WO2013029648A1

WO2013029648A1 - New aminoacid compounds for treating tumours and autoimmune diseases

Info

Publication number: WO2013029648A1
Application number: PCT/EP2011/064724
Authority: WO
Inventors: Andrea ZACCAGNINI; Mauro STADERINI; Gennaro Carnevale; Alfonso Pompella; Simona PIAGGI
Original assignee: Arke' Organics S.R.L.
Priority date: 2011-08-26
Filing date: 2011-08-26
Publication date: 2013-03-07

Abstract

A compound of formula (I) or a pharmaceutically acceptable salt or solvate thereof, wherein: - R' is H or F or trifluoromethyl; - R'' is OH or O-alkyl; - R''' is H or F or trifluoromethyl or phenyl or pyridyl; - R'''' is methylene or acylsulfanyl or phenacylsulfanyl or aryloylsulfanyl or thien-2-ylsulfanyl, thiazol-2-ylsulfanyl; - n is 0 or 1, as well as pharmaceutical composition containing at least of such compounds for treating neoplasms, including tumours and carcinomas, and autoimmune diseases, including rheumatoid arthritis.

Description

TITLE

NEW AMINOACID COMPOUNDS FOR TREATING TUMOURS AND AUTOIMMUNE DISEASES

DESCRIPTION Field of the invention

The present invention discloses new aminoacid derivatives showing antineoplastic and antiproliferative activity .

Background of the invention - Technical problems

Despite the progresses of the last two decades in the development of chemical entities used in therapeutic regimens for treating neoplasms such as tumours, the unmet clinical needs in this area are still relevant, and the efforts to obtain new drugs are still ongoing.

Summary of the invention

It is therefore a feature of the present invention to provide new compounds having antineoplastic and antiproliferative activity.

It is also a feature of the present invention to provide a composition comprising such compounds.

The present invention covers the use of such compounds and of any pharmaceutically acceptable salt or solvate thereof, or of a composition comprising it/them, to treat neoplasms and autoimmune disorders, in particular rheumatoid arthritis.

These and other objects are achieved by a compound of formula: '

[I]

or a pharmaceutically acceptable salt or solvate thereof, wherein:

— R' is H or F or trifluoromethyl (CF₃) ;

— R' ' is OH or O-alkyl;

— R' ' ' is H or F or trifluoromethyl or phenyl or pyridyl;

— R' ' ' ' is methylene or acylsulfanyl or phenacylsulfanyl, aryloylsulfanyl or thienylsulfanyl or

thiazolylsulfanyl .

— n is 0 or 1.

The disclosed compounds are of possible use for treatment of neoplasms, including tumours and carcinomas, like neoplasms of the genitourinary tract, neoplasms of the central nervous system, neoplasms of the endocrine glands, neoplasms of the head and neck, neoplasms of the respiratory tract including lungs, neoplasms of the alimentary canal, neoplasms of the female reproductive organs, neoplasms of the hematopoietic system such leukaemia, lymphomas and myelomas.

The disclosed compounds are also expected to be useful in treatment of autoimmune diseases, including rheumatoid arthritis .

Some of the compounds of the present invention can be acidic in nature and may form salts by reaction with a variety of inorganic and organic bases.

The pharmaceutical composition may comprise any pharmaceutically acceptable carrier, adjuvant or vehicle.

Pharmaceutically acceptable salts may be obtained by reaction with inorganic bases such as hydroxides, carbonates and bicarbonates of alkaline and alkaline-earth metals such as sodium, potassium, lithium or calcium. Sodium and potassium salts are preferred.

Pharmaceutically acceptable salts may be obtained also by reaction of a compound of formula [I] with primary, secondary or tertiary amines such as methylamine, ethylamine, diethanolamine, isopropylamine, diisopropyl- amine .

Some other compounds of the present invention can be basic in nature and may form pharmaceutically acceptable salts by reaction with hydrochloric acid, hydrobromic acid, phosphoric acid, citric acid, fumaric acid, L- tartaric acid, maleic acid, malonic acid, succinic acid. The hydrochlorides are preferred.

The compounds disclosed in this invention may be prepared by general methods that are well known by the ones skilled in the art. The sequence of the steps of the synthesis depends upon the relative lability of the substituted building blocks.

