DIAMINOCYCLOHEXANE PLATINUM COMPLEXES, PROCESS FOR PREPARING SAME AND PHARMACEUTICAL COMPOSITIONS CONTAINING SAME

Abstract: Diaminocyclohexane platinum complexes having the structural formula wherein; X is a monovalent anion selected from the group con­ sisting of ascorbate, isoascorbate, monosaccharate, saccha­ rate-4-lactone, shikimate, isethionate, 2-aminoethylsulfate, azetidinecarboxylate, proline, hydroxyproline, pipecolinate, cyclopropanecarboxylate, cyclobutanecarboxylate, cyclopen­ tanecarboxylate, cyclopentenecarboxylate, cyclohexanecar­ boxylate, cyclohexenecarboxylate, bicine, glycine, 2-amino­ ethanesulfonate, 2-chloroethanesulfonate and Y is a divalent anion selected from the group consisting of iminodiacetate, isocitratelactone, furanedicarboxylate, cyclo­ propane-1,1-dicarboxylate, isocitratomonoethylester, N-meth­ yliminodiacetate, N-(2-hydroxyethyl)-iminodiacetate, N-benzyl­ iminodiacetate, N-phenyliminodiacetate, N-(2-acetamido)-im­ inodiacetate, cyclohexane-1,1-diacetate, trans-1,2-cyclopro­ panedicarboxylate, trans-1,2-cyclobutanedicarboxylate, trans-­ 1,2-cyclopentanedicarboxylate and trans-1,2-cyclohexanedi­ carboxylate and pharmaceutical compositions for the treat­ ment of tumors. Disclosed is also a process for preparing the new complexes.

Description: Diaminocyclohexane platinum complexes, process for preparing same and pharmaceutical compositions containing same The present invention relates to certain novel 1,2-diaminocyclohexane platinum complexes, pharmaceutical compositions for use in the treatment of tumors and a process for preparing the complexes. Recently, certain platinum complexes have been shown by Rosenberg et al and others to be highly active anti-tumor agents (see U.S. Patents 4,177,263 and 4,140,707). For example, the complex cis-dichlorodiammineplatinum-II or "cisplatin" is the chemotherapeutic agent of choice in the treatment of many and varied tumors. There are several drawbacks, however, associated with the use of the platinum complexes' to treat tumors. Generally, the platinum complexes have a relatively low solubility in water thereby rendering it difficult to formulate a composition which can effectively deliver the reagent to the site of the tumor in the body. Moreover, many of the platinum complexes are highly nephrotoxic thereby further restricting their use in the absence of precautionary measures to avoid damage to the kidneys when administered to animals afflicted with tumors. Recently, considerable activity has centered on the use of 1,2-diaminocyclohexane complexes of platinum as anti-tumor agents. Sce, for example, U.S. Patents Nos. 3,892,790; 3,904,663; 4,115,418; 4,140,707; 4,169,846; 4,175,133; 4,228,090 and 4,256,652 and German Offenlegungschrift 30 22 917. Although the complexes described therein posscss antitumor activity, virtually all are difficultly soluble in water and are also relatively highl nephrotoxic. German Offenlegungschrift 30 22 957 describes 1,2-diaminocyclohexane/ascorbate complexes of platinum, useful as anti-tumor agents, which comprise mixtures of complexes having varying ratios of ascorbate ion to platinum. Thus, the publication discloses complexes of the formula: EMI2.1 wherein n varies from 1.2 to 1.5. The complexes are described as poorly water soluble. The hereinbelow described method enables the production of complexes of DACH-platinum complexes of the following formulae which contain stoichiometric ratios of anion to platinum: EMI2.2 wherein: X is a monovalent anion selected from the group consisting of ascorbate, isoascorbate, mono saccharate, saccharate-4-lactone, shikimate, isethionate, 2-aminoethylsulfate, azetidine carboxylate, proline, hydroxyproline, 'pipecolinate, cyclopropanecarboxylate, cyclobutanecarboxylate, cyclopentanecar boxylate, cyclopentenecarboxylate, cyclo hexanecarboxylate, cyclohexenecarboxylate, bicine, glycine, 2-aminoethanesulfonate, 2-chloroethanesulfonate, and Y is a divalent anion selected from the group consisting of iminodiacetate, iso citratelactone, furanedicarboxylate, cyclo propane-l, l-dicarboxylate and isocitra tomonoethyl esther, N-methyliminodiacetate, N-(2-hydroxyethyl)-iminodiacetate, N