup.1H NMR (Me-d3-OD) 7.20 (1H, br s); 7.18 (1H, s); 6.90 (2H, br m); 6.40 (1H, s); 4.10 (2H, t); 3.73 (4H, m); 3.63 (2H, t); 3.20 (1H, m); 3.18 (2H, s); 2.60 (4H, m); 1.25 (6H, d). MS: [M+H].sup.+ 484.

Example 50

Synthesis of (2,4-dihydroxy-5-isopropylphenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone

(259) ##STR00225##

50A: Synthesis of 5-bromo-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester

(260) A mixture of 5-bromo-2,3-dihydro-1H-isoindole (1.26 g; 6.4 mmol), di-tert-butyl dicarbonate (1.53 g; 1.1 equiv.) and 4-dimethylaminopyridine (catalytic amount) in DMF (20 ml) was stirred at room temperature overnight then evaporated. The residue was partitioned between EtOAc and brine, the EtOAc layer was separated, dried (MgSO.sub.4) and evaporated. Purification by flash column chromatography using a Biotage SP4 (40S, 40 ml/m in) eluting with 0% to 5% MeOH/DCM gave 695 mg of 5-bromo-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester as a brown gum. .sup.1HNMR (DMSO-d6) 7.55 (1H, d), 7.48 (1H, d), 7.30 (1H, dd), 4.63-4.51 (4H, m), 1.46 (9H, s).

50B. Synthesis of 5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester

(261) 0.69 ml of n-Butyl lithium (2.5M solution in hexane) was added dropwise to a stirred solution of 5-bromo-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester (429 mg; 1.44 mmol) in anhydrous THF (10 ml) at −78° C. under an atmosphere of nitrogen. The reaction was stirred for 50 minutes then 1-methyl-4-piperidone (212 μl; 1.2 equiv.) was added and stirred at −78° C. for a further 60 minutes then warmed to room temperature. The reaction was quenched with saturated ammonium chloride solution then extracted with EtOAc. The EtOAc layer was washed with saturated NaHCO.sub.3, brine, dried (MgSO.sub.4) and evaporated. Purification by flash column chromatography on SiO.sub.2, gradient elution from 0% to 10% 2M methanolic ammonia/DCM gave 11 mg of 5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester as a colourless oil.

50C. Synthesis of 4-(2,3-dihydro-1H-isoindol-5-yl)-1-methyl-piperidin-4-ol

(262) A solution of 5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydroisoindole-2-carboxylic acid tert-butyl ester (107 mg; 0.32 mmol) in THF (4 ml) was treated with concentrated hydrochloric acid (1.5 ml) then heated at reflux for 4 hours, then evaporated and re-evaporated with toluene to give 4-(2,3-dihydro-1H-isoindol-5-yl)-1-methyl-piperidin-4-ol dihydrochloride as a brown gum.

50D. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone

(263) A solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (145 mg; 1.2 equiv.) in DCM (5 ml) was treated with EDC (80 mg; 1.3 equiv.) and HOAt (66 mg; 1.5 equiv.) then stirred at room temperature for 30 minutes. This solution was then added to a mixture of 4-(2,3-dihydro-1H-isoindol-5-yl)-1-methyl-piperidin-4-ol dihydrochloride (112 mg; 0.32 mmol) and triethylamine (90 μl; 2 equiv.) in THF (5 ml) and DMF (2 ml), the reaction was then stirred at room temperature overnight. The reaction mixture was diluted with EtOAc, washed with water, 1N NaOH and brine, the EtOAc layer was separated, dried (MgSO.sub.4) and evaporated. Purification by flash column chromatography on SiO.sub.2, gradient elution from 0% to 5% 2M methanolic ammonia/DCM gave 104 mg of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone as a yellow glass.

50E. Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone

(264) Hydrogenation of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone (as described in method A5) afforded 72 mg of the title compound as a cream solid. .sup.1H NMR (Me-d3-OD) 7.35 (2H, m), 7.18 (1H, br m) 7.08 (1H, s), 6.25 (1H, s), 4.78 (4H, m), 3.10 (1H, m), 2.65 (2H, m), 2.45 (2H, m), 2.25 (3H, s), 2.00 (2H, m), 1.65 (2H, m), 1.10 (6H, d). MS: [M+H].sup.+ 411.

Example 51

Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-{5-[4-(4-methyl-piperazin-1-yl)-piperidin-1-yl]-1,3-dihydro-isoindol-2-yl}methanone

(265) ##STR00226##

51A. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-bromo-1,3-dihydro-isoindol-2-yl)-methanone

(266) A solution of benzyloxy-5-isopropenyl-benzoic acid (2.85 g; 7.6 mmol), 5-bromo-2,3-dihydro-1H-isoindole (1.5 g; 1 equiv.), EDC (1.75 g; 1.2 equiv.) and HOBt (1.25 g; 1.2 equiv.) in DMF (25 ml) was stirred at room temperature overnight then evaporated. The residue was dissolved in EtOAc, washed with 2M HCl then saturated NaHCO.sub.3, dried (MgSO.sub.4) and evaporated. Purification using a Biotage SP4 (40S, 40 ml/min) eluting with 1:4-1:3-1:2 EtOAc/P.E. gave 2.45 g of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-bromo-1,3-dihydro-isoindol-2-yl)-methanone as a light brown solid.

51B. (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-{5-[4-(4-methyl-piperazin-1-yl)-piperidin-1-yl]-1,3-dihydro-isoindol-2-yl}-methanone

(267) A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-bromo-1,3-dihydro-isoindol-2-yl)-methanone (200 mg; 0.36 mmol) and 1-methyl-4-(piperidin-4-yl)piperazine (80 mg; 1.2 equiv.) in toluene (5 ml) was treated with (2-biphenyl)-di-tert-butylphosphine (6 mg; 5 mol %), tris(dibenzylidene)palladium(0) (10 mg; 2.5 mol %) and sodium tert-butoxide (50 mg; 1.4 equiv.) then heated at 120° C. for 30 minutes in a CEM explorer microwave synthesiser. The reaction mixture was diluted with DCM, washed with brine, dried (MgSO.sub.4) and evaporated. Purification by flash column chromatography (Biotage SP4-25S, 25 ml/min) eluting with DMAW 240-120-90 followed by evaporation of product containing fractions gave 105 mg of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-{5-[4-(4-methyl-piperazin-1-yl)-piperidin-1-yl]-1,3-dihydro-isoindol-2-yl}-methanone as the acetic acid salt.

51C. (2,4-dihydroxy-5-isopropyl-phenyl)-{5-[4-(4-methyl-piperazin-1-yl)-piperidin-1-yl]-1, 3-dihydro-isoindol-2-yl}-methanone hydrochloride

(268) A solution of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-{5-[4-(4-methyl-piperazin-1-yl)-piperidin-1-yl]-1,3-dihydro-isoindol-2-yl}-methanone acetic acid salt in methanol (10 ml) was treated with 10% palladium on carbon (wet), hydrogenated at room temperature and pressure overnight then filtered and evaporated. The crude compound was purified by flash column chromatograph (Biotage SP4-25S, 25 ml/min) eluting with DMAW 240-120-90-60. Product containing fractions were evaporated, treated with saturated HCl/EtOAc then evaporated and re-evaporated with methanol and dried under high vacuum at 60° C. overnight. (2,4-dihydroxy-5-isopropyl-phenyl)-5-[4-(4-methyl-piperazin-1-yl)-piperidin-1-yl]-1,3-dihydro-isoindol-2-yl)-methanone hydrochloride was isolated as a cream solid (62 mg). .sup.1H NMR (DMSO-d6) 12.40-12.00 (2H, br m), 9.75-9.55 (1H, br m), 7.45-7.05 (3H, m), 7.03 (1H, s), 6.45 (1H, s), 4.70-4.55 (4H, m), 3.85-3.65 (6H, m), 3.60-3.40 (5H, m), 3.15-3.05 (1H, m), 3.0-2.78 (5H, m), 2.30-2.20 (2H, m), 2.05-1.90 (2H, m), 1.15 (6H, d). MS: [M+H].sup.+ 479.

Example 52

Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-piperazin-1-yl-phenyl)-1,3-dihydro-isoindol-2-yl]-methanone

(269) ##STR00227##

52A. Synthesis of 4-{4-[2-(2,4-bis-benzyloxy-5-isopropenyl-benzoyl)-2,3-dihydro-1H-isoindol-5-yl]-phenyl}piperazine-1-carboxylic acid tert-butyl ester

(270) A mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-bromo-1,3-dihydro-isoindol-2-yl)-methanone (240 mg, 0.43 mmol), t-butyl-4-[4-(4,4,5,5-tetramethyl-1,3,2-dioxaborolan-2-yl)phenyl]piperazine carboxylate (210 mg, 1.25 equiv.), bis(tri-t-butylphosphine)palladium(0) (12.5 mg, 2.5 mol %) and potassium carbonate (350 mg, 6 equiv.) in toluene/water/ethanol (1 ml: 1 ml: 4 ml) was heated at 135° C. for 30 minutes in a CEM explorer microwave synthesiser. The reaction mixture was diluted with EtOAc, washed with saturated NaHCO.sub.3, dried (MgSO.sub.4) and evaporated. Purification by flash column chromatography (Biotage SP4-25S, 25 ml/min) eluting with 1:3 then 1:1 EtOAc/P.E. Evaporation of product containing fractions gave 85 mg of 4-{4-[2-(2,4-bis-benzyloxy-5-isopropenyl-benzoyl)-2,3-dihydro-1H-isoindol-5-yl]-phenyl}piperazine-1-carboxylic acid tert-butyl ester. MS: [M+H].sup.+ 736.

52B. Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-piperazin-1-yl-phenyl)-1,3-dihydro-isoindol-2-yl]-methanone

(271) Hydrogenation of 4-(4-[2-(2,4-bis-benzyloxy-5-isopropenyl-benzoyl)-2,3-dihydro-1H-isoindol-5-yl]-phenyl)piperazine-1-carboxylic acid tert-butyl ester (as described in method A5), followed by BOC deprotection (as described in example 70) afforded 10 mg of the title compound as the hydrochloride salt after flash column chromatography (Biotage SP4, 25S) eluting with DMAW 240-120-90 and evaporation from saturated HCl/EtOAc. .sup.1H NMR (Me-d3-OD) 7.63 (2H, d), 7.55 (2H, m) 7.45-7.30 (1H, m), 7.25 (1H, s), 7.20 (2H, d), 5.03 (4H, m), 3.55 (4H, m), 3.47 (4H, m), 3.23 (1H, m), 1.25 (6H, d). MS: [M+H].sup.+ 458.

Example 53

Synthesis of 2,4-dihydroxy-5-isopropyl-phenyl)-[5-(1-dimethylamino-2-hydroxy-ethyl)-1,3-dihydro-isoindol-2-yl]-methanone, and dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-1-hydroxy-ethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(272) ##STR00228##

53A. Synthesis of 5-bromo-2,4-dimethoxybenzoic acid methyl ester

(273) A solution of 5-bromo-2,4-dihydroxybenzoic acid (24.9 g, 107 mmol) in acetone (355 ml), was treated with methyl iodide (39.9 ml, 640 mmol) and K.sub.2CO.sub.3 (88 g, 640 mmol) then heated at reflux overnight. The salts were filtered off and washed with acetone. The filtrate was evaporated to dryness and the product was purified by flash column chromatography (100% DCM) to yield 5-bromo-2,4-dimethoxybenzoic acid methyl ester as a colourless solid (28 g). .sup.1H NMR (Me-d.sub.3-OD) 7.98 (1H, s), 6.74 (1H, s), 3.99 (3H, s), 3.94 (3H, s), 3.85 (3H, s). MS: [M+H].sup.+ 275/277.

53B. Synthesis of -isopropenyl-2,4-dimethoxy-benzoic acid methyl ester

(274) To potassium isopropylidene trifluoroborate (4.87 g, 32.7 mmol) and 5-bromo-2,4-dimethoxybenzoic acid methyl ester (7.5 g, 27.3 mmol) in THF (195 ml) was added Cs.sub.2CO.sub.3 (26.6 g, 81.8 mmol) in water (39 ml). The reaction was degassed and Pd(PPh.sub.3).sub.4 (1.58 g, 1.36 mmol) added. The reaction was heated at reflux for three days then quenched by adding water and extracted with EtOAc (×2). The combined organic layers were washed with brine, dried (MgSO.sub.4), filtered and evaporated to leave an orange solid. The product was taken up in EtOAc again and the precipitate filtered. The filtrate was evaporated to dryness to yield 5-isopropenyl-2,4-dimethoxy-benzoic acid methyl ester (6.2 g). .sup.1H NMR (Me-d.sub.3-OD) 7.68 (1H, s), 6.66 (1H, s), 5.10-5.08 (1H, m), 5.02-5.00 (1H, m), 3.93 (3H, s), 3.92 (3H, s), 3.84 (3H, s), 2.08-2.06 (3H, m). MS: [M+H].sup.+ 237.

53C. Synthesis of 5-isopropyl-2,4-dimethoxy-benzoic acid methyl ester

(275) A solution of 5-isopropenyl-2,4-dimethoxy-benzoic acid methyl ester (6.0 g, 25.4 mmol) in MeOH (85 ml) was shaken with 10% Pd/C under an atmosphere of H.sub.2 at room temperature for 3 hours. The catalyst was filtered through GF/A paper but a little fine powder passed through. The filtrate was passed through a small pad of silica and evaporated to dryness to yield a colourless solid. The product was purified by flash column chromatography (DCM:Petrol gradient elution) to yield 5-isopropyl-2,4-dimethoxy-benzoic acid methyl ester a colourless solid (5.5 g). .sup.1H NMR (Me-d.sub.3-OD) 7.68 (1H, s), 6.64 (1H, s), 3.94 (3H, s), 3.91 (3H, s), 3.84 (3H, s), 3.23 (1H, sept), 1.20 (6H, d). MS: [M+H].sup.+ 239.

53D. Synthesis of 5-isopropyl-2,4-dimethoxy-benzoic acid

(276) 5-isopropyl-2,4-dimethoxy-benzoic acid methyl ester (5.5 g, 23.1 mmol) and NaOH (1.38 g, 34.6 mmol) in THF (46 ml) and water (46 ml) was warmed to 50° C. overnight. The reaction was cooled and diluted with water and EtOAc. The aqueous layer was neutralized with HCl (1N, aq.). The product was extracted with EtOAc (×3) and the combined organic layers were washed with brine and dried over MgSO.sub.4. The product was filtered and evaporated to dryness to yield 5-isopropyl-2,4-dimethoxy-benzoic acid as a pale peach solid (4.7 g). .sup.1H NMR (DMSO-d.sub.6) 12.1 (1H, br s), 7.62 (1H, s), 6.71 (1H, s), 3.95 (3H, s), 3.91 (3H, s), 3.19 (1H, sept), 1.18 (6H, d). MS: [M+H].sup.+ 225.

(277) ##STR00229##

53E. Synthesis of (5-bromo-1,3-dihydro-isoindol-2-yl)-(5-isopropyl-2,4-dimethoxyphenyl)methanone

(278) To a mixture of 5-isopropyl-2,4-dimethoxybenzoic acid (2.45 g, 10.9 mmol), HOBt (1.61 g, 11.9 mmol) and EDC (1.85 g, 11.9 mmol) in anhydrous DMF (33 ml) under N.sub.2 was added 5-bromo-2,3-dihydro-1H-isoindole (1.97 g, 9.95 mmol) and stirred at room temperature overnight. The reaction was quenched by diluting with NaOH (1M, aq.) and extracting the product with EtOAc (×2). The combined organic layers were washed with brine and dried over MgSO.sub.4. The product was filtered and evaporated to dryness to leave a brown oil. The product was purified by flash column chromatography using gradient elution (ether/petrol) to yield (5-bromo-1,3-dihydro-isoindol-2-yl)-(5-isopropyl-2,4-dimethoxyphenyl)-methanone as a beige solid (3 g). .sup.1H NMR (Me-d.sub.3-OD) 7.60-7.13 (3H, m), 7.14 (1H, s), 6.71 (1H, s), 4.89 (2H, d), 4.64 (2H, d), 3.93 (3H, s), 3.90 (3H, s), 3.27 (1H, sept), 1.20 (6H, d). MS: [M+H].sup.+ 404/406.

53F. Synthesis of 5-isopropyl-2,4-dimethoxy-phenyl)-(5-vinyl-1,3-dihydro-isoindol-2-yl)-methanone

(279) To (5-bromo-1,3-dihydro-isoindol-2-yl)-(5-isopropyl-2,4-dimethoxyphenyl)methanone (2.2 g, 5.44 mmol), and 2-vinyl-4,4,5,5-tetramethyl-1,3,2-dioxaborolane (1.2 ml, 6.53 mmol) in MeOH (25 ml) and toluene (25 ml) was added Na.sub.2CO.sub.3 in water (25 ml). The reaction was degassed, Pd(PPh.sub.3).sub.4 (0.38 g, 0.05 mmol) added then heated at 80° C. overnight. The reaction was worked up by adding water and extracting with EtOAc (×3). The combined organic layers were washed with brine and dried over MgSO.sub.4. The product was filtered and evaporated to dryness then purified by flash column chromatography, gradient elution (ether:petrol) to yield 5-isopropyl-2,4-dimethoxy-phenyl)-(5-vinyl-1,3-dihydro-isoindol-2-yl)-methanone as a yellow oil (1.6 g). .sup.1H NMR (Me-d.sub.3-OD) 7.47-7.15 (3H, m), 7.15 (1H, s), 6.82-6.72 (1H, m), 6.71 (1H, s), 5.79 (1H, dd), 5.24 (1H, dd), 4.90 (2H, d), 4.64 (2H, d), 3.93 (3H, s), 3.91 (3H, s), 3.27 (1H, sept), 1.23 (6H, d). MS: [M+H].sup.+ 352.