Compounds of formula [I] in which R' ' ' ' is methylene can be synthesized according to the general scheme:

(ID (HI) (IV) wherein a 2-aminoacrylic acid ester (III) is coupled with the chloride (II) of an optionally substituted phenylacetic or benzoic acid to obtain the final product (IV) . 2-aminoacrylic acid esters are commercially available or can be prepared following the protocols known from the literature (see, for instance, Rao, S. Nagaraja; O'Ferrall, R.A. More, Journal of the American Chemical Society, vol. 112; nb . 7; (1990); p. 2729 -2735) . Compounds of formula [I] in which R' ' ' ' is any of the previously listed group comprised of acylsulfanyl, phenacylsulfanyl, aryloylsulfanyl can be synthesized according to the general scheme:

wherein an L-Cystine diester [V] is treated with an optionally substituted phenylacetic or benzoic acid [VI] and a suitable peptide coupling agent such dicyclohexylcarbodiimide (DCC) , N, N' -carbonyldiimidazole (CDI), 1- ( 3-dimethylaminopropyl ) -3-ethyl carbodiimide (EDC) r 0- (Benzotriazol-l-yl ) -N, N, N ' , N ' -tetramethyluronium tetrafluoroborate (TBTU) . All the reagents are mixed in a suitable solvent, typically dichloromethane, dioxane, chloroform, trifluoromethylbenzene, anhydrous tetrahydrofuran or 2-methyl-tetrahydrofuran, and stirred from 1 to 72 hours at room temperature, for instance at 25°C. Unreacted acid and base residues may be removed by extractive washes and the desired product [IX] can be crystallized or purified by chromatography on silica gel.

Subsequently, the L-cystine diester diamide obtained by the above-described treatment step is treated in a suitable solvent such as dichloromethane, dioxane, chloroform, trifluoromethylbenzene, anhydrous tetrahydrofuran or 2-methyl-tetrahydrofuran with DL- dithiothreitol (DTT) and an organic base, typically, triethylamine, diisopropylethylamine, pyridine, N- methylmorpholine, and stirred at room temperature (for instance, 25°C) from 1 to 72 hours. After an aqueous workup, the organic layer is concentrated under vacuum to obtain a product that may be purified by crystallization or chromatography on silica gel, or that may be used as such in the subsequent step.

Afterwards, the L-cysteine ester amide is treated with an acyl chloride, typically in a solvent selected among the above-mentioned ones. An organic buffer may be added, comprising a salt of an amine, in particular a salt of a tertiary amine, such as trimethylamine, triethylamine, diisopropylethylamine, pyridine, and also comprising a free base thereof.

Compounds of formula [I], where R' ' ' ' is thienylsulfanyl and/or thiazolylsulfanyl, in particular thienyl-2-sulfanyl and/or thiazolyl-2-sulfanyl, can be s

wherein X may be C or N respectively for thienyl-2- sulfanyl and/or thiazolyl-2-sulfanyl .

S-thienyl or S-thiazolyl L-cysteine [X] is treated with an organic base, typically pyridine, triethylamine, diisopropylethylamine, N-methylmorpholine, and with the chloride of an optionally substituted phenylacetic or benzoic acid in a solvent such as dichloromethane, dioxane, chloroform, trifluoromethylbenzene, anhydrous tetrahydrofuran or 2-methyl-tetrahydrofuran, and is stirred from 1 to 72 hours at room temperature (for instance, 25°C) . Unreacted acid and base may be removed by extractive washes and the desired product [XI] can be crystallized or purified by chromatography on silica gel.

The required phenylacetic or benzoic acids and acyl chlorides, 2-aminoacrylic acid methyl ester, L- cysteine esters, L-cystine diesters, S-thiazolyl-L-cysteine and S- thienyl-L-cysteine are commercially available or are prepared by methods well known by the ones skilled in the art .

EXAMPLES

The invention will be made clearer with the following description of exemplary embodiments thereof, exemplifying but not limitative.

Eight examples are provided, comprising six compounds according to the invention, along with the description of a synthesis route thereto.

- Table 1 -

Citotoxicity against A2780 human ovarian carcinoma cells is also shown. Example 1

0 [xii]

2-aminoacrylic acid methyl ester (450 mg, 4.45 mmol) was dissolved in dichloromethane (20 ml) . Triethylamine (900 mg, 9 mmol), dimethylaminopyridine (catalyst) and 4- biphenyl-carbonyl-chloride (520 mg 5.34 mmol) were added. The reaction was stirred for 18 hours, then water was added. The organic layer was first washed with a saturated sodium bicarbonate solution, then with water. Subsequently, the organic layer was dried over anhydrous sodium sulphate and then evaporated under vacuum. The residue was purified by chromatography on silica gel, eluting with hexanes first, then with ethyl acetate/methanol 95/5, obtaining 225 mg of product [XII] .