benzyliminodiacetate, N-phenyliminodiacetate, N- ( 2-acetamido)-iminodiacetate, cyclohexane 1, 1-diacetate, trans-i, 2-cyciopropanedicar- boxylate, trans-1,2-cyclobutanedicarboxylate, trans-1,2-cyclopentanedicarboxylate and trans-1,2-cyclohexanedicarboxylate The method of the invention for preparing the above-described complexes comprises: : a) reacting a water-soluble haloplatinatc(IlJ in an aqueous medium with DACH to produce a di-halo-(DACH) )platinum(II); b) reacting said product of step a) with a soluble sulfate salt in an aqueous medium to produce sulfato(DjtCII)-platinum(II); c) reacting the product of step b) with a soluble salt of X or Y to produce said complex of the above formula, and d) recovering said complex. The present invention also provides a pharmaceutical composition in unit dosage form suitable for administration to an animal afflicted with tumor cells sensitive to a platinum complex of the above formula comprising a therapeutically anti-tumor effective amount of the platinum complex and a pharmaceutically acceptable carrier therefor. The invention also provides a method for the treatment of an animal afflicted with tumor cells sensitive to a platinum complex of the above formula comprising administering to the animal a therapeutically, anti-tumor effective amount of the platinum complex. DETAILED DESCRIPTION OF THE INVENTION The complexes of the invention have the following structural formulae: EMI4.1 [bis-ascorbato] EMI5.1 [bis-isoascorbato] EMI5.2 [bis-monosaccharato] EMI5.3 [bis-saccharato 1,4 lactone] EMI6.1 [bis-shikimato] EMI6.2 [bis-isethionato] EMI6.3 [bis-2-aminocthylsulfato] EMI7.1 [bis-azetidinecarboxylato] EMI7.2 [bis-prolinato] EMI7.3 [bis-hydroxyprolinato] EMI8.1 [bis-pipecolinato] EMI8.2 [Iminodiacetato] EMI8.3 [isocitratolactone] EMI9.1 [furanedicarboxylate] EMI9.2 [cyclopropanedicarboxylato-] EMI9.3 [isocitratomonethylester] EMI9.4 (bis-cyclopropanecarboxylato) EMI10.1 (bis-cyclobutanecarboxylato) EMI10.2 (bis-cyclopentanecarboxylato) EMI10.3 (bis-cyclopentenecarboxylato) EMI10.4 (bis-cyclohexanecarboxylato) EMI11.1 (bis-cyclohexenecarboxylato) EMI11.2 (N-phenyliminodiacetato) EMI11.3 (cyclohexane-l,l-diacetato) EMI11.4 (1,2-cyclopropanedicarboxylato) EMI12.1 (trans-1,2-cyclobutanedicarboxylato) EMI12.2 (trans-1,2-cyclopentanedicarboxylato) EMI12.3 (trans-1,2-cyclohexanedicarboxylato) EMI12.4 (N-(2-acetamidoj-iminodiacetato) EMI13.1 (bis-2-chloroethanesulfonato) EMI13.2 (bis-2-aminoethanesulfonato) EMI13.3 (bis-glycinato) EMI13.4 (bis-bicinato) EMI14.1 (N-methyliminodiacetato) EMI14.2 (N-(2-hydroxyethyl)iminodiacetato) EMI14.3 (N-benzyliminodiacetato) The stoichiometric mono- and bis- anion-(DAClI) platinum complexes of the invention (i.e., containing one divalent anion or two monovalent anions per molecule of platinum) may be prepared by reacting any suitable water soluble haloplatinate (i.e., R2PtC14) with DACH to produce the intermediate, di-halo-(DACH)platinum(II). The reaction is preferably effected in water at room temperature for a time sufficient to drive the reaction to completion, generally from about 6 to 8 hours. The intermediate di-halo complex is normally a yellow solid precipitate which is advantageously isolated from the reaction mixture by filtering; followed by washing with water, methanol and acetone. Finally, the yellow solid is dried, preferably under vacuum or is left in solution and immediately reacted therein with a sulfate. It is preferred to utilize a sulfate of a metal or cation the halo salt of which is insoluble in water to facilitate isolation of the product. Thus, where the di-halo (DACH) platinum is the di-chloro complex, it is preferred to employ Ag2SO4 thereby facilitating removal of the insoluble by-product, AgCl. The reaction is preferably carried out in an aqueous medium such as water or in the reaction medium from the first step of the method at room temperature. The reaction generally gocs to completion in from about 18 to about 24 hours. Following removal of the insoluble by-product halide, the intermediate sulfato (DACH-platinum(II)) is isolated by evaporating the yellow filtrate to dryness, e.g., using a