53G. Synthesis of (5-isopropyl-2,4-dimethoxy-phenyl)-(5-oxiranyl-1,3-dihydro-isoindol-2-yl)-methanone

(280) To (5-isopropyl-2,4-dimethoxy-phenyl)-(5-vinyl-1,3-dihydro-isoindol-2-yl)-methanone (0.80 g, 2.28 mmol) in DCM (22 ml) was added mCPBA (0.61 g, 2.73 mmol) at 0° C. The reaction was stirred at room temperature for an hour. The reaction was diluted with NaOH (1M, aq.) and extracted the product with EtOAc. The EtOAc layer was washed with NaOH again. The organic layer was washed with brine and dried over MgSO.sub.4. The product was filtered and evaporated to dryness to yield crude (5-isopropyl-2,4-dimethoxy-phenyl)-(5-oxiranyl-1,3-dihydro-isoindol-2-yl)-methanone as a very pale yellow oil. MS: [M+H].sup.+ 368.

(281) ##STR00230##

53H. Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(1-dimethylamino-2-hydroxy-ethyl)-1,3-dihydro-isoindol-2-yl]-methanone (Compound 121H-i) and (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-1-hydroxy-ethyl)-1,3-dihydro-isoindol-2-yl]-methanone (Compound 121H-ii)

(282) (5-isopropyl-2,4-dimethoxy-phenyl)-(5-oxiranyl-1,3-dihydro-isoindol-2-yl)-methanone (˜120 mg, crude) was dissolved in dimethylamine in EtOH (20 ml, ˜33%, 5.6 M) and heated at 60° C. overnight. The reaction was evaporated to dryness and the product crudely purified by flash column chromatography MeOH:DCM (1:5) to yield impure material which was used without further purification. To a mixture of [(5-(1-dimethylamino-2-hydroxy-ethyl)-1,3-dihydro-isoindol-2-yl]-(5-isopropyl-2,4-dimethoxy-phenyl)-methanone and [5-(2-dimethylamino-1-hydroxy-ethyl)-1,3-dihydro-isoindol-2-yl]-(5-isopropyl-2,4-dimethoxy-phenyl)-methanone (˜100 mg) was added DCM (5 ml) and then boron tribromide (3 eq.) under N.sub.2. The reaction was left to stir at room temperature until completion. The reaction was quenched with ice and diluted with water and EtOAc. The aqueous layer was extracted with EtOAc (×2). The combined organic layers were washed with brine, dried over MgSO.sub.4 then filtered and evaporated to dryness, to leave a yellow residue which was purified by preparative HPLC to yield the two resorcinol isomers.

(283) (2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(1-dimethylamino-2-hydroxy-ethyl)-1,3-dihydro-isoindol-2-yl]-methanone, (Compound 121H-i) .sup.1H NMR (Me-d.sub.3-OD) 7.42-7.30 (3H, m), 7.19 (1H, s), 6.39 (1H, s), 4.98-4.87 (4H, m), 4.03-3.97 (1H, m), 3.94-3.86 (1H, m), 3.68 (1H, br s), 3.22 (1H, sept), 2.40 (6H, s), 1.23 (6H, d). MS: [M+H].sup.+ 384.

(284) (2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-1-hydroxy-ethyl)-1,3-dihydro-isoindol-2-yl]-methanone, (Compound 121H-ii) .sup.1H NMR (Me-d.sub.3-OD) 7.39-7.25 (3H, m), 7.18 (1H, s), 6.38 (1H, s), 6.94-6.88 (5H, m), 3.22 (1H, sept), 2.77-2.68 (1H, m), 2.61-2.51 (1H, m), 2.42 (6H, s), 1.23 (6H, d). MS: [M+H].sup.+ 384.

Example 54

Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(piperazine-1-carbonyl)-1,3-dihydro-isoindol-2-yl]-methanone hydrochloride

(285) ##STR00231##

54A. Synthesis of 4-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carbonyl]-piperazine-1-carboxylic acid tert-butyl ester

(286) A solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid (Preparation D6) (0.5 g, 0.96 mmol), EDC (0.22 g, 1.15 mmol), HOBT (0.196 g, 1.15 mmol) and BOC piperazine (0.117 ml, 1.06 mmol) in DMF (10 ml) was stirred at room temperature for 48 hours, then evaporated under vacuum. The crude material was dissolved in ethyl acetate and extracted twice with saturated NaHCO.sub.3, organics washed with brine, dried (MgSO.sub.4), filtered then evaporated under vacuum and purified by flash column chromatography (80% EtOAc-P.E. as eluant) to give 0.5 g of 4-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carbonyl]-piperazine-1-carboxylic acid tert-butyl ester. MS: [M+H].sup.+ 688.

54B. Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(piperazine-1-carbonyl)-1,3-dihydro-isoindol-2-yl]-methanone hydrochloride

(287) Hydrogenation as Method A5 to give (0.2 g, 0.30 mmol) 4-[2-(2,4-dihydroxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carbonyl]-piperazine-1-carboxylic acid tert-butyl ester [used crude] dissolved in EtOAc then treated with saturated EtOAc/HCl, stirred at ambient for 3 hours, reaction diluted with ether, solid filtered to give 0.19 g of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(piperazine-1-carbonyl)-1,3-dihydro-isoindol-2-yl]-methanone hydrochloride. .sup.1H NMR (Me-d.sub.3-OD) 7.50-7.42 (3H, m), 7.18 (1H, s), 6.39 (1H, s), 5.00-4.95 (4H, br s), 3.92-3.79 (4H, br s), 3.35-3.28 (4H, br s), 3.26-3.15 (1H, m), 1.23 (6H, d). MS: [M+H].sup.+ 410.

Example 55

Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(288) ##STR00232##

55A. Synthesis of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid methoxy-methyl-amide

(289) A solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid (Preparation D6) (1.76 g, 3.39 mmol), EDC (0.78 g, 4.06 mmol), HOBT (0.55 g, 4.06 mmol), Et.sub.3N (1 ml, 6.78 mmol) and N,O-dimethylhydroxylamine hydrochloride (0.36 g, 3.72 mmol) in DMF (20 ml) was stirred at room temperature for 48 hours, then evaporated under vacuum. The crude material was dissolved in ethyl acetate and extracted twice with saturated NaHCO.sub.3, organics washed with brine, dried (MgSO.sub.4), filtered then evaporated to give 1.84 g of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid methoxy-methyl-amide. MS: [M+H].sup.+ 563.

55. Synthesis of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carbaldehyde

(290) A solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carboxylic acid methoxy-methyl-amide (0.226 g, 0.4 mmol) in THF (5 ml) cooled to 0° C., treated with 1M LiAlH.sub.4/THF (0.3 ml, 0.3 mmol), stirred 1 hour, further LiAlH.sub.4 (0.05 ml) added then stirred for 30 minutes. The reaction was quenched with saturated KHSO.sub.4 solution, extracted with EtOAc, dried (MgSO.sub.4), filtered and evaporated to give 0.2 g of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole 5-carbaldehyde. MS: [M+H].sup.+ 504.

55C. Synthesis of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(291) To a solution of 2-(2,4-bis-benzyloxy-5-isopropyl-benzoyl)-2,3-dihydro-1H-isoindole-5-carbaldehyde (0.316 g, 0.63 mmol) and n-methyl piperazine (63 mg, 0.63 mmol) in CH.sub.2Cl.sub.2 (10 ml) was added AcOH (38 mgs 0.63 mmol) and NaBH(OAc).sub.3 (0.28 g, 1.33 mmol), then stirred at ambient for 5 hours. The reaction was quenched with water, layers separated and aqueous washed CH.sub.2Cl.sub.2. The organics were combined, washed with brine, dried (MgSO.sub.4), filtered and evaporated to give 0.32 g of (2,4-bis-benzyloxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone. MS: [M+H].sup.+ 588.

55D. Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(292) Hydrogenation was carried out using Method A5 but with the addition of K.sub.2CO.sub.3 (2 equiv.) in a MeOH/H.sub.2O [9.1]. After evaporation of methanol the reaction was diluted with water, neutralised using 1M HCl and extracted with CH.sub.2Cl.sub.2 (×2). Organics dried (MgSO.sub.4), filtered and evaporated under vacuum then purified by preparative HPLC to give 21 mg of (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone. MS: [M+H].sup.+ 410. .sup.1H NMR (Me-d.sub.3-OD) 7.37-7.23 (3H, br s), 7.19 (1H, s), 6.39 (1H, s), 4.94-4.87 (4H, br s), 3.57 (2H, s), 3.27-3.16 (1H, m), 2.67-2.39 (8H, m), 2.31 (3H, s), 1.23 (6H, d).

Example 56

Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]methanone

56A. Synthesis of 4-hydroxyisoindoline hydrobromide

(293) ##STR00233##

(294) A suspension of dimethyl 3-methoxyphthalate (69.45 g, 0.31 mol) [prepared as per J. Chem. Soc., Perkin Trans. 1, 1989, 391] in water (300 ml) was treated with potassium hydroxide (43.7 g, 0.78 mol) and the mixture was stirred and held at reflux for 4 hours. Upon cooling to room temperature the methanol liberated during the course of the reaction was removed in vacuo, the mixture acidified to pH 2 or below by the addition of 5M hydrochloric acid and evaporated gently in vacuo to induce crystallization. The solid material was filtered off, washed with a little ice cooled water, sucked dry under reduced pressure and dried in a vacuum oven at 50° C. overnight to afford 3-methoxyphthalic acid (51.0 g, 84%) as a colourless solid. .sup.1H NMR (DMSO-d.sub.6) 13.05 (2H, br s), 7.48 (2H, m), 7.33 (1H, m), 3.82 (3H, s). MS: [M+H].sup.+ 197.

(295) Acetic anhydride (70 ml) was added to a mixture of 3-methoxyphthalic acid (51.0 g, 0.26 mol) in anhydrous tetrahydrofuran (250 ml) and the mixture was stirred and held at reflux for 4 hours. Upon cooling to room temperature the solvent was removed in vacuo and the resulting solid material was dried in a vacuum oven at 50° C. overnight to afford 3-methoxyphthalic anhydride (45.9 g, 99%) as a colourless solid. .sup.1H NMR (DMSO-d.sub.6) 7.97 (1H, dd), 7.63 (1H, d), 7.60 (1H, d), 4.02 (3H, s). MS: [M+H].sup.+ 179.

(296) A mixture of 3-methoxyphthalic anhydride (24.0 g, 134.8 mmol) and formamide (120 ml) was stirred and held at 210° C. for 5 hours and was then allowed to cool to room temperature overnight. Water (100 ml) was added and the solid material filtered off under reduced pressure. The crude product was washed sequentially with 50% aqueous acetone (50 ml) and diethyl ether (200 ml) and sucked dry under reduced pressure to afford 3-methoxyphthalimide (8.95 g, 37%) as an off-white solid. .sup.1H NMR (DMSO-d.sub.6) 11.08 (1H, br s), 7.78 (1H, dd), 7.45 (1H, d), 7.36 (1H, d), 3.93 (3H, s). MS: [M+H].sup.+ 178.

(297) A stirred solution of 3-methoxyphthalimide (8.95 g, 50.56 mmol) in anhydrous tetrahydrofuran (200 ml) at 0° C. was treated dropwise with a solution of borane in tetrahydrofuran (1M, 150 ml, 0.15 mol) and the resulting mixture was stirred and held at reflux for 16 hours. The mixture was cooled to 0° C., methanol (60 ml) was added dropwise followed by 5M hydrochloric acid (60 ml) and the mixture was stirred and held at reflux for 4 hours. Upon cooling to room temperature the organic solvent was removed in vacuo and the mixture diluted with water (250 ml) and extracted with dichloromethane (3×250 ml). The aqueous layer was basified to pH 12 or above by the addition of 5M sodium hydroxide, extracted with dichloromethane (3×250 ml) and the combined extracts were evaporated to dryness in vacuo to afford 4-methoxyisoindoline (4.44 g, 59%) as a green oil which was used without further purification. .sup.1H NMR (DMSO-d.sub.6) 7.18 (1H, t), 6.83 (1H, d), 6.78 (1H, d), 4.07 (2H, s), 4.02 (2H, s), 3.78 (3H, s). MS: [M+H].sup.+ 150.

(298) 4-Methoxyisoindoline (4.4 g, 29.53 mmol) in 48% aqueous hydrobromic acid (50 ml) was stirred and held at reflux for 16 hours. Upon cooling to room temperature the solvent was removed in vacuo to afford 4-hydroxyisoindoline hydrobromide (5.0 g, 78%) as a pale orange solid. .sup.1H NMR (DMSO-d.sub.6) 9.95 (1H, br s), 9.37 (2H, br s), 7.19 (1H, t), 6.84 (1H, d), 6.80 (1H, d), 4.48 (2H, t), 4.40 (2H, t). MS: [M+H].sup.+ 136.

56B. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(4-hydroxy-1,3-dihydro-isoindol-2-yl)-methanone

(299) ##STR00234##

(300) A mixture of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (8.1 g, 21.65 mmol), 4-hydroxylsoindoline hydrobromide (4.91 g, 22.73 mmol), N-ethyl-N′-(3-dimethylaminopropyl)carbodiimide hydrochloride (5.0 g, 25.98 mmol), 1-hydroxybenzotriazole (3.5 g, 25.98 mmol) and triethylamine (6 ml, 43.3 mmol) in N,N-dimethylformamide (50 ml) was stirred at room temperature for 16 hours. The solvent was removed in vacuo and the residue was treated with a saturated aqueous solution of sodium hydrogen carbonate (200 ml). The mixture was filtered, the solid material was washed copiously with water, sucked dry under reduced pressure and dried in a vacuum oven at 50° C. overnight to afford (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(4-hydroxy-1,3-dihydro-isoindol-2-yl)-methanone (10.25 g, 96%) as a pale tan solid. .sup.1H NMR (DMSO-d.sub.6) (mixture of amide rotamers) 9.68 and 9.60 (1H, 2×br s), 7.45-7.25 (10H, m), 7.20-7.00 (3H, m), 6.82 and 6.72 (1H, 2×d), 6.68 (1H, m), 5.23 and 5.22 (2H, 2×s), 5.18 (2H, s), 5.11 (1H, s), 5.09 (1H, s), 4.77 and 6.67 (2H, 2×s), 4.53 and 4.44 (2H, 2×s), 2.04 (3H, s). MS: [M+H].sup.+ 492.

56C. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]-methanone

(301) A mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(4-hydroxy-1,3-dihydro-isoindol-2-yl)-methanone (2 g; 4.07 mmol), 4-(3-chloropropyl)morpholine (1.66 g; 2.5 equiv.) and caesium carbonate (8.3 g; 6.25 equiv) in DMF was heated at 90° C. overnight then evaporated. The residue was dissolved in EtOAc, washed with brine, dried (MgSO.sub.4) and evaporated. Purification of the crude material using a Biotage SP4 (40S, 40 ml/min), using gradient elution form 0% to 10% MeOH/EtOAc gave 1.8 g of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]-methanone as a pale yellow gum. MS: [M+H].sup.+ 619.

56D. Synthesis of (2,4-dihydroxy-5-isopropyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]methanone

(302) ##STR00235##

(303) Hydrogenation of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[4-(3-morpholin-4-yl-propoxy)-1,3-dihydro-isoindol-2-yl]-methanone (as described in method A5) followed by treatment with saturated HCl/EtOAc and trituration with hot acetone afforded 890 mg of the title compound (hydrochloride salt) as a cream solid. .sup.1H NMR (DMSO-d.sub.6) 10.78 (1H, br s), 10.05 (1H, br s), 9.55 (1H, br s), 7.30 (1H, t), 7.08 (1H, s) 6.98-6.90 (2H, m), 6.45 (1H, s), 4.80 (2H, s), 4.75 (2H, s), 4.15 (2H, t), 3.95 (2H, br m), 2.80 (2H, br m), 3.50-3.35 (2H, br m), 3.25 (2H, br m,), 3.18-3.02 (3H, br m), 2.20 (2H, br m), 1.15 (6H, d). MS: [M+H].sup.+ 441.

Examples 57 to 74

(304) By following the methods described above, the following compounds were prepared.