HNMR (DMSOd6, 200 Mhz) s: 3.728 (3H); d: 5.795 (2H, j 40.36) ; m: 7.988-7.400 (9H) .

Example 2

S-thiazolyl-L-cysteine (250 mg, 1.22 mmol) was dissolved in THF (20 ml) . Triethylamine (135 mg, 1.342 mmol) and 2 , 2-dimethylphenylacetic acid chloride (222.83 mg, 1.22 mmol) were added. The reaction was stirred at room temperature (for instance, 25°C) for 18 hours, then ethyl acetate was added.

The organic phase was washed with a 1 N solution of hydrochloric acid, then it was dried over anhydrous sodium sulphate and evaporated under vacuum. The residue was purified by chromatography on silica gel, eluting with hexanes first, then with ethyl acetate/methanol 95/5, obtaining 120 mg of product [XIII] .

HNMR (DMSOd6, 200 Mhz) s: 1.392 (6H); m: 3.405- (2H) ; m: 4.425 (1H); m: 7.250-7.700 (7H) . Example 3

S-thienyl-L-cysteine (250 mg, 1.23 mmol) was dissolved in THF (20 ml) . Triethylamine (248 mg., 2.26 mol), dimethylaminopyridine (catalyst) and 4-biphenyl-carbonyl- chloride (269 mg, 1.47 mmol) were added. The reaction was stirred at room temperature (for instance, 25°C) for 18 hours, then ethyl acetate was added.

The organic phase was washed with a 1 N solution of hydrochloric acid, then it was dried over anhydrous sodium sulphate and evaporated under vacuum. The residue was purified by chromatography on silica gel, eluting with hexanes first, then with ethyl acetate/methanol 95/5, obtaining 120 mg of product.

HNMR ( DMSO d6, 200 Mhz) : m: 2.775-3.012 (2H); m: 3.984 (1H) ; m: 7.356-8.033 (12H) .

To a solution of 2-aminoacrilyc acid methyl ester (lg, 9.9 mmol) in DCM (10 ml) were added triethylamine (2 g, 20 mmol.), DMAP (catalyst) and 2 , 4-trifluoromethyl-benzoyl chloride (1.46 g, 11.88 mmol) . The reaction was stirred at room temperature (for instance, 25°C) for 18 hours, then water was added.

The organic layer was washed with a saturated sodium bicarbonate solution, then with water. The organic layer was then it was dried over anhydrous sodium sulphate then evaporated under vacuum. The residue was purified by chromatography on silica gel, eluting with hexanes first, then with ethyl acetate/methanol 95/5, obtaining 560 mg of product [XV] .

HNMR (DMSOd6, 200 MHz) s: 3.751 (3H); d: 6.051 (J: 67.866, 2H) m: 7.820-8.174 (3H) .

4-biphenyl carboxylic acid (1.65 g, 8.3 mmol) was dissolved in THF (20 ml) . Then triethylamine (1.13 g, 11.2 mmol) e 0- (Benzotriazol-l-yl ) -N, N, N ', N ' -tetramethyluronium tetrafluoroborate (3.59 g, 11.2 mmol) were added. The reaction was stirred at room temperature (for instance, 25°C) for 45 min. Then L-cystine dimethyl ester was added (1 g, 3.72 mmol) and the reaction was stirred at room temperature (for instance, 25°C) for 18 hours. The organic solution was treated with a saturated solution of sodium bicarbonate and the organic layer was washed with water, then it was dried over anhydrous sodium sulphate and evaporated under vacuum. The product was purified by chromatography over silica gel eluting with hexanes/ethyl acetate 6/4 obtaining 1.5 g of product [XVI] .