(305) TABLE-US-00017 Exam- ple Number Compound Chemical Name Method N.M.R. Data M.S. 57 [5-(2-Amino- ethoxy)-1,3- dihydro-isoindol- 2-yl]-(2,4- dihydroxy-5- isopropyl- phenyl)- methanone As for Example 34, A2 (from 2,4-bis- benzyloxy-5- isopropenyl- benzoic acid (Preparation B5) and 5- hydroxy- isoindoline), alkylation using 3-(BOC- amino)ethyl tosylate, then A5. Final BOC deprotection using saturated HCl/EtOAC (Example 18). .sup.1H NMR (Me- d.sub.3-OD) 8.55 (1 H, s), 7.30- 7.20 (1 H, m), 7.15 (1 H, s), 7.05-6.95 (2 H, m), 6.40 (1 H, s), 4.95-4.80 (4 H, m) 4.25 (2 H, t), 3.35 (2 H, t), 3.25- 3.15 (1 H, m), 1.25 (6 H, d) MS: [M + H].sup.+ 357 58 (2,4-Dihydroxy-5- isopropyl- phenyl)-(5- hydroxy-1,3- dihydro-isoindol- 2-yl)-methanone Isolated as a bi-product from synthesis of Example 57. .sup.1H NMR (Me- d.sub.3-OD) 7.20 (1 H, s), 7.15- 7.05 (1 H, m), 6.80- 6.70 (2 H, m), 6.40 (1 H, s), 4.95-4.80 (4 H, m), 3.25-3.15 (1 H, m), 1.25 (6 H, d) MS: [M + H].sup.+ 314 59 (2,4-Dihydroxy-5- isopropyl- phenyl)-{5-[4-(2- hydroxy-ethyl)- piperazin-1-yl]- 1,3-dihydro- isoindol-2-yl}- methanone As for Example 51, using N-(2- hydroxyethyl)- piperazine in the Buchwald reaction. .sup.1H NMR (DMSO-d.sub.6) 10.40 (1 H, br s), 9.65 (1 H, br s), 7.40-7.15 (1 H, m), 7.05 (1 H, s), 7.05- 6.90 (2 H, m), 6.45 (1 H, s), 4.80-4.60 (4 H, m), 3.85-3.70 (4 H, m), 3.65- 3.55 (2 H, m), 3.25-3.05 (7 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 426 60 (2,4-Dihydroxy-5- isopropyl- phenyl)-[5-(4- morpholin-4-yl- piperidin-1-yl)- 1,3-dihydro- isoindol-2-yl]- methanone As for Example 51, using 4- morpholino- piperidine in the Buchwald reaction. .sup.1H NMR (DMSO-d.sub.6) 11.10 (1 H, br s), 9.65 (1 H, br s), 7.30-7.05 (3 H, m), 7.03 (1 H, s), 6.45 (1 H, s), 4.80- 4.65 (4 H, m), 4.0-3.95 (2 H, m), 3.90-3.75 (4 H, m), 3.50- 3.40 (2 H, m), 3.40-3.30 (1 H, m), 3.15-3.03 (3 H, m), 2.90- 2.75 (2 H, m), 2.25-2.15 (2 H, m), 1.95-1.80 (2 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 466 61 0 (2,4-Dihydroxy-5- isopropyl- phenyl)-[5-(1- methyl-piperidin- 4-ylamino)-1,3- dihydro-isoindol- 2-yl]-methanone As for Example 51, using 4- amino-1- methyl- piperidine in the Buchwald reaction. .sup.1H NMR (DMSO-d.sub.6) 10.60 (1 H, br s), 9.65 (1 H, br s), 7.20 (1 H, m), 7.03 (1 H, s), 6.95-6.80 (2 H, m), 6.45 (1 H, s), 4.80- 4.65 (4 H, m), 3.45 (2 H, m), 3.25 (1 H, m), 3.10 (1 H, m), 3.00 (2 H, m), 2.70 (3 H, d), 2.15-2.05 (2 H, m), 1.90-1.75 (2 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 410 62 (2,4-Dihydroxy-5- isopropyl- phenyl)-[5-(4- isopropyl- piperazin-1-yl)- 1,3-dihydro- isoindol-2-yl]- methanone As for Example 51, using i- propyl- piperazine in the Buchwald reaction. .sup.1H NMR (DMSO-d.sub.6) 10.70 (1 H, br s), 9.65 (1 H, br s), 7.25-7.10 (1 H, m), 7.05 (1 H, s), 7.00- 6.90 (2 H, m), 6.45 (1 H, s), 4.80-4.60 (4 H, m), 3.80 (2 H, m), 3.55-3.40 (3 H, m), 3.23- 3.05 (5 H, m), 1.33 (6 H, d), 1.15 (6 H d) MS: [M + H].sup.+ 424 63 (2,4-Dihydroxy-5- isopropyl- phenyl)-(5- piperazin-1-yl- 1,3-dihydro- isoindol-2-yl)- methanone As for Example 51, using Boc- piperazine in the Buchward reaction. Boc deprotection using saturated HCl/dioxane (Example 18). .sup.1H NMR (DMSO-d.sub.6) 9.70 (1 H, br s), 9.25 (2 H, br s), 7.23 (1 H, br m), 7.05 (1 H, s), 7.00-6.90 (2 H, m), 6.45 (1 H, s), 4.80-4.60 (4 H, m), 3.35 (4 H, m), 3.20 (4 H, m), 3.10 (1 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 382 64 4-[2-(2,4- Dihydroxy-5- isopropyl- benzoyl)-2,3- dihydro-1H- isoindol-5- ylamino]- piperidine-1- carboxylic acid tert-butyl ester As for Example 51, using 1- Boc-4-amino- piperidine in the Buchwald reaction. .sup.1H NMR (Me- d.sub.3-OD) 7.20 (1 H, s), 7.05 (1 H, m), 6.65- 6.55 (2 H, m), 6.35 (1 H, s), 4.85-4.75 (4 H, m), 4.05 (2 H, m), 3.50 (1 H, m), 3.20 (1 H, m), 3.00 (2 H, m), 2.00 (2 H, m), 1.5 (9 H, s), 1.30 (2 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 496 65 (2,4-dihydroxy-5- isopropyl- phenyl)-[5- (piperidin-4- ylamino)-1,3- dihydro-isoindol- 2-yl]-methanone BOC deprotection using saturated HCl/EtOAC (Example 18). .sup.1H NMR (DMSO-d.sub.6) 7.05 (1 H, s), 7.00 (1 H, m), 6.55- 6.45 (2 H, m), 6.40 (1 H, s), 4.70-4.60 (4 H, m), 3.25 (1 H, m), 3.10 (1 H, m), 2.95 (2 H, m), 2.45 (2 H, m), 1.85 (2 H, m), 1.75 (3 H, s), 1.20 (2 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 396 66 (2,4-dihydroxy-5- isopropyl- phenyl)-[4-(4- methyl-piperazin- 1-yl)-1,3-dihydro- isoindol-2-yl]- methanone As for Example 51, using (2,4- bis-benzyloxy- 5-isopropenyl- phenyl)-(4- bromo-1,3- dihydro- isoindol-2-yl)- methanone (Prep: A2 between 2,4- bis-benzyloxy- 5-isopropenyl- benzoic acid and 4-bromo- 1,3-dihydro-1H- isoindoline) and N-methyl- piperazine in the Buchwald reaction. .sup.1H NMR (Me- d.sub.3-OD) 7.35- 7.18 (2 H, m), 7.10-6.95 (2 H, m), 6.95-6.85 (2 H, m), 6.40 4.85 (4 H, m), 3.25 (1 H, m), 3.20-3.05 (4 H, m), 3.05-2.80 (4 H, m), 2.60 (3 H, m), 2.00 (3 H, s), 1.25 (6 H, d) MS: [M + H].sup.+ 396 67 (2,4-dihydroxy-5- isopropyl- phenyl)-[4- (piperidin-4- ylamino)-1,3- dihydro-isoindol- 2-yl]-methanone As for Example 65, using 1- Boc-4-amino- piperidine in the Buchwald reaction, followed by Boc deprotection using saturated HCl/EtOAc (Example 18). .sup.1H NMR (DMSO-d.sub.6) 7.05 (1 H, s), 7.00 (1 H, m), 6.55- 6.45 (2 H, m), 6.40 (1 H, s), 4.70-4.60 (4 H, m), 3.25 (1 H, m), 3.10 (1 H, m), 2.95 (2 H, m), 2.45 (2 H, m), 1.85 (2 H, m), 1.75 (3 H, s), 1.20 (2 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 396 68 (2,4-Dihydroxy-5- isopropyl- phenyl)-(5- dimethylamino- methyl-1,3dihydro- isoindol-2-yl)- methanone A2 and A5. From 2,4- bis-benzyloxy- 5-isopropenyl- benzoic acid (B5, and (2,3- dihydro-1H- isoindol-5- ylmethyl)- dimethyl-amine (Preparation A1) .sup.1H NMR (Me- d.sub.3-OD) 7.26- 7.12 (3 H, m), 7.07 (1 H, s), 6.27 (1 H, s), 4.85-4.77 (4 H, br s), 3.40 (2 H, s), 3.15-3.05 (1 H, m), 2.15 (6 H, s), 1.11 (6 H, d) MS: [M + H].sup.+ 355 69 (2,4-Dihydroxy-5- isopropyl- phenyl)-[5-(4- methyl- piperazine-1- carbonyl)-1,3- dihydro-isoindol- 2-yl]-methanone A2 and A5. From 2- (2,4-bis- benzyloxy-5- isopropyl- benzoyl)-2,3- dihydro-1H- isoindole-5- carboxylic acid (D6) and N- methyl piperazine .sup.1H NMR (Me- d.sub.3-OD) 7.60- 7.38 (3 H, m), 7.19 (1 H, s), 6.39 (1 H, s), 4.96 (4 H, m), 3.85-3.71 (2 H, br s), 3.54-3.4 (2 H, br s), 3.26- 3.15 (1 H, m), 2.59-2.39 (4 H, br d), 2.34 (3 H, s), 1.23 (6 H, d) MS: [M + H].sup.+ 424 70 (2,4-Dihydroxy-5- isopropyl- phenyl)-{5-[2- (2,2-dimethyl- propylamino)- ethoxy]-1,3- dihydro-isoindol- 2-yl}-methanone As for the synthesis of (2,4-dihydroxy- 5-isopropyl- phenyl)-[5-(2- isopropylamino- ethoxy)-1,3- dihydro- isoindol-2-yl]- methanone except using trimethyl acetaldehyde instead of acetone. Purified by preparative HPLC. .sup.1H NMR (Me- d.sub.3-OD) 7.28 (1 H, br s); 7.20 (1 H, s); 7.00 (2 H, br m); 6.40 (1 H, s); 4.35 (2 H, t); 3.50 (2 H t); 3.20 (1 H, m); 3.00 (2 H, s); 1.23 (6 H, d); 1.10 (9 H, s) MS: [M + H].sup.+ 427 71 0 [5-(2-Cyclopentyl- amino-ethoxy)- 1,3-dihydro- isoindol-2-yl]- (2,4-dihydroxy-5- isopropyl- phenyl)- methanone As for the synthesis of (2,4-dihydroxy- 5-isopropyl- phenyl)-[5-(2- isopropylamino- ethoxy)-1,3- dihydro- isoindol-2-yl]- methanone except using cyclopentanone instead of acetone. Purified by preparative HPLC. .sup.1H NMR (DMSO-d.sub.6) 10.05 (1 H, br s); 9.60 (1 H, br s); 7.23 (1 H, br s); 7.05 (1 H, s); 6.95 (1 H, br s); 6.88 (1 H, br d); 6.40 (1 H, s); 4.72 (4 H, br m); 4.02 (2 H, t); 3.10 (2 H, m); 2.93 (2 H, t); 1.78 (2 H, m); 1.63 (2 H, m); 1.48 (2 H, m); 1.35 (2 H, m); 1.15 (6 H, d) MS: [M + H].sup.+ 425 72 (2,4-Dihydroxy-5- isopropyl- phenyl)-(5- piperidin-1- ylmethyl-1,3- dihydro-isoindol- 2-yl)-methanone As for the synthesis of (2,4-dihydroxy- 5-isopropyl- phenyl)-[5-(4- methyl- piperazin-1- ylmethyl)-1,3- dihydro- isoindol-2-yl]- methanone (Example 56) except using piperidine instead of N- methyl- piperazine. .sup.1H NMR (Me- d.sub.3-OD) 7.35- 7.24 (3 H, m), 7.19 (1 H, s), 6.39 (1 H, s), 4.94-4.49 (4 H, br s), 3.54 (2 H, s), 3.27-3.18 (1 H, m), 2.51- 2.41 (4 H, br s), 1.66-1.58 (4 h br m), 1.53- 1.42 (2 H, br s), 1.23 (6 H, d). MS: [M + H].sup.+ 395. 73 (2,4-Dihydroxy-5- isopropyl- phenyl)-[5-(4- hydroxypiperidin- 4-yl)-1,3-dihydro- isoindol-2-yl]- methanone As for the synthesis of (2,4-dihydroxy- 5-isopropyl- phenyl)-[5-(4- hydroxy-1- methyl- piperidin-4-yl)- 1,3-dihydro- isoindol-2-yl]- methanone (Example 50) except using N- benzyloxy- carbonyl- piperidin-4-one in step 2. .sup.1H NMR (Me- d.sub.3-OD) 7.47 (2 H, m), 7.30 (1 H, br m) 7.20 (1 H, s), 6.40 (1 H, s), 4.90 (4 H, d), 3.22 (1 H, m), 3.15 (2 H, m), 2.95 (2 H, m), 2.05 (2 H, m), 1.75 (2 H, m), 1.25 (6 H, d) MS: [M + H].sup.+ 397 74 (5-chloro-6- hydroxy-1,3- dihydro-isoindol- 2-yl)-(2,4- dihydroxy-5- isopropyl- phenyl)- methanone Isolated as a bi-product during the preparation of Example 33. .sup.1H NMR (DMSO-d.sub.6) 10.00 (1 H, s), 9.58 (1 H, s), 7.48-7.38 (1 H, m), 7.02 (1 H, s), 7.97-6.85 (1 H, m), 6.40 (1 H, s), 4.68 (4 H, brs), 3.10 (1 H, m), 1.15 (6 H, d) MS: [M + H].sup.+ 348

Example 75

(5-Chloro-2,4-dihydroxy-phenyl)-[5-(4-methyl-piperazin-1-yl)-1, 3-dihydro-isoindol-2-yl]-methanone

75A. 5-(4-Methyl-piperazin-1-yl)-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester

(306) ##STR00254##

(307) 5-Bromo-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester (2.97 g, 10 mmol) was azeotropically dried by evaporation from toluene. Tris(dibenzylideneacetone)dipalladium (0) (228 mg, 0.25 mmol), 2-(di-tert-butylphosphino)biphenyl (149 mg, 0.50 mmol) and sodium tert-butoxide (1.34 g, 13.9 mmol) were added and the flask was purged with nitrogen. Toluene (25 mL) then N-methylpiperazine (1.33 mL, 12 mmol) were added and the mixture was heated to 80° C. for 2 hours. After allowing to cool to r.t. the mixture was diluted with ether, filtered through Celite and concentrated to give a residue that was purified by flash chromatography on silica (2M methanolic ammonia/dichloromethane, 1% to 3% gradient). This afforded the title compound as a brown solid (1.45 g, 46%). .sup.1H NMR (MeOH-d.sub.4) 7.15 (1H, m), 6.94-6.88 (2H, m), 4.60-4.54 (4H, m), 3.20-3.17 (4H, m), 2.63-2.60 (4H, m), 2.34 (3H, s), 1.52 (9H, s). MS: [M+H].sup.+ 318.

75B. 5-(4-Methyl-piperazin-1-yl)-2,3-dihydro-1H-isoindole dihydrochloride

(308) ##STR00255##

(309) 5-(4-Methyl-piperazin-1-yl)-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester (247 mg, 0.78 mmol) was treated with 4M HCl in dioxane (4 mL, 4 mmol) for 24 hours. Concentration in vacuo afforded the title compound quantitatively, which was used directly in the coupling reaction. .sup.1H NMR (DMSO-d.sub.6) 11.13 (1H, br.s), 9.99 (2H, br.s), 7.27 (1H, d), 7.02-7.00 (2H, m), 4.43-4.37 (4H, m), 3.82-3.75 (2H, m), 3.49-3.43 (2H, m), 3.15-3.10 (4H, m), 2.79-2.78 (3H, s), 1.52 (9H, s). MS: [M+H].sup.+ 218.

75C. (5-Chloro-2,4-dihydroxy-phenyl)-[5-(4-methyl-piperazin-1-yl)-1,3-dihydro-isoindol-2-yl]-methanone

(310) ##STR00256##

(311) A solution of 5-chloro-2,4-dihydroxy-benzoic acid (176 mg, 0.93 mmol) in DMF (5 mL) was treated with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (179 mg, 0.93 mmol) then HOBt (126 mg, 0.93 mmol). After 45 min, the solution of the activated acid was added to a mixture 5-(4-methyl-piperazin-1-yl)-2,3-dihydro-1H-isoindole dihydrochloride (290 mg, 0.78 mmol) and triethylamine (0.28 mL, 2 mmol) then the mixture was stirred at r.t. for 3 h. The solvent was removed in vacuo then the residue was partitioned between ethyl acetate and water (×3). Each extract was washed with saturated sodium bicarbonate solution and brine then dried (MgSO.sub.4), combined and concentrated. Some insoluble material remained and this was dissolved in 1N hydrochloric acid and methanol then combined with the organic extracts. The pH was adjusted to 14 with solid sodium hydroxide and the mixture allowed to stand overnight. The pH was adjusted to 7 with 1N hydrochloric acid and the resulting precipitate was filtered off then subjected to purification by preparative HPLC to afford the title compound as a red solid. This was converted to its hydrochloride salt by treatment with 4M HCl in dioxane, concentration in vacuo and trituration with ether which gave a brown solid (91 mg, 27%). .sup.1H NMR (DMSO-d.sub.6) 11.10 (1H, br.s), 10.50 (1H, br.s), 7.26-7.15 (2H, m), 7.02-6.93 (2H, m), 6.69 (1H, s), 4.72-4.61 (4H, m), 3.78-3.72 (2H, m), 3.45 (2H, br.s), 3.12 (4H, br.s), 2.78 (3H, s). MS: [M+H].sup.+ 386/388.

Example 76

(2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-yl)-1,3-di-hydro-isoindol-2-yl]-methanone

76A. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-bromo-1,3-dihydro-isoindol-2-yl)-methanone

(312) ##STR00257##

(313) Coupling of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (5.0 g, 13.4 mmol) (Preparation B9) and 5-bromo-2,3-dihydro-1H-isoindole (Preparation C20) was completed according to method A4, using CH.sub.2Cl.sub.2 as the reaction solvent to give the title compound (8.34 g) as a beige solid. 76B. Synthesis of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-methyl-piperazin-1-yl)-1,3-dihydro-isoindol-2-yl]-methanone

(314) ##STR00258##

(315) To a mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-bromo-1,3-dihydro-isoindol-2-yl)-methanone (8.30 g, 15.0 mmol), 2-(di-t-butylphosphino)biphenyl (223 mg, 0.75 mmol), tris(dibenzylideneacetone)dipalladium (344 mg, 0.38 mmol), sodium tert-butoxide (2.17 g, 22.5 mmol) and 1-methyl-piperazine (2.16 mL, 19.5 mmol) under a N.sub.2 atmosphere was added anhydrous toluene (100 mL). The mixture was taken to 80° C. and heated at this temperature for 16 h. The mixture was allowed to cool to ambient temperature, diluted with ether (150 mL) and filtered through a plug of Celite, washing with ether. The filtrate was reduced in vacuo and the residue purified by column chromatography using an eluant of CH.sub.2Cl.sub.2-DMAW120 (1:0-0:1) to give the title compound (9.39 g) as a red gum.

76C. (2,4-dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-yl)-1,3-di-hydro-isoindol-2-yl]-methanone

(316) ##STR00259##

(317) A mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-methyl-piperazin-1-yl)-1,3-dihydro-isoindol-2-yl]-methanone (8.61 g, 15.0 mmol) and 10% Pd/C (1.0 g) in methanol (200 mL) was stirred vigorously under a hydrogen atmosphere (˜1 atm) for 18 h at ambient temperature. The mixture was filtered through a plug of Celite and reduced in vacuo to give a purple oil. This residue was purified by column chromatography using an eluant of DMAW120 to give the title compound as its acetate salt. This salt was taken up in MeOH (30 mL) and to the solution was added saturated HCl in EtOAc (20 mL). This mixture was stirred at ambient for 2 h and the solid formed collected by filtration and dried in vacuo to give the title compound as its hydrochloride salt (2.64 g) as a white solid.

Example 77

(5-Chloro-2,4-dihydroxy-phenyl)-[5-(4-methyl-piperazin-1-yl)-1,3-dihydro-isoindol-2-yl]-methanone

(318) ##STR00260##

(319) A solution of 5-chloro-2,4-dihydroxy-benzoic acid (176 mg, 0.93 mmol) in DMF (5 mL) was treated with 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (179 mg, 0.93 mmol) then HOBt (126 mg, 0.93 mmol). After 45 minutes, the solution of the activated acid was added to a mixture 5-(4-methyl-piperazin-1-yl)-2,3-dihydro-1H-isoindole dihydrochloride (290 mg, 0.78 mmol) and triethylamine (0.28 mL, 2 mmol) then the mixture was stirred at r.t. for 3 h. The solvent was removed in vacuo then the residue was partitioned between ethyl acetate and water (×3). Each extract was washed with saturated sodium bicarbonate solution and brine then dried (MgSO.sub.4), combined and concentrated. Some insoluble material remained and this was dissolved in 1N hydrochloric acid and methanol then combined with the organic extracts. The pH was adjusted to 14 with solid sodium hydroxide and the mixture allowed to stand overnight. The pH was adjusted to 7 with 1N hydrochloric acid and the resulting precipitate was filtered off then subjected to purification by preparative HPLC to afford the title compound as a red solid. This was converted to its hydrochloride salt by treatment with 4M HCl in dioxane, concentration in vacuo and trituration with ether which gave a brown solid (91 mg, 27%). .sup.1H NMR (DMSO-d.sub.6) 11.10 (1H, br.s), 10.50 (1H, br.s), 7.26-7.15 (2H, m), 7.02-6.93 (2H, m), 6.69 (1H, s), 4.72-4.61 (4H, m), 3.78-3.72 (2H, m), 3.45 (2H, br.s), 3.12 (4H, br.s), 2.78 (3H, s). MS: [M+H].sup.+ 386/388.

Example 78

Alternative Synthesis of (2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone

78A. 5-bromo-2-trityl-2,3-dihydro-1H-isoindole

(320) ##STR00261##

(321) Trityl chloride (2.30 g, 8.23 mmol) was added to a solution of 5-bromo-2,3-dihydro-1H-isoindole (1.64 g, 8.23 mmol) and triethylamine (1.4 mL, 9.9 mmol) in dichloromethane (20 mL). After 18 hours the solvent was removed in vacuo, the residue taken up in ethyl acetate and washed with water (×2) and brine, dried (MgSO.sub.4) and concentrated. The crude material was purified by flash chromatography on silica eluting with 1% triethylamine/10% ethyl acetate/petrol to give 5-bromo-2-trityl-2,3-dihydro-1H-isoindole as a reddish-brown solid (3.10 g, 85%). .sup.1H NMR (CDCl.sub.3) 7.91-7.84 (1H, m), 7.57 (6H, d), 7.45-7.41 (1H, m), 7.33-7.14 (9H, m), 6.95 (1H, d), 3.90 (2H, s), 3.86 (2H, s). MS: Ph.sub.3C.sup.+ 243.

78B. 1-methyl-4-(2-trityl-2,3-dihydro-1H-isoindol-5-yl)-piperidin-4-ol

(322) ##STR00262##

(323) Under nitrogen, a solution of 5-bromo-2-trityl-2,3-dihydro-1H-isoindole (2.03 g, 4.6 mmol) in THF (20 mL) was cooled to −78° C. n-Butyllithium solution (2.5M in hexanes, 2.0 mL, 5 mmol) was added over 5 minutes, then after 10 minutes, 1-methyl-4-piperidone was added dropwise. After a further hour, the cooling bath was removed and the reaction quenched with sodium bicarbonate solution. The mixture was extracted with ethyl acetate then the organic phase was washed with brine, dried (MgSO.sub.4) and concentrated. The residue was purified by flash chromatography on silica (gradient elution with 2M methanolic ammonia/dichloromethane, 0% to 5%) to afford 1-methyl-4-(2-trityl-2,3-dihydro-1H-isoindol-5-yl)-piperidin-4-ol as a pink foam (1.25 g, 57%). .sup.1H NMR (MeOH-d.sub.4) 7.56 (6H, dd), 7.28 (6H, t), 7.25-7.21 (2H, m), 7.15 (3H, t), 7.03 (1H, d), 3.92 (2H, s), 3.91 (2H, s), 2.70 (2H, d), 2.53 (2H, td), 2.33 (3H, s), 2.06 (2H, td), 1.70 (2H, d). MS: [M+H].sup.+ 475.

78C. 4-(2,3-Dihydro-1H-isoindol-5-yl)-1-methyl-piperidin-4-ol dihydrochloride

(324) ##STR00263##

(325) A mixture of 1-methyl-4-(2-trityl-2,3-dihydro-1H-isoindol-5-yl)-piperidin-4-ol (1.42 g, 3.0 mmol), 5N hydrochloric acid (5 mL) and methanol (10 mL) was placed under nitrogen then heated to reflux for 80 minutes. After cooling, the mixture was concentrated in vacuo to remove methanol, diluted with water and washed with ethyl acetate (×2). The aqueous phase was concentrated to dryness to afford the title compound in quantitative yield as a black solid. .sup.1H NMR (MeOH-d.sub.4) 7.62 (1H, s), 7.57 (1H, d), 7.45 (1H, d), 4.64 (2H, s), 4.63 (2H, s), 3.49-3.46 (4H, m), 2.95 (3H, s), 2.40-2.32 (2H, m), 1.97 (2H, dd).

78D. (2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone

(326) ##STR00264##

(327) 2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid (1.65 g, 4.4 mmol), 1-[3-(dimethylamino)propyl)]-3-ethylcarbodimide hydrochloride (843 mg, 4.4 mmol) and 1-hydroxybenzotriazole (595 mg, 4.4 mmol) were dissolved in DMF (20 mL). After 35 minutes, the solution was added to a suspension of 4-(2,3-dihydro-1H-isoindol-5-yl)-1-methyl-piperidin-4-ol dihydrochloride (1.22 g, 4.0 mmol) in DMF (5 mL) and triethylamine (1.4 mL, 10 mmol). The mixture was stirred for 3 hours then concentrated in vacuo. The residue was taken up in ethyl acetate and washed with a mixture of water (adjusted to pH 14 with 2N sodium hydroxide solution) and brine. The aqueous phase was extracted twice further with ethyl acetate then the combined organic extracts were washed with sodium bicarbonate solution and brine, dried (MgSO.sub.4) and concentrated. The crude product was purified by flash chromatography (gradient elution with 2M methanolic ammonia/dichloromethane, 2% to 10%) to afford the title compound as a brown foam (1.62 g, 69%). .sup.1H NMR (methanol-d.sub.4) 7.51-7.14 (14H, m), 6.85 (0.5H, s), 6.84 (0.5H, s), 5.16 (2H, s), 5.15 (2H, s), 5.10-5.08 (1H, m), 5.07-5.05 (1H, m), 4.87 (1H, s), 4.86 (1H, s), 4.61 (2H, br.s), 2.78-2.70 (2H, m), 2.57 (1H, td), 2.54 (1H, td), 2.36 (1.5H, s), 2.34 (1.5H, s), 2.16-2.05 (5H, m including 2.09 (3H, s)), 1.78-1.70 (2H, m). MS: [M+H].sup.+ 589.

78E. (2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone

(328) ##STR00265##

(329) (2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-hydroxy-1-methyl-piperidin-4-yl)-1,3-dihydro-isoindol-2-yl]-methanone (Example 50F) (1.62 g, 2.75 mmol) was dissolved in methanol (50 mL) and hydrogenated at 50° C. over 10% palladium on charcoal using an H-cube hydrogenation apparatus, under free hydrogen conditions. Concentration afforded the title compound (1.14 g, 100%) as a yellow solid, the NMR and mass spectrometric data of which were as set out in Example 50E.

Example 79

(2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-ethoxy)-7-methyl-1,3-dihydro-isoindol-2-yl]-methanone

79A. 7-Methyl-2,3-dihydro-1H-isoindol-5-ol hydrobromide

(330) ##STR00266##

(331) Using the method of preparation C2,5-methoxy-3-methyl-phthalic acid dimethyl ester (prepared according to Tam and Coles, Synthesis 1988, 383) was hydrolysed to 5-methoxy-3-methyl-phthalic acid. .sup.1H NMR (DMSO-d.sub.6) 12.95 (2H, br.s), 7.15 (1H, d), 7.04 (1H, d), 3.80 (3H, s), 2.29 (3H, s). MS: [M−H].sup.+ 209.

(332) 5-Methoxy-3-methyl-phthalic acid was converted to 5-methoxy-3-methyl-phthalic anhydride. .sup.1H NMR (DMSO-d.sub.6) 7.40 (1H, d), 7.34-7.33 (1H, m), 3.94 (3H, s), 2.58 (3H, s).

(333) 5-Methoxy-3-methyl-phthalic anhydride was used to prepare 6-methoxy-4-methyl-isoindole-1,3-dione. .sup.1H NMR (DMSO-d.sub.6) 11.05 (1H, br.s), 7.13 (1H, d), 7.10 (1H, d), 3.88 (3H, s), 2.55 (3H, s).

(334) Reduction of 6-methoxy-4-methyl-isoindole-1,3-dione according to the method of preparation C2 afforded 6-methoxy-4-methyl-isoindole. .sup.1H NMR (DMSO-d.sub.6) 6.64 (1H, s), 6.57 (1H, s), 4.05 (2H, s), 3.96 (2H, s), 3.70 (3H, s), 2.16 (3H, s). MS: [M+H].sup.+ 164.

(335) 6-Methoxy-4-methyl-isoindole was demethylated to give the title compound as its hydrobromide salt. .sup.1H NMR (DMSO-d.sub.6) 9.52 (1H, br.s), 9.29 (2H, br.s), 6.59 (1H, s), 6.56 (1H, s), 4.41 (2H, t), 4.34 (2H, t), 2.17 (3H, s).

79B. (2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-(5-hydroxy-7-methyl-1,3-dihydro-isoindol-2-yl)-methanone

(336) ##STR00267##

(337) 2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid (248 mg, 0.66 mmol), 1-[3-(dimethylamino)propyl)]-3-ethylcarbodiimide hydrochloride (127 mg, 0.66 mmol) and 1-hydroxybenzotriazole (89 mg, 0.661 mmol) were dissolved in DMF (5 mL). After 20 minutes, 7-methyl-2,3-dihydro-1H-isoindol-5-ol hydrobromide (152 mg, 0.66 mmol) and triethylamine (0.14 mL, 0.99 mmol) were added. After a further 3.5 hours the mixture was concentrated in vacuo and the residue was treated with 1N hydrochloric acid and ethyl acetate. The aqueous phase was removed, brine was added and the title compound was collected by filtration as a grey solid (168 mg, 57%). .sup.1H NMR (DMSO-d.sub.6) 9.30 (0.47H, s), 9.24 (0.53H, s), 7.48-7.25 (10H, m), 7.09 (0.47H, s), 7.08 (0.53H, s), 6.99 (0.47H, s), 6.98 (0.53H, s), 6.56 (0.47H, s), 6.50 (0.53H, s), 6.48 (0.47H, s), 6.44 (0.53H, s), 5.24 (0.47H, s), 5.22 (0.53, s), 5.18 (2H, s), 5.10-5.07 (2H, m), 4.70 (0.47H, s), 4.61 (0.53H, s), 4.46 (0.47H, s), 4.36 (0.53H, s), 2.17 (1.41H, s), 2.04 (3H, s), 1.99 (1.59H, s). MS: [M+H].sup.+ 506.

79B. (2,4-Bis-benzyloxy-5-isopropenyl-Phenyl)-[5-(2-dimethylamino-ethoxy)-7-methyl-1,3-dihydro-isoindol-2-yl]-methanone

(338) ##STR00268##

(339) A mixture of (2,4-bis-benzyloxy-5-isopropenyl-phenyl)-(5-hydroxy-7-methyl-1,3-dihydro-isoindol-2-yl)-methanone (164 mg, 0.32 mmol), potassium carbonate (112 mg, 0.81 mmol) and 2-(dimethylamino)ethyl chloride hydrochloride (93 mg, 0.64 mmol) in DMF (5 mL) was heated at 60° C. for 17 hours then 90° C. for 6 hours. Further portions of potassium carbonate (112 mg, 0.81 mmol) and 2-(dimethylamino)ethyl chloride hydrochloride (93 mg, 0.64 mmol) were added and the mixture maintained at 60° C. for 72 hours and finally, a further 24 hours at 90° C. The mixture was concentrated in vacuo then the residue was partitioned between ethyl acetate and 0.5N aqueous sodium hydroxide. The organic phase was washed with brine (×2), dried (MgSO.sub.4) and concentrated to give a residue which was purified by preparative HPLC (acidic method) to afford the title compound as a formate salt (37 mg, 20%). .sup.1H NMR (MeOH-d.sub.4) 8.51 (1H, br.s), 7.43-7.27 (7H, m), 7.24-7.20 (3H, m), 7.17 (0.5H, s), 7.16 (0.5H, s), 6.85 (0.5H, s), 6.84 (0.5H, s), 6.81 (0.5H, s), 6.77 (0.5H, s), 6.74 (0.5H, s), 6.62 (0.5H, s), 5.16 (1H, s), 5.14 (3H, s), 5.09 (1H, m), 5.06 (1H, m), 4.83 (1H, s), 4.74 (1H, s), 4.60 (1H, s), 4.48 (1H, s), 4.28 (1H, t), 4.23 (1H, t), 3.41 (1H, t), 3.37 (1H t), 2.84 (3H, s), 2.81 (3H, s), 2.27 (1.5H, s), 2.09 (3H, s), 2.07 (1.5H, s). MS: [M+H].sup.+ 577.

79C. (2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(2-dimethylamino-ethoxy)-7-methyl-1,3-dihydro-isoindol-2-yl]-methanone

(340) ##STR00269##

(341) (2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-[5-(2-dimethylamino-ethoxy)-1,3-dihydro-isoindol-2-yl]-methanone (37 mg, 0.06 mmol) was hydrogenated in methanol at 50° C. over 10% palladium on charcoal using an H-cube hydrogenation apparatus, under free hydrogen conditions. The product was purified by preparative HPLC (basic method) to give the title compound as an off-white solid (9 mg, 35%). .sup.1H NMR (MeOH-d.sub.4) 7.18 (1H, s), 6.77-6.65 (2H, br.m), 6.37 (1H, s), 4.85 (water obscuring CH.sub.2), 4.77 (2H, s), 4.08 (2H, t), 3.20 (1H, sept), 2.81 (2H, t), 2.39 (6H, s), 2.22 (3H, br.s), 1.21 (6H, d). MS: [M+H].sup.+ 399.

Example 80

(2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

Step 1

4-Acetoxy-2-hydroxy-benzoic acid methyl ester

(342) ##STR00270##

(343) Resorcinol methyl ester (50 g, 0.298 mol) and N,N-dimethyl-4-aminopyridine (0.27 g, 0.0022 mol, 0.74 mol %) were added to toluene 0.2 L followed by acetic anhydride (30 mL, 0.318 mol). The solution was heated to 50° C. for 2 h. The solvent was removed by evaporation at 50° C. to a small volume and the residue was azeotroped once with toluene. To the residual oil was immediately added toluene (100 mL) whilst still warm and the solution used for Step 2 without further purification.