The product of example 5 (1.5 g, 2.38 mmol) was dissolved in THF (20 ml) under inert atmosphere. Then triethylamine (0.48 g, 4.76 mmol) e DL-Dithiothreitol (DTT) (0.73 g 4.76 mmol) were added. The reaction was stirred at room temperature (for instance, 25°C) for 18 hours, then water was added. The organic layer was washed again with water then evaporated under vacuum. The residue was used without further purification. The crude thiol (500 mg, 1.58 mmol) was treated in THF (20 ml) with pivaloyl chloride (228 mg, 1.9 mmol), trimethylamine hydrochloride (151 mg, 1.58 mmol) and triethylamine (240 mg, 2.37 mmol) . The reaction was stirred at room temperature (for instance, 25°C) for 18 hours. Then water was added, the organic layer was washed with brine, dried over anhydrous sodium sulphate and evaporated under vacuum. The residue was purified by chromatography over silica gel, eluting with hexanes/Ethyl acetate 6/4, obtaining 220 mg of product [XVII] .

HNMR (DMSOd6, 200 MHz) s: 1.142 (9H); m: 3.083-3.201 (2H) s: 3.674 (3H) m: 4.220-4.271 (1H) m: 7.366-7.678 (9H); d 7.864-7.909 (1H) .

[XVIII]

4-Pyridin-4-yl-benzoic acid (1.65 g, 8.3 mmol) was dissolved in THF (20 ml) . Then triethylamine (1.13 g, 11.2 mmol) e 0- (Benzotriazol-l-yl ) -N, N, N ' , N ' -tetramethyluronium tetrafluoroborate (3.59 g, 11.2 mmol) were added. The reaction was stirred at room temperature (for instance, 25°C) for 45 min. Then L-cystine dimethyl ester was added (1 g, 3.72 mmol) and the reaction was stirred at room temperature (for instance, 25°C) for 18 hours. The organic solution was treated with a saturated solution of sodium bicarbonate and the organic layer was washed with water, then it was dried over anhydrous sodium sulphate and evaporated under vacuum. The product was purified by chromatography over silica gel eluting with hexanes/ethyl acetate 6/4 obtaining 1.5 g of product [XVIII] . Example 8

The product obtained in example 7 (1.5 g, 2.38 mmol) was dissolved in THF (20 ml) under inert atmosphere. Then triethylamine (480 mg, 4.76 mmol) e DL-Dithiothreitol (DTT) (0.73 g 4.76 mmol) were added. The reaction was stirred at room temperature (for instance, 25°C) for 18 hours, then water was added. The organic layer was washed again with water then evaporated under vacuum. The residue was used without further purification.

500 mg (1.58 mmol) of the crude thiol were treated in THF (20 ml) with pivaloyl chloride (228 mg, 1.9 mmol), trimethylamine hydrochloride (151 mg, 1.58 mmol) and triethylamine (240 mg, 2.37 mmol) . The reaction was stirred at room temperature (for instance, 25°C) for 18 hours. Then water was added, the organic layer was washed with brine, dried over anhydrous sodium sulphate and evaporated under vacuum. The residue was purified by chromatography over silica gel, eluting with hexanes/Ethyl acetate 6/4, obtaining 220 mg of product [XIX] .

HNMR (DMSOd6, 200 MHz) s: 1.145(9H) ; m: 3.086-3.220 (2H), s: 3.651 (3H) m: 4.195-4.259 (1H) m: 7.30-7.81 (8H) ; d: 7.890-7.910 (1H) . A2780 human ovarian carcinoma cells growth inhibition

Ovarian carcinoma A2780 cells (Banca Biologica e Cell Factory 1ST - Istituto Nazionale per la Ricerca sul Cancro, L . go R. Benzi, 10 - 16132 Genova) were grown in the appropriate complete media (RPMI) and seeded at 4 x 103 cells/well in 96-wells Elisa plates. 24 hours later, incubations were performed in the presence of the test compounds at various concentrations, for additional 48 hrs of culture. At the end of the incubation times, cytotoxicity was evaluated by colorimetric water-soluble tetrazolium-1 assay (WST-1, Roche Diagnostics S.p.A, Milan, Italy) following the manufacturer's instructions. WST-1 was added to each well and after an additional 4 h of incubation the coloured tetrazolium salt was spectrophotometrically read at 450 nm in a Victor3 1420 multilabel counter ( Perkin-Elmer , Waltham, MA) .

Representative compounds disclosed in this invention have been tested against A2780 human ovarian carcinoma cells. The results are summarized in Table 2. - Table 2 -

As known, most compounds used for treating neoplasms revealed themselves suitable for treating autoimmune diseases as well. Therefore, the above compound are expected to be useful for treating autoimmune diseases, for example rheumatoid arthritis.