Step 2

5-Acetyl-2,4-dihydroxy-benzoic acid methyl ester

(344) ##STR00271##

(345) The toluene solution from Step 1 was cooled in an ice bath under N.sub.2 and triflic acid (26 mL) added slowly over 30 min. On stirring a fine white solid was formed which dissolved on stirring for 16 h at RT to give a yellow solution. To the solution was added acetyl chloride (2 mL) and the solution stirred at RT for a further 1 h. This solution was cannulated into a stirred cooled (0° C.) solution of EtOAc (600 mL) and NaOAc.3H.sub.2O (40 g) dissolved in water (400 mL). The organic phase was washed with water (twice, 200 mL), saturated brine and was evaporated to a small volume without drying. The residue was azeotroped with heptane (twice, 100 mL) and heptane (100 mL) was added and the crystalline solid removed by filtration, washed well on sinter with heptane and dried to give 49.5 g (79%).

(346) Final Purification of Combined Batches

(347) The combined batches of solid (96.3 g) was heated to boiling with 10% IPA/heptane (250 mL) then cooled to RT and finally to 0° C., filtered and the residue dried 72 h (oil pump) to give (88.04 g, 91.5%), pure by hplc, tlc and NMR.

(348) .sup.1H NMR (DMSO-d.sub.6) 12.58 (1H, s), 11.22 (1H, s), 8.33 (1H, s), 6.45 (1H, s), 3.90 (3H, s), 2.62 (3H, s).

Step 3

5-Acetyl-2,4-dihydroxy-benzoic acid methyl ester

Alternative Procedure

(349) ##STR00272##

(350) Resorcinol methyl ester (50 g, 0.298 mol) and Amberlyst 15 resin (40 g) were suspended in toluene 150 mL (under a nitrogen atmosphere) and the solution was heated in an oil bath at 70° C. (internal temp 56° C.). Acetyl chloride (22 mL, 308 mmol) was added in 5 mL portions over 30 mins giving evolution of gaseous HCl (which was scrubbed by passing the nitrogen stream through aqueous NaOH). The solution was stirred at 70° C. for 4.5 h then heated in an oil bath temp (internal temperature 96° C.) for 3.5 h. The solution was cooled to 50° C. and EtOAc (100 mL) was added and the solution filtered whilst at this temperature. The residual resin was washed with EtOAc (50 mL) and the combined filtrates were concentrated to slurry of crystalline solid (total weight of 128 g for solid plus solvent). To the slurry was added heptane (100 mL) and after 10 mins at RT the solid was removed by filtration. The residue was washed with heptane:toluene (2:1, 60 mL) then with petroleum ether bp 40-60° C. and dried in vacuo to give crop 1 29 g (46.4%) (NMR showed 3% of material resulting from saponification of the methyl ester).

(351) The filtrate was evaporated to a small volume and 20% EtOAc in heptane (100 mL) was added. After standing at RT 16 h a second crop of 4.75 g (7.6%) was obtained (NMR identical to crop 1).

Step 4

5-Acetyl-2,4-bis-benzyloxy-benzoic acid methyl ester

(352) ##STR00273##

(353) Benzyl bromide (70 ml, 0.59 mol) was added to a stirred mixture of methyl 5-acetyl-2,4-dihydroxybenzoate (60.7 g, 0.29 mol) and anhydrous potassium carbonate (87.8 g, 0.64 mol) in acetonitrile (800 ml) and the mixture was stirred and held at reflux for 16 hours. Upon cooling to room temperature the mixture was poured onto water (3 L) and stirred vigorously for 2 hours. The solids were collected by filtration, rinsed with water (2 L), sucked dry under reduced pressure and dried to constant mass in a vacuum oven at 60° C. overnight to afford methyl 5-acetyl-2,4-bis-benzyloxybenzoate (112.1 g, 99%) as a cream solid. .sup.1H NMR (DMSO-d.sub.6) 8.21 (1H, s), 7.55 (4H, m), 7.43 (4H, m), 7.37 (2H, m), 7.04 (1H, s), 5.38 (4H, s), 3.79 (3H, s), 2.48 (3H, s). MS: [M+H].sup.+ 391.

Step 5

2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid methyl ester

(354) ##STR00274##

(355) Potassium tert-butoxide (29.1 g, 0.26 mol) was added to a stirred suspension of methyltriphenylphosphonium bromide (92.8 g, 0.26 mol) in anhydrous tetrahydrofuran (1 L) and the mixture was stirred at room temperature for 10 minutes whereupon methyl 5-acetyl-2,4-bis-benzyloxybenzoate (78.0 g, 0.2 mol) was added and the mixture stirred at room temperature for a further 30 minutes. Methanol (100 ml) was added to quench excess phosphorus ylide and the solvent was removed in vacuo to afford an orange oil that crystallized on standing. The residue was recrystallized from methanol (330 ml). The solids were collected by suction filtration, washed with methanol (50 ml) and sucked dry under reduced pressure to afford methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate as pale yellow needles. The mother liquor deposited a second crop of material upon standing overnight (combined yield: 56.55 g, 73%) .sup.1H NMR (DMSO-d.sub.6) 7.59 (1H, s), 7.52 (2H, d), 7.64-7.32 (8H, m), 6.97 (1H, s), 5.28 (2H, s), 5.22 (2H, s), 5.09 (1H, s), 5.04 (1H, s), 3.76 (3H, s), 2.02 (3H, s). MS: [M+H].sup.+ 389.

(356) A further crop of the ester could be obtained as follows. The crystallization residues were evaporated to dryness in vacuo and the oily solid was treated with 5% ethyl acetate in heptane (250 ml). Ethyl acetate was added in small portions to the vigorously stirred mixture until the residue deposited a large quantity of solid triphenylphosphine oxide. The solids were removed by filtration and the filtrate evaporated to dryness in vacuo to afford an orange oil. Recrystallization from methanol (as described above) afforded further methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate as a pale yellow crystalline solid (total yield 85-90%).

Step 6

2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid

(357) ##STR00275##

(358) Potassium hydroxide (10.96 g, 0.19 mmol) was added to a stirred suspension of methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate (61.0 g, 0.16 mol) in methanol (750 ml) and water (250 ml) and the mixture was stirred and held at reflux for 16 hours. Upon cooling the organic solvent was removed in vacuo and the mixture acidified to pH 2 or below by the addition of 2M hydrochloric acid (200 ml). The mixture was diluted with water (2 L) and extracted with ethyl acetate (2 L), the organic layer was separated and the solvent removed in vacuo to afford 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (58.8 g, 100%) as a colourless solid. .sup.1H NMR (DMSO-d.sub.6) 7.52 (2H, d), 7.47-7.29 (9H, m), 6.82 (1H, s), 5.20 (2H, s), 5.17 (2H, s), 5.06 (1H, s), 5.04 (1H, s), 2.03 (3H, s). MS: [M+H].sup.+ 375.

Step 7

Di-prop-2-ynyl-carbamic acid benzyl ester

(359) ##STR00276##

(360) To a cooled (0° C.) solution of dipropargylamine (46.7 g, 502 mmol) in EtOAc (200 mL) and 10% aqueous K.sub.2CO.sub.3 (700 mL, 507 mmol) was slowly added a solution of N-(benzyloxycarbonyloxy)succinimide (125 g, 502 mmol) in EtOAc (500 mL) over 20 mins. The solution was stirred at 0° C. for 2 h then at RT 16 h. The phases were separated and the organic phase was washed with 10% aqueous K.sub.2CO.sub.3 (700 mL, 507 mmol) and then with saturated brine (500 mL) and was diluted to 1000 mL with EtOAc to give a 0.5M solution.

Step 8

5-Hydroxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester

(361) ##STR00277##

(362) A solution of propargyl alcohol (26.4 mL, 424 mmol) in toluene (120 mL) was degassed. The 0.5M-diyne solution above (440 mL, 220 mmol) was evaporated and the residue dissolved in toluene (80 mL). This protected diyne solution and Wilkinson's catalyst (2.26 g, 2.44 mmol, 1.11% were added in 14 equal portions over a 2 h period with constant monitoring of the internal temperature such that the temperature remained 50-100° C. The solution was allowed to cool to 50° C. over 30 min when the solution was evaporated (to remove excess propargyl alcohol). The residue was heated with toluene (500 mL) and charcoal (Darco 4-12 mesh, 20 g) at 100° C. for 30 min and then filtered hot through a bed of Celite and the brown solution was evaporated. The residue was dissolve in EtOAc (400 mL) at 80° C. when silica gel (chromatography grade 65 g) was added and heating continued for 20 mins. The solution was filtered whilst hot and then evaporated (with seeding) to give a pale brown solid. 10% EtOAc/heptane (v/v, 100 mL) was added and the solid removed by filtration. The solid was washed on the sinter with heptane (100 mL) and the dried (50° C., oil pump, 16 h) to give the title compound 59.0 g (95%). .sup.1H NMR (400 MHz, Me-d3-OD): 7.51-7.16 (m, 8H), 5.21 (s, 2H), 4.74 (s, 2H), 4.70 (s, 2H), 4.61 (s, 2H).

Step 9

5-Methanesulfonyloxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester

(363) ##STR00278##

(364) To a solution of 5-hydroxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester (65.75 g, 0.232 mol) in THF (470 mL) and EtOAc (770 mL) was added Et.sub.3N (39 mL, 0.28 mol). The solution was cooled in an ice-bath and a solution of methanesulphonyl chloride (19 mL, 0.245 mol) dissolved in EtOAc (50 mL) was added (so that the internal temp <12° C.). After stirring for 2 h in the ice-bath further additions of methanesulphonyl chloride (1.9 mL and 0.95 mL) and Et.sub.3N (3.9 mL) were made (so that by tlc there was no remaining starting material after a further 1 h of stirring). NaHCO.sub.3 (550 mL) was added and the solution stirred for 20 mins then saturated brine (200 mL) was added and the phases were separated. The organic phase was dried (MgSO.sub.4) and evaporated with seeding to give a damp solid which was used in the next step without thorough drying.

Step 10

5-(4-Methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester dihydrochloride salt

(365) ##STR00279##

(366) The solid from Step 9 (assume 0.232 mol) was dissolved in acetone (700 mL) and this solution was added over 45 mins to a cooled (internal temp 15-17° C.) suspension of K.sub.2CO.sub.3 (48 g) and N-methylpiperazine (50 mL, 0.45 mol) in acetone (330 mL). The suspension was stirred at 15° C. for 3 h (complete removal of starting material by tlc) when the solution was evaporated to a small volume and the residue partition between EtOAc (1000 mL) and a mixture of water (500 mL) and saturated brine (50 mL). The organic phase was washed with a mixture of water (500 mL) and saturated brine. (150 mL) and finally washed with saturated brine (300 mL). The solution was dried (MgSO.sub.4) and filtered and to this solution was added 1M-HCl in MeOH (430 mL, 0.43 mol). The suspension was cooled (0° C. for 30 mins) and the solid removed by filtration which was washed with EtOAc and then heptane on the sinter and the solid dried (oil-pump, RT 72 h) to give crop 1 of the title compound 66.34 g (65%) as a colourless solid. .sup.1H NMR (400 MHz, Me-d3-OD): 7.64-7.51 (m, 2H), 7.51-7.29 (m, 6H), 5.23 (s, 2H), 4.79 (dd, J=16.2, 6.1 Hz, 4H), 4.49 (s, 2H), 3.66 (s, 8H), 3.03 (s, 3H).

Alternative Step 10A

5-(4-Methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester dihydrochloride

(367) ##STR00280##

(368) Step 10A can be used as an alternative route to replace steps 9 and 10 above.

(369) To a suspension of manganese dioxide (15.5 g, 178 mmol) in DCM (100 mL) was added 5-hydroxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester (3.35 g, 11.8 mmol) and after 6 h stirring at RT a further addition of manganese dioxide (5 g, 57 mmol) was made. After a further 1 h stirring at RT Celite (7 g) was added and the solution was filtered through a bed of Celite™ giving a clear pale yellow solution. The Celite™ was washed with DCM and the volume of the combined organic solution adjusted to 100 mL by evaporation. N-Methylpiperazine (1.31 mL, 11.8 mmol) and acetic acid (0.68 mL) were added followed by sodium triacetoxyborohydride (4.98 g, 23.5 mmol). The yellow solution was stirred 16 h giving a colourless solution. To the solution was added 2M-HCl (10 mL, 20 mmol) giving an effervescence. After 30 min water (10 mL) and K.sub.2CO.sub.3 (5.5 g, 39.8 mmol) were added and the organic phase was dried (Na.sub.2SO.sub.4). After filtration 4M-HCl in dioxan (6 mL) was added with stirring and the suspension was evaporated to dryness. The residue was dissolved in MeOH with warming and after evaporation the solid was washed on a sinter with EtOAc then petrol (bp 40-60° C.) followed by drying in vacuo at 50° C. to give the title compound 3.61 g (70%). .sup.1H NMR (400 MHz, Me-d3-OD): 7.65-7.51 (2H, m), 7.51-7.27 (6H, m), 5.23 (2H, s), 4.83-4.69 (4H, m), 4.49 (2H, s), 3.66 (8H, d), 3.03 (3H, s)

Step 11

5-(4-Methyl-piperazin-1-ylmethyl)-2,3-dihydro-1H-isoindole

(370) ##STR00281##

(371) To 5-(4-Methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester dihydrochloride salt (Step 10, 59.8 g, 136.7 mmol) was added EtOAc (400 mL) and 10% aqueous K.sub.2CO.sub.3 (400 mL). The organic phase was washed with saturated brine (200 mL) and then dried (MgSO.sub.4). The solution was filtered and was evaporated to an oil (which crystallised on standing with petroleum ether (bp 40-60° C.)). The solid was dried in vacuo to give a colourless solid: 48.8 g (133.5 mmol).

(372) A portion of the solid (24.4 g, 66.8 mmol) was dissolved in MeOH (170 mL) and after degassing the solution and purging with nitrogen 10% Pd/C (1.22 g) was added and the mixture hydrogenated at 1 atmosphere for 2.5 h. The solution was filtered and the solution evaporated and the residue was azeotroped twice with toluene at 30-40° C. The residue was dissolved in DMF (92 mL) and the solution was immediately degassed and purged with N.sub.2.

(373) (NB The product at this stage is sensitive to air and darkens on contact with oxygen. The DMF solution was used immediately but can be stored by degassing and storing under an atmosphere of N.sub.2)

Step 12

(2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(374) ##STR00282##

(375) A solution of the resorcinol acid (Step 6, 23.7 g, 63.4 mmol) and 1-hydroxybenzotriazole (10.21 g, 66.7 mmol) were dissolved in DMF (92 mL) and to this solution was added N-ethyl-N′-(3-dimethylaminopropyl)carbodiimide hydrochloride (12.8 g, 66.8 mmol). The solution was stirred at RT for 40 mins and this solution was added to the solution of the amine from Step 11 (66.8 mmol) together with DMF (5 mL) washings. The solution was degassed and the solution stirred at RT for 16 h. To the solution was added 10% K.sub.2CO.sub.3 (500 mL) and EtOAc (500 mL) and the organic phase was washed sequentially with 10% K.sub.2CO.sub.3 (500 mL), water (4×100 mL) and saturated brine (200 mL). The solution was evaporated to a small volume and 20% EtOAc in heptane (250 mL) was added and stored at 0° C. The solid which had formed was removed by filtration, washed with heptane twice and was dried in vacuo to give the title compound 35.05 g (94.4%). .sup.1H NMR (400 MHz, Me-d3-OD): 7.49-7.10 (m, 14H), 6.86 (d, J=2.5 Hz, 1H), 5.17 (d, J=2.5 Hz, 4H), 5.09 (d, J=11.3 Hz, 2H), 4.88 (s, 2H), 4.63 (s, 2H), 3.54 (d, J=16.0 Hz, 2H), 2.50 (s, 7H), 2.28 (d, J=7.6 Hz, 3H), 2.11 (s, 3H).

Step 13

(2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(376) ##STR00283##

(377) The product from Step 12 (4.7 g) was dissolved in 1:1 MeOH/water (98 mL) and after purging with N.sub.2 10% Pd/C and K.sub.2CO.sub.3 (2.38 g, 17.2 mmol) were added and the suspension was hydrogenated for 16 h under an atmosphere of H.sub.2. The solution was filtered and the solvent evaporated. To the residue was added aqueous 2M-HCl (40 mL) and the solution was washed with 1:1 EtOAc/petrol (40 mL×2) and then the pH adjusted to pH 8.5 by addition of NaOH and EtOAc (50 mL) added. The solution was heated to 60° C. and the aqueous phase removed. The hot organic phase was washed with water (30 mL) and then evaporated to a small volume (ca. 5 mL) and allowed to stand at RT 16 h with seeding. To the crystalline material was added 1:1 EtOAc/petrol (10 mL) and the mixture was filtered and dried to give the title compound as the free base 1.76 g. .sup.1H NMR (400 MHz, Me-d3-OD): 7.29 (s, 3H), 7.19 (s, 1H), 6.39 (s, 1H), 4.91 (s, 4H), 3.56 (s, 2H), 3.28-3.15 (m, 1H), 2.53 (s, 8H), 2.31 (s, 3H), 1.23 (d, J=6.9 Hz, 7H).

Optional Step 14

Purification of (2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(378) In some batches of product, the title compound (X═H in the formula) can contain small amounts of the impurity 2,4-Dihydroxy-5-(2-hydroxypropy-2-yl)-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone (X═OH in the formula). The impurities can be removed by the following method.

(379) ##STR00284##

(380) Acetic anhydride (1.04 ml, 11.0 mmol) was added to a stirred suspension of impure 2-(2,4-dihydroxy-5-isopropylbenzoyl)-5-(4-methylpiperazin-1-ylmethyl)-1,3-dihydroisoindole (2.05 g, 5.0 mmol) in toluene (20 ml) and the resulting mixture was stirred and held at 100° C. for 16 hours. Upon cooling to room temperature the solvent was removed in vacuo to afford a brown oil which was dissolved in methanol (20 ml). Concentrated hydrochloric acid (1 ml) was added and the mixture was stirred and held at reflux for 5 hours. Upon cooling to room temperature, the organic solvent and volatile material were removed in vacuo and the aqueous residue was diluted with water (25 ml) and basified to pH 8 with vigorous stirring by the careful addition of 10% aqueous potassium carbonate solution. 50% Ethyl acetate in heptane (50 ml) was added and the mixture was stirred vigorously at room temperature for 16 hours. The solid material was collected by suction filtration, rinsed with 50% ethyl acetate in heptane (50 ml), sucked dry under reduced pressure and dried overnight in a vacuum oven at 50° C. to afford 2-(2,4-dihydroxy-5-isopropylbenzoyl)-5-(4-methylpiperazin-1-ylmethyl)-1,3-dihydroisoindole (1.85 g, 90%) as an off-white solid. .sup.1H NMR (DMSO-d.sub.6) 10.07 (1H, br s), 9.60 (1H, br s), 7.24 (3H, m), 7.06 (1H, s), 6.40 (1H, s), 4.76 (4H, br s), 3.44 (2H, s), 3.10 (1H, m), 2.32 (8H, m), 2.14 (3H, s), 1.15 (6H, d). MS: [M+H].sup.+ 410.

Example 81

(2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone L-lactate salt (form FL1)

(381) The product of Example 1 (1.24 g, 3.303 mmol) was suspended in ethanol (3 mL) and EtOAc (5 mL) and a solution of L-lactic acid (0.285 g, 3.13 mmol) dissolved in ethanol (3 mL) was added. The solution was heated until clear and then was filtered. EtOAc (5 mL) was used to wash the filter and the combined filtrates were stirred at RT for 2 h with seeding. The crystalline mass which formed was removed by filtration, was washed with EtOAc and then dried in vacuum at 50° C. to give the title compound 1.29 g. .sup.1H NMR (400 MHz, Me-d3-OD): 7.30 (s, 3H), 7.18 (s, 1H), 6.39 (s, 1H), 4.91 (s, 4H), 4.08 (q, J=6.8 Hz, 1H), 3.70-3.63 (m, 2H), 3.28-3.15 (m, 1H), 3.01 (s, 4H), 2.68 (m, 7H), 1.36 (d, J=6.8 Hz, 3H), 1.23 (d, J=6.9 Hz, 6H).

Example 82

(2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone L-lactate salt

(382) Example 82 describes a synthetic route containing essentially the same process steps as the route described in Examples 80 and 81 but wherein the process conditions are more suited to larger scale reactions.

Step 1

4-Acetoxy-2-hydroxy-benzoic acid methyl ester

(383) To a heated solution (50° C.) of resorcinol methyl ester (16.5 Kg, 98.1 mol) and N,N-dimethyl-4-aminopyridine (89.1 g, 0.73 mol, 7.4 mol %) in toluene (66 L) was slowly added (over 2 h) acetic anhydride (9.9 L, 104.9 mol). The solution was heated to 50° C. for a further 1.5 h and then the solvent was removed by evaporation at 50° C. to a small volume and the residue was azeotroped once with toluene. To the residual oil was immediately added toluene (33 L) whilst still warm and the solution used for Step 2 without further purification.

Step 2

5-Acetyl-2,4-dihydroxy-benzoic acid methyl ester

(384) The toluene solution from Step 1 was cooled in an ice bath under N.sub.2 and triflic acid (9.44 L) added slowly over 3 h. On stirring a fine white solid was formed which dissolved on warming to RT over 20 h and then stirring at RT for 37 h to give a yellow solution. To the solution was added acetyl chloride (726 mL) and the solution stirred at RT for a further 1 h. This solution was cannulated into a stirred cooled (0° C.) solution of EtOAc (217.8 L) and NaOAc.3H.sub.2O (14.52 Kg) dissolved in water (145 L). The organic phase was washed with saturated brine (twice, 72.6 L), and was evaporated to 5.5 Kg. Toluene:Isopropanol (2:3) was added and the crystalline solid removed by filtration and dried to give 12.6 Kg (61% over 2 steps), mp 124-126° C.

Step 3

5-Acetyl-2,4-bis-benzyloxy-benzoic acid methyl ester

(385) To a stirred solution of benzyl bromide (16.14 L, 136 mol) and anhydrous potassium carbonate (20.25 Kg, 147.6 mol) in acetonitrile (184.5 L) was added methyl 5-acetyl-2,4-dihydroxybenzoate (14 Kg, 66.6 mol, step 2) in 6 portions over 5 h. The mixture was stirred and held at reflux for 20 hours, cooled to room temperature the mixture was poured onto water (682 L) and stirred vigorously for 2 hours. The solids were collected by centrifugation and dried under reduced pressure to constant mass in a vacuum oven at 60° C. overnight to afford methyl 5-acetyl-2,4-bis-benzyloxybenzoate (23.5 Kg, 97.3%) as a cream solid mp 114-115° C.

Step 4

2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid methyl ester

(386) A solution of potassium tert-butoxide (6.72 Kg, 60.1 mol) in anhydrous THF (60 L) was added over 3 h to a stirred suspension of methyltriphenylphosphonium bromide (21.43 Kg, 60.1 mol) and methyl 5-acetyl-2,4-bis-benzyloxybenzoate (21.3 Kg, 54.6 mol, step 3) in anhydrous tetrahydrofuran (213 L) at 15° C. The mixture was stirred at 15° C. for 70 mins and the warmed to 20° C. over 60 mins. Methanol (27.3 L) was added to quench excess phosphorus ylide and the solvent was concentrated in vacuo followed by addition of EtOAc and water. The organic phase was treated with activated charcoal, filtered and evaporated to a small volume. The residue was crystallised from boiling MeOH and the solids were collected by suction filtration, washed with methanol and dried under reduced pressure to afford methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate 18.1 Kg (85%) as pale yellow needles mp 92-94° C. (99.6% pure by hplc).

Step 5

2,4-Bis-benzyloxy-5-isopropenyl-benzoic acid

(387) Potassium hydroxide (0.527 Kg, 9.4 mol) was added to a stirred suspension of methyl 2,4-bis-benzyloxy-5-isopropenyl-benzoate (3.1 Kg, 8 mol, step 4) in methanol (18.6 L) and water (12.4 L) and the mixture was stirred and held at reflux for 3 hours. The methanol was removed under partial vacuum from the vessel, and to the remaining solution was added toluene (62 L). The solution was heated to 40° C. and to the mixture was added conc HCl (1.36 L). The biphasic mixture is heated to 50° C. and the phases separated. The organic phase was washed with water (31 L) at 50° C. and the organic phase was evaporated under reduced pressure to give 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid 2.851 Kg (95% yield) as a colourless solid.

Step 6

Di-prop-2-ynyl-carbamic acid benzyl ester

(388) To a cooled (5° C.) solution of K.sub.2CO.sub.3 (4 Kg, 29.0 mol) in water (17.5 L) and toluene (12.5 L) was added dipropargylamine (2.50 Kg, 26.88 mol). Benzyloxychloroformate (4.8 Kg, 28.14 mol) was added at a rate such that T<10° C. The solution was stirred at 5° C. for 10 mins and then allowed to warm to RT. The aqueous phase was separated and the organic phase was washed with 0.2M HCl (12.5 L), sat NaHCO.sub.3 (13.5 L) and brine (17 L) and the resultant solution used in step 7 (assayed to contain 6.23 Kg, 102% based on an evaporated portion).

Step 7

5-Hydroxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester

(389) A solution of propargyl alcohol (2.11 Kg, 37.7 mol) in toluene (32.48 L) was degassed and heated to 55° C. The solution of di-prop-2-ynyl-carbamic acid benzyl ester (4.06 Kg, 17.86 mol, step 6) in toluene and Wilkinsons catalyst (0.162 Kg) were added in 10 equal portions such that temperature <65° C. (the exotherm was allowed to subside before the next addition was made). The solution was then stirred at 55° C. for 1 h and then cooled to 20° C. DCM (8.12 L) was added and the mixture was concentrated to a small volume. Toluene (8 L) was added and the solution evaporated to constant weight giving the title compound 5.72 Kg (113%).

Step 8

5-Methanesulfonyloxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester

(390) To a cooled solution (5° C.) of 5-hydroxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester (11 Kg, 38.8 mol, step 7) and Et.sub.3N (7.04 L, 50.6 mol) in DCM (55 L) was added methanesulphonyl chloride (2.97 L, 38.4 mol) so that the internal temp <10° C. After stirring for 0.5 h at 5° C. the solution was used below in step 9.

Step 9

5-(4-Methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester dihydrochloride salt

(391) ##STR00285##

(392) The solid from Step 8 (assume 0.232 mol) was dissolved in acetone (700 mL) and this solution was added over 45 mins to a cooled (internal temp 15-17° C.) suspension of K.sub.2CO.sub.3 (48 g) and N-methylpiperazine (50 mL, 0.45 mol) in acetone (330 mL). The suspension was stirred at 15° C. for 3 h (complete removal of starting material by tlc) when the solution was evaporated to a small volume and the residue partition between EtOAc (1000 mL) and a mixture of water (500 mL) and saturated brine (50 mL). The organic phase was washed with a mixture of water (500 mL) and saturated brine (150 mL) and finally washed with saturated brine (300 mL). The solution was dried (MgSO.sub.4) and filtered and to this solution was added 1M-HCl in MeOH (430 mL, 0.43 mol). The suspension was cooled (0° C. for 30 mins) and the solid removed by filtration which was washed with EtOAc and then heptane on the sinter and the solid dried (oil-pump, RT 72 h) to give crop 1 of the title compound 66.34 g (65%) as a colourless solid. .sup.1H NMR (400 MHz, Me-d3-OD): 7.64-7.51 (m, 2H), 7.51-7.29 (m, 6H), 5.23 (s, 2H), 4.79 (dd, J=16.2, 6.1 Hz, 4H), 4.49 (s, 2H), 3.66 (s, 8H), 3.03 (s, 3H).

Step 9

5-(4-Methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester

(393) DCM (33 L) and N-methylpiperazine (21.45 L, 193.4 mol) were stirred at 25° C. and the solution from step 8 added over a minimum of 30 mins such that temperature 20-30° C. After stirring the solution for a further 30 mins water (55 L) was added and the organic phase was washed with water (2×55 L). The product was extracted into 0.8M HCl (66 L) and the layers separated. The aqueous phase was washed with DCM (55 L) and then basified with 2M NaOH to pH 10-11 and the product was extracted into EtOAc (2×55 L). The combined organic phase were filtered to remove solids and the evaporated followed by azeotroping with toluene and drying to constant weight to give the title compound, 6.63 kg (47% yield, 98% pure by hplc).

Step 10

5-(4-Methyl-piperazin-1-ylmethyl)-2,3-dihydro-1H-isoindole

(394) To a degassed solution of 5-(4-Methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester (Step 9, 1.3 Kg, 3.55 mol) dissolved in EtOH (13 L) was added 10% Pd/C (0.065 Kg). Hydrogen was passed through the mixture at 30° C. for 4 h or until complete by NMR. The solution was then stirred for 1 h under an atmosphere of N.sub.2 and then filtered to remove the catalyst through a GF/F filter followed by filtration through a Cuno filter. The filtrate was evaporated to a small volume, azeotroped with toluene (3.9 L) and dried to constant weight yielding the title compound as a red/black oily solid (0.78 Kg) which was stored under nitrogen until required.

Step 11

(2,4-Bis-benzyloxy-5-isopropenyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(395) 1,1′-Carbonyldiimidazole (4.82 Kg, 29.8 mol) was added to a solution of 2,4-bis-benzyloxy-5-isopropenyl-benzoic acid (10.58 Kg, 28.3 mol, step 5) in DMF (21.2 L) at 25° C. After 20 mins at 25° C. a solution of 5-(4-Methyl-piperazin-1-ylmethyl)-2,3-dihydro-1H-isoindole (7.2 Kg, 31.1 mol, step 10) in DMF (7.2 L) maintaining a temperature below 35° C. and the solution stirred at 25° C. for a minimum of 12 h. The solid which had formed was removed by filtration, washed with isopropyl acetate (2×21.6 L) and dried at 35° C. to constant weight to give the title compound 8.7 Kg (77% yield, purity by hplc 97.5%).

Step 12

(2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone

(396) The product from Step 11 (0.9 Kg, 1.53 mol) was dissolved in isopropanol (6.8 L) and water (1.04 L) and after purging with N.sub.2 10% Pd/C (90 g) and K.sub.2CO.sub.3 (0.212 Kg, 1.53 mol) were added and the suspension was hydrogenated for 60 to 70 mins under an 3 Barr pressure of H.sub.2. The solution was diluted with water (0.5 L) and filtered. To the filtrate was added aqueous HCl (30% hydrochloric acid, 0.85 Kg diluted with water 5.42 Kg) and the solution was concentrated at 60° C. under vacuum (removing 10 L isopropanol). Water (0.45 L) was added to the solution and concentration continued (until a further 10 L isopropanol had been removed). The aqueous phase was washed with EtOAc (4.61 L), diluted with acetonitrile (4.06 L) and neutralized to pH 7.5-8.5 by addition of conc ammonia solution (0.35 Kg). The suspension was stirred for 2.5 h and then the solid was removed by filtration. The residue was washed with acetonitrile (2×0.8 L) and dried at 40° C. to constant weight to give the title compound 588 g (94% yield).

Step 13

(2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone L-lactate salt (form FL1)

(397) The product of Step 12 (646 g, 1.58 mol) was dissolved in ethanol (5.17 L) and the solution filtered. A solution of L-lactic acid (142 g, 1.58 mol) dissolved in ethanol (2.59 L) was filtered and added to the solution of the filtered solution (above) and then to the mixture was added EtOAc (7.75 L). The suspension was stirred at RT for 12 h and then cooled to 5° C. for a further 2 h. The solid which had formed was removed by filtration, washed with EtOAc (2×2.58 L) and heptane (2×1.94 L) and dried to constant weight at 35° C. giving the title compound (581 g, 74% yield).

Example 83

Alternative synthesis of 5-(4-Methyl-piperazin-1-ylmethyl)-2,3-dihydro-1H-isoindole

83A. Synthesis of 1-methyl-4-prop-2-ynyl-piperazine

(398) ##STR00286##

(399) To 1-methylpiperazine (37.7 ml, 337 mmol) and K.sub.2CO.sub.3 (46.6 g, 337 mmol) in acetone (380 ml) was added propargyl bromide (25 ml, 225 mmol, 80% in toluene) in acetone (70 ml) dropwise at 0° C. under N.sub.2. The internal temperature of the reaction was kept <10° C. The reaction was stirred at room temperature for 3 hours. The reaction was filtered, and the salts were washed with small portions of acetone (×2). The filtrates were combined evaporated to concentration (gently). To the residue was added water and the product was extracted with DCM (×3). The combined organic layers were washed with brine and dried over MgSO.sub.4. The product was filtered and evaporated to dryness to yield 1-methyl-4-prop-2-ynyl-piperazine as a yellow oil.

83B. Synthesis of 5-(4-Methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindole-2-carboxylic acid tert-butyl ester

(400) ##STR00287##

(401) A solution of N-boc-dipropargylamine (36.3 ml, 226 mmol, 86% pure) in EtOAc (30 ml) was made up and degassed by bubbling through N.sub.2, in a separating funnel. Tris(triphenylphosphine)rhodium(1) chloride (1.39 g, 1.50 mmol, 1 mol %) was added to pre-degassed EtOAc (15 ml) in a second separating funnel. (NB CpRu(COD)Cl) can also be used as an alternative catalyst).

(402) In the main reactor flask, 1-propargyl-4-methylpiperazine (32.3 ml, 150 mmol, 90% pure) was diluted with EtOAc (75 ml) and was degassed by bubbling N.sub.2 through the mixture The mixture was cooled in a ice-water bath and then the tris(triphenylphosphine)rhodium(1) chloride (1.39 g, 1 mol %) in EtOAc was added. Slow addition of N-boc-dipropargylamine/EtOAc was undertaken to yield a mild exotherm. The internal temperature rose to 25° C. and remained at this temperature. After addition was approximately one third complete (˜45 minutes), the exotherm tailed off (despite the continual slow addition of N-boc-dipropargylamine/EtOAc). Another portion of tris(triphenylphosphine) rhodium(1) chloride catalyst (1.39 g, 1 mol %) in EtOAc (15 ml, pre-degassed) was made up and added very slowly to the reaction. After a couple of minutes a new exotherm started and grew to 30° C. The reaction temperature was cooled gently by the addition of a small amount of ice to the water bath. Once the exotherm began to subside, slow addition of N-boc-dipropargylamine/EtOAc was continued. The entire addition was carried out over a 2 hour period. The reaction mixture was then left at room temperature overnight before diluting with EtOAc and washing with NH.sub.4Cl (×2) (aqueous, saturated) to remove excess 1-propargyl-4-methylpiperazine. The mixture was diluted with a small amount of water to dissolve the salts. The organic layer was washed with water, brine and dried over MgSO.sub.4. The product was filtered and evaporated to dryness to leave a brown oil.