The foregoing description of embodiments of the invention will so fully reveal the invention according to the conceptual point of view, so that others, by applying current knowledge, will be able to modify and/or adapt for various applications such embodiments without further research and without parting from the invention, and it is therefore to be understood that such adaptations and modifications will have to be considered as equivalent to the specific embodiment. The means and the materials to perform the different functions described herein could have a different nature without, for this reason, departing from the field of the invention. It is to be understood that the phraseology or terminology that is employed herein is for the purpose of description and not of limitation.

Claims

1. A compound of formula:

R'

or a pharmaceutically acceptable salt or solvate thereof, wherein:

— R' is selected among H, F, trifluoromethyl ;

— R' ' is selected among OH, O-alkyl;

— R' ' ' is selected among H, F, trifluoromethyl,

phenyl, pyridyl;

— R' ' ' ' is selected among methylene, acylsulfanyl, phenacylsulfanyl , aryloylsulfanyl ,

thien-2-ylsulfanyl , thiazol-2-ylsulfanyl ;

— n is selected between 0 and 1.

2. A compound of formula [I] in which R' ' ' ' is methylene according to the general scheme:

(ID (III) (IV) wherein a 2-aminoacrylic acid ester (III) is coupled with the chloride (II) of an optionally substituted phenylacetic or benzoic acid to obtain the final product ( IV) .

3. A compound of formula [I] in which R' ' ' ' is any of acylsulfanyl or phenacylsulfanyl, or aryloylsulfanyl according to the general scheme:

wherein an L-Cystine diester [V] is treated with an optionally substituted phenylacetic or benzoic acid [VI] and a suitable peptide coupling agent such dicyclohexylcarbodiimide (DCC) , Ν,Ν'- carbonyldiimidazole (CDI), l-(3- dimethylaminopropyl ) -3-ethyl carbodiimide (EDC) , 0- (Benzotriazol-l-yl) -Ν,Ν,Ν',Ν ' -tetramethyluronium tetrafluoroborate (TBTU) .

4. A compound of formula [I] in which R' ' ' ' is any of thienylsulfanyl and/or thiazolylsulfanyl, in particular thienyl-2-sulfanyl and/or thiazolyl-2-

5 . A pharmaceutical composition comprising a compound according to claim 1, and/or a pharmaceutically acceptable salt and/or solvate thereof, for treating a neoplasm.

6. A pharmaceutical composition according to claim 2, wherein said neoplasm is selected from the group comprised of:

— a neoplasm of the genitourinary tract; — a neoplasm of the central nervous system;

— a neoplasm of the endocrine glands;

— a neoplasm of the head and of the neck;

— a neoplasm of the respiratory tract, in particular a neoplasm of the lungs ;

— a neoplasm of the alimentary canal;

— a neoplasm of the female reproductive organs;

— a neoplasm of the hematopoietic system;

— lymphoma;

— myeloma.

7. A pharmaceutical composition according to claim 2, wherein said neoplasm of the hematopoietic system is leukaemia .

8. A pharmaceutical composition comprising a compound according to claim 1, or a pharmaceutically acceptable salt or solvate thereof, for treating an autoimmune disease .

9. A pharmaceutical composition according to claim 5, wherein said autoimmune disease is selected from the group comprised of:

— rheumatoid arthritis.

10. A pharmaceutical composition according to claim 1, wherein said pharmaceutically acceptable salt is an alkaline or an alkaline-earth salt.

11. A pharmaceutical composition according to claim 1, wherein said pharmaceutically acceptable salt is selected among a sodium salt and a potassium salt.

12. A pharmaceutical composition according to claim 1, wherein said pharmaceutically acceptable salt is obtainable by reaction of said compound with an organic amine.

13. A pharmaceutical composition according to claim 1, wherein said pharmaceutically acceptable salt is obtainable by reaction of said compound with an organic amine selected from the group comprised of methylamine ; ethylamine; diethanolamine ; isopropylamine ; diisopropylamine .

14. A pharmaceutical composition according to claim 1, wherein said pharmaceutically acceptable salt is obtainable by reaction of said compound with an acid selected from the group comprised of hydrochloric acid; hydrobromic acid; phosphoric acid; citric acid; fumaric acid; L-tartaric acid; maleic acid; malonic acid; succinic acid.

15. A pharmaceutical composition according to any of claims 2 to 12, comprising a pharmaceutically acceptable carrier, adjuvant or vehicle.