(403) To the oil residue obtained was added n-heptane. The oil/heptane was left to stand (˜10 minutes) until a red precipitate formed. The precipitate was filtered and washed with fresh n-heptane (×2). The filtrates were dried to yield the product as a red oil.

(404) The desired product was further purified by forming the toluenesulphonic acid (TsOH) salt. Thus, the crude product was taken up in MeOH (20 ml) and the TsOH.H.sub.2O (1 eq to estimated purity by NMR) was added. The solution was evaporated to dryness, and then dissolved in toluene (×1) and re-evaporated. The resulting product was taken up in ether. After a few minutes, a precipitate and solution formed. The precipitate was filtered and washed with more ether (×2) until the filtrate was colourless. The yellow solid was dried to yield the product as the TsOH salt. MS: [M+H].sup.+ 332.

83C. Synthesis of 5-(4-Methyl-piperazin-1-ylmethyl)-2,3-dihydro-1H-isoindole

(405) ##STR00288##

(406) The isoindoline tosylate salt was taken up in DCM (0.3 M) and TFA (12 eq.) added slowly at 0° C. The reaction was stirred overnight at room temperature. The reaction was evaporated to dryness and then with toluene/MeOH(×3) to yield the product as a mixture of acid addition salts. MS: [M+H].sup.+ 232.

(407) The compound of Example 83C can be used in the method of Example 80 Step 12.

Example 84

Alternative synthesis of 5-hydroxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester

84A, Methyl 2-benzyl-2,3-dihydro-1H-isoindole-5-carboxylate

(408) ##STR00289##

(409) Benzylamine (3.21 g, 30.0 mmol) in anhydrous tetrahydrofuran (25 ml) was added to a stirred mixture of methyl 3,4-bis-(bromomethyl)benzoate (9.66 g, 30.0 mmol) (obtained from Fluorochem) and triethylamine (9 ml, 64.7 mmol) in anhydrous tetrahydrofuran (50 ml) and the resulting mixture was stirred at room temperature for 3 hours. The solvent was removed in vacuo at 40° C. and the residue partitioned between ethyl acetate (100 ml) and water (100 ml). The organic layer was washed with a further portion of water (100 ml), separated and the solvent removed in vacuo at 40° C. to afford methyl 2-benzyl-2,3-dihydro-1H-isoindole-5-carboxylate as a pale orange solid that was used immediately without further purification as described below. .sup.1H NMR (DMSO-d.sub.6) 7.82 (2H, m), 7.40-7.25 (6H, m), 3.90 (3H, s), 3.88 (2H, s), 3.84 (4H, s). MS: [M+H].sup.+ 268.

84B. (2-Benzyl-2,3-dihydro-1H-isoindol-5-yl)-methanol

(410) ##STR00290##

(411) Methyl 2-benzyl-2,3-dihydro-1H-isoindole-5-carboxylate (from above) was dissolved in anhydrous tetrahydrofuran (75 ml) and added dropwise over 15 minutes to a rapidly stirred suspension of lithium aluminium hydride (1.71 g, 45.0 mmol) in anhydrous tetrahydrofuran (75 ml). The mixture was stirred at room temperature for 2 hours whereupon excess lithium aluminium hydride was destroyed by the slow dropwise addition of 1M sodium sulphate solution (12 ml). The solids were removed by filtration, rinsed with ethyl acetate (2×50 ml) and sucked dry. The solvent was removed in vacuo to afford (2-benzyl-2,3-dihydro-1H-isoindol-5-yl)-methanol (7.15 g, 99%) as a tan solid. .sup.1H NMR (DMSO-d.sub.6) 7.40-7.30 (4H, m), 7.28 (1H, m), 7.17-7.10 (3H, m), 5.10 (1H, t), 4.47 (2H, d), 3.85 (2H, s), 3.82 (2H, s), 3.80 (2H, s). MS: [M+H].sup.+ 240.

84C. (2,3-Dihydro-1H-isoindol-5-yl)-methanol

(412) ##STR00291##

(413) 10% Palladium on activated carbon (200 mg) was added to a solution of (2-benzyl-2,3-dihydro-1H-isoindol-5-yl)-methanol (2.39 g, 10.0 mmol) in ethanol (60 ml) and the resulting mixture was placed in a Parr apparatus, heated to 50° C. and shaken under a hydrogen atmosphere at 60 psi for 30 hours. Upon cooling to room temperature the mixture was filtered under gravity, the solids were rinsed with ethanol (2×10 ml) and the solvent removed in vacuo to afford (2,3-dihydro-1H-isoindol-5-yl)-methanol (1.49 g, 100%) as an off-white solid. .sup.1H NMR (DMSO-d.sub.6) 7.20 (1H, s), 7.18 (1H, d), 7.12 (1H, d), 5.10 (1H, br s), 4.46 (2H, s), 4.05 (4H, s). MS: [M+H].sup.+ 150.

84D. 5-Hydroxymethyl-1,3-dihydro-isoindole-2-carboxylic acid benzyl ester

(414) ##STR00292##

(415) A mixture of (2,3-dihydro-1H-isoindol-5-yl)-methanol (1.34 g, 9.0 mmol) in anhydrous tetrahydrofuran (50 ml) was warmed gently to aid dissolution and allowed to cool to room temperature. Triethylamine (1.5 ml, 10.8 mmol) was added and the stirred mixture was treated dropwise with benzyl chloroformate (1.35 ml, 9.5 mmol) and stirred at room temperature for 3 hours. The solvent was removed in vacuo and the residue partitioned between ethyl acetate (30 ml) and 2M hydrochloric acid (30 ml). The organic layer was washed with water (30 ml), separated and the solvent removed in vacuo to afford a pink oil that solidified upon standing. The solids were triturated with 10% ethyl acetate in hexane (10 ml), filtered, rinsed with heptane (10 ml) and sucked dry to afford the title compound (2.5 g, 98%) as a pale pink solid. .sup.1H NMR (DMSO-d.sub.6) 7.45-7.21 (8H, m), 5.20 (1H, t), 5.17 (2H, s), 4.71 (2H, br s), 4.64 (2H, br s), 4.50 (2H, d). MS: [M+H].sup.+ 284.

(416) The title compound can be used in Step 9 of Example 80.

Example 85

Crystal Structure Studies

(417) The compound of formula (VIII), 2,4-Dihydroxy-5-isopropyl-phenyl)-[5-(4-methyl-piperazin-1-ylmethyl)-1,3-dihydro-isoindol-2-yl]-methanone, and its salts exist in a number of different crystalline forms. The L-lactate salts have been identified and characterised using the methods described below.

(418) General Methods

(419) Single Crystal Diffraction Methodology

(420) Crystallographic data were collected at room temperature (20° C.) using synchrotron radiation (λ=0.775 Å) from ESRF ID23.1 beamline equipped with φ goniometer and an ADSC Quantum 315 CCD detector. Images were collected in two φ scans with φ=0-180° and Δφ=1°, one with high radiation dose and one with low dose. Detector to crystal distance was 110 mm. Data collection was controlled by ProDC software and images were processed and scaled by Dtrek.

(421) The crystal structures were solved using direct methods implemented in SHELXS-97 and refined by SHELXL-97. Hydrogen atoms were generated on geometrical grounds while the location of heteroatom bound hydrogen atoms was confirmed by inspection of Fo-Fc difference maps. The positional and thermal parameters of hydrogen atoms were constricted to ride on corresponding non-hydrogen atoms. The thermal motion of non-hydrogen atoms was modelled by anisotropic thermal factors.

(422) Powder Diffraction Methodology

(423) Samples for X-ray powder diffraction (XRPD) data collection were gently ground by marble mortar and loaded into a crystallographic capillary (from Hampton Research, Quartz or Glass Type 10, 0.4 or 0.7 mm diameter). Diffraction patterns were collected at room temperature using CuKα radiation (λ=1.5418 Å) from a Rigaku rotating anode RU3HR, Osmic blue confocal optics, ¼ c goniometer and a Rigaku HTC image plate detector. 2D images were collected while spinning φ axis with a detector to crystal distance of 250 mm. Data collection was controlled by CrystalClear software and 2D images were converted to 1D plot (2θ vs. Intensity) by Datasqueeze (intensity averaged over the azimuthal angle 0≦χ≦360° for 2θ range 3-30° in 0.02° steps). In house program AstexXRPD was used for manipulation and visualisation of 1D XRPD patterns.

(424) Determination of Salt Stoichiometry by Titration Experiments

(425) In the following examples, where they relate to salts and the stoichiometry of the salt is given, the stoichiometry was determined using the following titration method.

(426) A solution (KCl/HCl solution) of 150 mM KCl and 20 mM HCl was freshly prepared for each batch of titration experiments. An aliquot of 1 ml of the solution was titrated and the potentiometric titration curve thus produced was used as the control curve. All titrations were performed at 25° C. and with 300 mM KOH in 2 μl steps using a Mettler Toledo MP220 pH meter. Electrode potential readings for 4 standard buffers were recorded before and after daily batch of measurement. Samples of Compound (VIII) salts of (1-3 mg) were dissolved in 1 ml of KCl/HCl solution and titrated with vigorous stirring using a small magnetic stirrer. The recorded electrode potentials were converted into pH values using a calibration curve from the 4 standard buffers. Sample and control titration data were processed to produce a Bjerrum plot in the pH range 2-12. The Bjerrum plot calculation and analysis method is described in the review “Physicochemical Profiling (Solubility, Permeability and Charge State)”, A. Avdeef (Current Topics in Medicinal Chemistry 2001, p277-351).

(427) The stoichiometry of the Compound (VIII) salts was deduced from the starting nH (number of protons at pH=2), (i.e. free base starts with −2 protons, mono-salt with −1 protons (Compound (VIII).sup.+ acid.sup.−)), while double salts (Compound (VIII).sup.2+ acid.sup.2− or Compound (VIII).sup.2+ 2* acid.sup.−) start at nH=0.

(428) Compound (VIII) L-Lactate 1:1 Salt Crystal Forms

(429) (85A) Compound (VIII) L-Lactate—Form FL1

(430) The L-Lactate salt form FL1 was prepared as described in Example 81 above.

(431) Form FL1 is stable in air and at 40° C. and 75% RH for at least one month. The XRPD pattern for form FL1 is shown in FIG. 1 and the main peaks are listed in Table EX85A.

(432) TABLE-US-00018 TABLE EX85A Main XRPD peaks for Compound (VIII) Lactate - Form FL1 2θ/° d/Å I/% 6.18 14.30 15 6.53 13.52 50 8.39 10.54 19 11.08 7.98 7 13.10 6.75 85 14.13 6.26 33 14.40 6.15 23 15.21 5.82 4 16.21 5.46 6 16.81 5.27 100 17.22 5.15 45 18.65 4.75 23 19.52 4.54 33 19.82 4.48 34 20.49 4.33 7 20.76 4.27 13 21.13 4.20 17 22.02 4.03 12 22.33 3.98 44 22.84 3.89 40 23.09 3.85 25 23.94 3.71 14 25.19 3.53 7 26.41 3.37 14 26.95 3.31 5 27.81 3.21 14
(85B) Compound (VIII) L-Lactate—Form FL2

(433) Form FL2 was observed in precipitation experiments of methanol solutions of form FL1. Single crystal X-ray analysis showed that form FL2 is hydrated. It is nominally a tri-hydrate because there are 3 crystal water positions in the asymmetric unit, but they are not 100% occupied at room temperature and laboratory humidity. A saturated solution of form FL1 in methanol:water 9:1 was prepared at room temperature. Slow precipitation with approximately 4 volumes of acetone gave form FL2 which is stable in air. The XRPD pattern for form FL2 is shown in FIG. 2 and the main peaks are listed in Table 14 below. A crystal packing diagram is shown in FIG. 3 and the atom coordinates are listed in Table EX85B(i) below.

(434) TABLE-US-00019 TABLE EX85B(i) Main XRPD peaks for Compound (VIII) Lactate salt - form FL2 2θ/° d/Å I/% 8.03 11.00 29 10.71 8.26 53 11.98 7.38 90 13.13 6.74 49 15.39 5.75 29 16.09 5.50 32 16.61 5.33 42 17.26 5.13 37 18.17 4.88 20 18.82 4.71 56 20.40 4.35 40 21.01 4.22 49 21.53 4.12 27 22.34 3.98 100 22.56 3.94 73 23.71 3.75 82 24.30 3.66 8 24.65 3.61 12 26.56 3.35 13 27.70 3.22 21 28.29 3.15 16

(435) TABLE-US-00020 TABLE EX85B(II) Unit cell parameters and coordinates in cif format for crystal structure of Compound (VIII) Lactate salt - form FL2 space group: P2.sub.1 unit cell at 293K with a, b, c & β having 5% s.u.: a = 5.8 b = 16.6 c = 14.9 beta = 98 alpha = gamma = 90 Coordinates in cif format: loop.sub.— _atom_site_label _atom_site_type_symbol _atom_site_fract_x _atom_site_fract_y _atom_site_fract_z _atom_site_U_iso_or_equiv _atom_site_adp_type _atom_site_occupancy _atom_site_symmetry_multiplicity _atom_site_calc_flag _atom_site_refinement_flags _atom_site_disorder_assembly _atom_site_disorder_group C1 C −0.643(2) 1.1037(6) 0.6763(7) 0.097(3) Uani 1 1 d . . . H1A H −0.6995 1.0577 0.6395 0.117 Uiso 1 1 calc . . . H1B H −0.5231 1.1308 0.6484 0.117 Uiso 1 1 calc . . . N2 N −0.5563(16) 1.0791(5) 0.7694(6) 0.096(2) Uani 1 1 d . . . C3 C −0.692(3) 1.1148(8) 0.8352(8) 0.124(4) Uani 1 1 d . . . H3A H −0.7713 1.0734 0.8651 0.148 Uiso 1 1 calc . . . H3B H −0.5925 1.1454 0.8805 0.148 Uiso 1 1 calc . . . C4 C −0.8553(19) 1.1667(7) 0.7825(7) 0.094(3) Uani 1 1 d . . . C5 C −0.8393(19) 1.1609(6) 0.6900(7) 0.092(3) Uani 1 1 d . . . C6 C −1.036(3) 1.2141(8) 0.8083(8) 0.110(3) Uani 1 1 d . . . H6 H −1.0636 1.2139 0.8682 0.132 Uiso 1 1 calc . . . C7 C −1.172(2) 1.2611(8) 0.7456(8) 0.105(3) Uani 1 1 d . . . C8 C −1.145(2) 1.2560(8) 0.6564(9) 0.111(3) Uani 1 1 d . . . H8 H −1.2387 1.2867 0.6138 0.133 Uiso 1 1 calc . . . C9 C −0.979(2) 1.2053(9) 0.6287(7) 0.109(3) Uani 1 1 d . . . H9 H −0.9640 1.2017 0.5677 0.130 Uiso 1 1 calc . . . C10 C −1.3561(18) 1.3173(8) 0.7739(9) 0.106(3) Uani 1 1 d . . . H10A H −1.4455 1.3402 0.7202 0.127 Uiso 1 1 calc . . . H10B H −1.4617 1.2864 0.8055 0.127 Uiso 1 1 calc . . . N11 N −1.2550(14) 1.3836(6) 0.8332(6) 0.096(2) Uani 1 1 d . . . C12 C −1.1136(17) 1.4353(6) 0.7839(7) 0.091(3) Uani 1 1 d . . . H12A H −1.2098 1.4591 0.7324 0.109 Uiso 1 1 calc . . . H12B H −0.9935 1.4035 0.7615 0.109 Uiso 1 1 calc . . . C13 C −1.0015(17) 1.5021(7) 0.8462(8) 0.100(3) Uani 1 1 d . . . H13A H −0.8991 1.4783 0.8961 0.121 Uiso 1 1 calc . . . H13B H −0.9092 1.5368 0.8128 0.121 Uiso 1 1 calc . . . N14 N −1.1853(15) 1.5509(5) 0.8822(6) 0.094(2) Uani 1 1 d . . . H14 H −1.2741 1.5755 0.8352 0.113 Uiso 1 1 calc . . . C15 C −1.3350(18) 1.4966(7) 0.9279(7) 0.095(3) Uani 1 1 d . . . H15A H −1.4599 1.5276 0.9479 0.114 Uiso 1 1 calc . . . H15B H −1.2441 1.4730 0.9808 0.114 Uiso 1 1 calc . . . C16 C −1.4358(17) 1.4308(7) 0.8658(8) 0.098(3) Uani 1 1 d . . . H16A H −1.5310 1.3959 0.8977 0.117 Uiso 1 1 calc . . . H16B H −1.5346 1.4542 0.8148 0.117 Uiso 1 1 calc . . . C17 C −1.068(2) 1.6140(9) 0.9439(9) 0.119(4) Uani 1 1 d . . . H17A H −1.1835 1.6447 0.9694 0.178 Uiso 1 1 calc . . . H17B H −0.9807 1.6492 0.9103 0.178 Uiso 1 1 calc . . . H17C H −0.9658 1.5886 0.9916 0.178 Uiso 1 1 calc . . . C18 C −0.382(2) 1.0287(9) 0.7999(8) 0.113(4) Uani 1 1 d . . . O19 O −0.345(2) 1.0216(8) 0.8837(6) 0.156(4) Uani 1 1 d . . . C20 C −0.228(2) 0.9847(6) 0.7418(7) 0.096(3) Uani 1 1 d . . . C21 C −0.069(3) 0.9286(9) 0.7863(9) 0.119(4) Uani 1 1 d . . . C22 C 0.064(2) 0.8867(9) 0.7367(9) 0.114(4) Uani 1 1 d . . . H22 H 0.1812 0.8547 0.7669 0.137 Uiso 1 1 calc . . . C23 C 0.038(2) 0.8879(7) 0.6447(8) 0.097(3) Uani 1 1 d . . . C24 C −0.1201(18) 0.9425(7) 0.5972(8) 0.096(3) Uani 1 1 d . B . C25 C −0.253(2) 0.9882(7) 0.6463(8) 0.100(3) Uani 1 1 d . . . H25 H −0.3632 1.0228 0.6160 0.120 Uiso 1 1 calc . . . O26 O −0.036(2) 0.9229(9) 0.8775(6) 0.169(5) Uani 1 1 d . . . H26 H −0.1427 0.9456 0.8980 0.253 Uiso 1 1 calc R . . O27 O 0.1658(15) 0.8404(5) 0.5948(6) 0.118(3) Uani 1 1 d . . . H27 H 0.2091 0.7999 0.6238 0.176 Uiso 1 1 calc R . . C28 C −0.141(4) 0.9478(11) 0.4948(10) 0.138(6) Uani 1 1 d . . . H28 H −0.0894 0.8953 0.4750 0.166 Uiso 1 1 calc . A 1 C29 C −0.029(11) 1.004(4) 0.449(3) 0.24(3) Uani 0.58(6) 1 d P B 1 H29A H −0.0741 0.9976 0.3847 0.363 Uiso 0.58 1 calc P B 1 H29B H 0.1361 0.9972 0.4628 0.363 Uiso 0.58 1 calc P B 1 H29C H −0.0703 1.0575 0.4662 0.363 Uiso 0.58 1 calc P B 1 C30 C −0.417(7) 0.950(3) 0.4621(19) 0.159(19) Uani 0.58(6) 1 d P B 1 H30A H −0.4911 0.9083 0.4918 0.239 Uiso 0.58 1 calc P B 1 H30B H −0.4462 0.9424 0.3978 0.239 Uiso 0.58 1 calc P B 1 H30C H −0.4773 1.0016 0.4772 0.239 Uiso 0.58 1 calc P B 1 C29 C −0.156(11) 1.040(2) 0.465(2) 0.14(2) Uani 0.42(6) 1 d P B 2 H29D H −0.0071 1.0655 0.4814 0.215 Uiso 0.42 1 calc P B 2 H29E H −0.2703 1.0675 0.4943 0.215 Uiso 0.42 1 calc P B 2 H29F H −0.1983 1.0438 0.4003 0.215 Uiso 0.42 1 calc P B 2 C30 C −0.295(12) 0.897(4) 0.446(2) 0.150(19) Uani 0.42(6) 1 d P B 2 H30D H −0.3403 0.9185 0.3870 0.224 Uiso 0.42 1 calc P B 2 H30E H −0.4300 0.8910 0.4766 0.224 Uiso 0.42 1 calc P B 2 H30F H −0.2234 0.8451 0.4418 0.224 Uiso 0.42 1 calc P B 2 O1L O −1.5549(12) 1.6174(6) 0.7786(6) 0.124(3) Uani 1 1 d . . . O2L O −1.7419(12) 1.7087(6) 0.6890(7) 0.125(3) Uani 1 1 d . . . C1L C −1.5569(17) 1.6742(7) 0.7238(8) 0.098(3) Uani 1 1 d . . . C2L C −1.3365(17) 1.6989(8) 0.6926(9) 0.108(4) Uani 1 1 d . . . H2L H −1.3065 1.7549 0.7117 0.129 Uiso 1 1 calc . . . C3L C −1.355(2) 1.6971(12) 0.5917(11) 0.143(5) Uani 1 1 d . . . H3L1 H −1.2130 1.7162 0.5734 0.214 Uiso 1 1 calc . . . H3L2 H −1.4813 1.7312 0.5662 0.214 Uiso 1 1 calc . . . H3L3 H −1.3842 1.6429 0.5706 0.214 Uiso 1 1 calc . . . O3L O −1.1538(13) 1.6538(7) 0.7316(8) 0.150(4) Uani 1 1 d . . . H3L H −1.0243 1.6711 0.7191 0.224 Uiso 1 1 d . . . O1W O −0.448(6) 1.237(6) 1.045(2) 0.45(5) Uani 0.78(6) 1 d P . . . O2W O −0.021(15) 0.8037(17) 0.9990(19) 0.74(7) Uani 1 1 d . . . O3W O −0.35(3) 0.773(9) 0.953(15) 0.77(8) Uani 0.22(6) 1 d P . . .
(85C) Compound (VIII) L-Lactate—Form FL3

(436) Form FL3 was observed in precipitation experiments of THF solutions of form FL1. Form FL3 transforms in air into form FL1. A saturated solution of form FL1 in THF was prepared at room temperature. Slow precipitation with approximately 4 volumes of heptane gave form FL3. The XRPD pattern of a fresh sample of form FL3 is shown in FIG. 4 and the main peaks are listed in Table EX85C(i) below. A sample of FL3 was dried in air for 2 days after which XRPD analysis showed that conversion to form FL1 had occurred.

(437) TABLE-US-00021 TABLE EX85C Main XRPD peaks for Compound (VIII) Lactate salt - form FL3 2θ/° d/Å I/% 5.53 15.98 100 8.36 10.56 5 11.07 7.98 41 13.16 6.72 12 13.85 6.39 8 16.69 5.31 39 17.17 5.16 21 18.00 4.92 49 18.49 4.80 11 19.28 4.60 14 19.79 4.48 5 20.34 4.36 7 21.05 4.22 21 21.47 4.14 7 21.93 4.05 4 22.47 3.95 16 22.84 3.89 23 24.56 3.62 4 26.28 3.39 6 27.06 3.29 3 27.47 3.24 3 29.11 3.07 6

BIOLOGICAL ACTIVITY

Example 86

Isothermal Titration Calorimetry

(438) The ability of the compounds of the invention to bind to human Hsp90 proteins was determined using isothermal titration calorimetry.

(439) Isothermal titration calorimetry (ITC) experiments were performed with a VP-ITC titration calorimeter (Microcal Inc., Northampton, Mass., USA). Cloning, expression, and purification of the Human Hsp90a N-terminal domain were performed according to published methods (Jez, J. M. et al, Chem. Biol. 2003 April; 10(4):361-8.) Solutions of the human Hsp90α N-terminal domain and compound were prepared in a buffer comprising 25 mM Tris, 100 mM NaCl, 1 mM MgCl.sub.2, 1 mM TCEP, 5% DMSO, pH 7.4. All solutions were filtered and degassed prior to a titration being carried out. The enthalpy change resulting from each injection of ligand was obtained through integration of the calorimetric signal. Data were analysed using Origin 7.0 (Microcal Software Inc., Northampton, Mass.). Heats of dilution were estimated using the final injections of each individual titration and subtracted before data fitting. Different ITC experimental formats were employed in order to obtain compound dissociation constants (Kd's) over a wide range of affinities. For weakly binding compounds a low c-value ITC method was used (Turnbull W. B. & Daranas A. H. J. Am. Chem. Soc. 2003 Dec. 3; 125(48):14859-66) in which the protein was present at 10-20 μM in the calorimetric cell and the compound concentration was 1-20 mM in the injection syringe. In this type of experiment the stoichiometry parameter (N) was locked at 1 for data fitting. For Kd's in the 20-0.004 μM range the experiment was configured such that the binding site concentration divided by the Kd (c-value) was between 5 and 1000. For the majority of these experiments the protein concentration in the calorimetric cell was in the range 4-100 μM and the ligand concentration in the injection syringe ranged from 50-1500 μM. In rare cases where compound solubility was limiting, the compound solution was placed in the calorimetric cell and titrated with protein from the injection syringe, maintaining a c-value between 5 and 1000. Competition ITC experiments were used to access Kd's <4 nM by performing the titration in the presence of a weaker binding competitor according to the method described in Sigurskjold B. W. Anal Biochem. 2000 Jan. 15; 277(2):260-6.

(440) The compounds of examples 5, 10, 11, 12, 13, 14, 16, 17, 18, 19, 21, 22, 23, 25, 26, 27, 28, 29, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 48, 49, 50, 51, 52, 53, 54, 55, 59, 60, 61, 62, 63, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74 and 75 were tested and were found to have K.sub.d values of less than 1 micromolar.

(441) The compounds of examples 5, 10, 12, 13, 14, 16, 17, 18, 19, 21, 22, 23, 25, 26, 27, 28, 29, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 48, 49, 50, 51, 53, 54, 55, 59, 60, 61, 62, 63, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74 and 75 have K.sub.d values of less than 0.1 micromolar and most of these compounds have K.sub.d values of less than 0.01 micromolar.

Example 87

(442) Methods of Testing for Pain Reducing or Pain Preventing Activity

(443) (I) Inflammatory Hyperalgesia Test

(444) Mechanical hyperalgesia can be examined in a rat model of inflammatory pain. Paw withdrawal thresholds to an increasing pressure stimulus are measured by the Randal-Sellito technique using an analgesymeter (Ugo Basile, Milan), in naïve animals prior to an intraplantar injection of complete Freund's complete adjuvant (FCA) into the left hind paw. 24 h later paw withdrawal thresholds are measured again prior to (predose) and then from 10 min to 6 h following drug or vehicle administration. Reversal of hyperalgesia in the ipsilateral paw is calculated according to the formula:

(445) $\%\mathrm{reversal}=\frac{\mathrm{postdose}\mathrm{threshold}-\mathrm{predose}\mathrm{threshold}}{\mathrm{naive}\mathrm{threshold}-\mathrm{predose}\mathrm{threshold}}\times 100$
(ii) Neuropathic Hyperalgesia Test

(446) Mechanical hyperalgesia can be examined in a rat model of neuropathic pain induced by partial ligation of the left sciatic nerve. Approximately 14 days following surgery mechanical withdrawal thresholds of both the ligated (ipsilateral) and non-ligated (contralateral) paw are measured prior to (predose) and then from 10 min to 6 h following drug or vehicle administration. Reversal of hyperalgesia at each time point is calculated according to the formula:

(447) $\%\mathrm{reversal}=\frac{\begin{array}{c}\mathrm{ipsilateral}\mathrm{threshold}\mathrm{postdose}-\\ \mathrm{ipsilateral}\mathrm{threshold}\mathrm{predose}\end{array}}{\begin{array}{c}\mathrm{contralateral}\mathrm{threshold}\mathrm{predose}-\\ \mathrm{ipsilateral}\mathrm{threshold}\mathrm{predose}\end{array}}\times 100$

(448) All experiments are carried out using groups of 6 animals. Stock concentrations of drugs are dissolved in distilled water and subsequent dilutions were made in 0.9% saline for subcutaneous administration in a volume of 4 mlkg.sup.−1. All drugs are made up in plastic vials and kept in the dark.

(449) Statistical analysis are carried out on withdrawal threshold readings (g) using ANOVA with repeated measures followed by Tukey's HSD test. Efficacy refers to the maximal reversal of hyperalgesia observed at the doses used.

(450) (iii) Testing the Effects of Compounds of Formula (0) a Rat Model of Bone Cancer Pain

(451) Adult female rats are given intra-tibial injections of MRMZ-1 rat mammary gland carcinoma cells (3 μl, 10.sup.7 cells/ml). The animals typically gradually develop mechanical hyperalgesia, mechanical allodynia (skin sensitivity to non-noxious stimuli) and hind limb sparing, beginning on day 12-14 following cell injection. A compound of formula (0) (e.g. at a dose of 10 and 30 μg/kg s.c.) is administered 3 times a week from the day of cell injection, and the extent of inhibition of hind limb sparing and mechanical allodynia is determined in comparison to vehicle-treated controls.

Example 88

(452) Determination of Antifungal Activity

(453) The antifungal activity of the compounds of the formula (I) is determined using the following protocol.

(454) The compounds are tested against a panel of fungi including Candida parapsilosis, Candida tropicalis, Candida albicans-ATCC 36082 and Cryptococcus neoformans. The test organisms are maintained on Sabourahd Dextrose Agar slants at 4° C. Singlet suspensions of each organism are prepared by growing the yeast overnight at 27° C. on a rotating drum in yeast-nitrogen base broth (YNB) with amino acids (Difco, Detroit, Mich.), pH 7.0 with 0.05 morpholine propanesulphonic acid (MOPS). The suspension is then centrifuged and washed twice with 0.85% NaCl before sonicating the washed cell suspension for 4 seconds (Branson Sonifier, model 360, Danbury, Conn.). The singlet blastospores are counted in a haemocytometer and adjusted to the desired concentration in 0.85% NaCl.

(455) The activity of the test compounds is determined using a modification of a broth microdilution technique. Test compounds are diluted in DMSO to a 1.0 mg/ml ratio then diluted to 64 μg/ml in YNB broth, pH 7.0 with MOPS (Fluconazole is used as the control) to provide a working solution of each compound. Using a 96-well plate, wells 1 and 3 through 12 are prepared with YNB broth, ten fold dilutions of the compound solution are made in wells 2 to II (concentration ranges are 64 to 0.125 μg/ml). Well 1 serves as a sterility control and blank for the spectrophotometric assays. Well 12 serves as a growth control. The microtitre plates are inoculated with 10 μl in each of well 2 to 11 (final inoculum size is 10.sup.4 organisms/ml). Inoculated plates are incubated for 48 hours at 35° C. The MIC values are determined spectrophotometrically by measuring the absorbance at 420 nm (Automatic Microplate Reader, DuPont Instruments, Wilmington, Del.) after agitation of the plates for 2 minutes with a vortex-mixer (Vorte-Genie 2 Mixer, Scientific industries, Inc., Bolemia, N.Y.). The MIC endpoint is defined as the lowest drug concentration exhibiting approximately 50% (or more) reduction of the growth compared with the control well. With the turbidity assay this is defined as the lowest drug concentration at which turbidity in the well is <50% of the control (IC.sub.50). Minimal Cytolytic Concentrations (MCC) are determined by sub-culturing all wells from the 96-well plate onto a Sabourahd Dextrose Agar (SDA) plate, incubating for 1 to 2 days at 35° C. and then checking viability.

Pharmaceutical Formulations

Example 89

(456) (i) Tablet Formulation

(457) A tablet composition containing a compound of the invention is prepared by mixing 50 mg of the compound with 197 mg of lactose (BP) as diluent, and 3 mg magnesium stearate as a lubricant and compressing to form a tablet in known manner.

(458) (ii) Capsule Formulation

(459) A capsule formulation is prepared by mixing 100 mg of a compound of the invention with 100 mg lactose and filling the resulting mixture into standard opaque hard gelatin capsules.

(460) (iii) Injectable Formulation I

(461) A parenteral composition for administration by injection can be prepared by dissolving a compound of the invention (e.g. in a salt form) in water containing 10% propylene glycol to give a concentration of active compound of 1.5% by weight. The solution is then sterilised by filtration, filled into an ampoule and sealed.

(462) (iv) Injectable Formulation II

(463) A parenteral composition for injection is prepared by dissolving in water a compound of the invention (e.g. in salt form) (2 mg/ml) and mannitol (50 mg/ml), sterile filtering the solution and filling into sealable 1 ml vials or ampoules.

(464) v) Injectable formulation III

(465) A formulation for i.v. delivery by injection or infusion can be prepared by dissolving the compound of the invention (e.g. in a salt form) in water at 20 mg/ml. The vial is then sealed and sterilised by autoclaving.

(466) vi) Injectable formulation IV

(467) A formulation for i.v. delivery by injection or infusion can be prepared by dissolving the compound of the invention (e.g. in a salt form) in water containing a buffer (e.g. 0.2 M acetate pH 4.6) at 20 mg/ml. The vial is then sealed and sterilised by autoclaving.

(468) (vii) Subcutaneous Injection Formulation

(469) A composition for sub-cutaneous administration is prepared by mixing a compound of the invention with pharmaceutical grade corn oil to give a concentration of 5 mg/ml. The composition is sterilised and filled into a suitable container.

(470) (viii) Lyophilised Formulation

(471) Aliquots of formulated compound of the invention (I) are put into 50 ml vials and lyophilized. During lyophilisation, the compositions are frozen using a one-step freezing protocol at (−45° C.). The temperature is raised to −10° C. for annealing, then lowered to freezing at −45° C., followed by primary drying at +25° C. for approximately 3400 minutes, followed by a secondary drying with increased steps if temperature to 50° C. The pressure during primary and secondary drying is set at 80 millitorr.

(472) TABLE-US-00022 (ix) 2% Topical Gel Formulation % w/w Compound 2.00 Hydroxypropyl Methyl cellulose (Methocel F4M) 2.50 Polyethyleneoxide (Polyox WSR -205) 0.25 Propylene glycol 10.00 Methylparaben 0.15 Propylparaben 0.05 Purified Water to 100.00

EQUIVALENTS

(473) The foregoing examples are presented for the purpose of illustrating the invention and should not be construed as imposing any limitation on the scope of the invention. It will readily be apparent that numerous modifications and alterations may be made to the specific embodiments of the invention described above and illustrated in the examples without departing from the principles underlying the Invention. All such modifications and alterations are intended to be embraced by this application.