rotary evaporator, as a yellow-brown solid which is washed, erg., with acetone and dried, preferably, in vacuum. Alternatively, the sulfato-DACH-platinum intermediate is left in solution and reacted therein with a soluble salt of the appropriate anion to yield the platinum complex of the invention It is preferred to utilize a metal or other cation salt of the anion whose by-product sulfate salt is insoluble in water to facilitate isolation of the platinum complex. For example, the use of a barium salt of the anion yiclds barium sulfate as a by-product which is readily removable from the reaction medium by filtration. The reaction is preferably conducted in an aqueous medium at about room temperature for a time sufficient to drive the reaction to completion, generally from about 0.3 to about 0.5 hours. The platinum complex may be isolated from the reaction medium by filtering the by-product barium sulfate. The yellow filtrate is evaporated to dryness at 45-500C under reduced pressure, e.g., using a rotary evaporator. The yellow brown solid is then dried, e.g., over P205 under vacuum. The reaction sequence for preparing the bis ascorbate complex of the invention is depicted by the following equations wherein: L =. DACH, and Asc = ascorbate K2PtC14 + L LPtCl2 + Ag2SO4 LPtSO4 + Ba(Asc)2 EMI16.1 LPtCl2 + 2KC1 LPtSO4 + 2AgCl LPt(Asc)2 + BaSO4 The method of invention is illustrated by the following non-limiting examples. The products of preparative examples 1 - 13, 22, 23 and 24 are summarized in Table 1. TABLE I COMTOUXD NO. COMPOUND EXEMPLE NO. 1 cis-bis-dichloro(DACH)Pt(II) 1 2 sulfato(DAClUPt(II) 2 3 cis-bis-ascorbato[DACH]Pt(II). 1H2O 3 4 cis-bis-ascorbato[trans-1-DACH]Pt(II).3H2O 4 5 cis-bis-ascorbato[trans-d-DACH]Pt(II).2H2O 5 6 cis-bis-ascorbato(cis-DACH]Pt(II).5H2O 6 7 cis-bis-isoascorbato(DACH)Pt(II).3H2O 7 8 cis-bis-D-monosaccharato(DACH)Pt(II).3H2O 8 9 cis-bis-D-saccharato-1,4-lactone(DACH)Pt(II) 10 cis-bis-shikimato(DACH)Pt(II).2H20 10 11 iminodiacetato(DACH)Pt(II). 2H20 11 12 DL-isocitratolactone(DACH)Pt(II).1H2O 12 13 cis-bis-isethionato(DACH)Pt(II).2H2O 13 14 cis-bis-2-aminoethylsulfato(DACH)Pt(II) 13 15 cis-bis-L-azetidinecarboxylato(DACH)Pt(II).1H2O 13 16 cis-bis-L-prolinato(D.4CHjPt(II) 13 17 cis-bis-L-hyroxyprolinato(DACH)Pt(II).1H2) 13 18 cis-bis-DL-pipecolinato(DACH)Pt(II).1H2) 13 19 furanedicarboxylato(DACH)Pt(II) 13 20 cyclopropane-l,l-dicarboxylato(DACH)Pt(II).H20 13 21 isocitratomonoethylester(DACH)Pt(II).5H2O 13 TABLE I (continued) COMPOUND NO. COMPOUND EXAMPLE NO. 22 Cis-bis-cyclohexanecarboxylato 24 (DACH)Pt(II) .iH2O 23 l,l-cyclohexanediaceta.to(DACH) 24 Pt(II).1H2O 24 Cis-bis-cyclohexenecarboxylato 24 (DACH)Pt(II). 1H2O 25 Cis-bis-cyclopentanecarboxylato 24 (DACH)Pt(II). 1H2O 26 Cis-bis-cyclopentenecarboxylato 24 (DACH)Pt(II). 2H2O 27 trans-DL-1,2-cyclopentanedicar- 24 boxylato(DACH)Pt(II) 28 N-Methyliminodiacetato(DACH)Pt 24 (11) .iH2O 29 Cis-bis-cyclopropanecarboxylato 24 (DACH)Pt(II) lH20 30 Cis-bis-cyclobutanecarboxylato 24 (DACH)Pt(II). 1H2O 31 Cis-bis-ethanesulfonato(DACH) 24 Pt(II) H20 32 Cis-bis-glycinato(DACH)Pt(II). 1H2O 24 33 Cis-bis-chloroethanesulfonato(DACH) 24 Pt(II).lH20. 34 Cis-bis-bicinato(DACH)Pt(II) 24 35 Iminodiacetato(trans-1-DACH)Pt(II).2H2O 23 36 Cis-bis-L-prolinato(trans-1-DACH) 22 Pt(II) 2H2O 37 N-Phenyliminodiacetato(DACH)Pt(II). 2H2O 24 38 N-Benzyliminodiacetato(DACH)Pt(II) 24 39 N-(2-Hydroxyethyl)-iminodiacetato 24 (DACH)Pt(II) 2H20 40 trans-1,2-cyclopropanedicarboxylato 24 (DACH)Pt(II) TABLE I (continued) COMPOUND NO. COMPOUND EXAMPLE NO. 41 trans-1,2-cyclobutanedicarboxy- 24 lato(DACH)Pt(II) 42 N-(2-Acetamido)-iminodiacetato(DACH) 24 Pt(II).1H2O 43 trans-1,2-cyclohexanedicarboxylato 24 (DACH)Pt(II) EXAMPLE 1 Cis-bis-dichloro(DACH)platinum(II) To an aqueous filtered solution of R'2PtcL4(3.5g; 8.4 mmole in 50 ml of water) 0.9416 g (8.4 mmole) of DACII was added. The reaction mixture was stirred at room temperature for 6-8 hours. A yellow solid was precipitated, filtered, washed with water, methanol and finally with acetone. The final product was dried under vacuum. Yield = 56%. Cis-bis-dichloro(trans-1-DACH)platinum (II); cisbis-dichloro(trans-d-DACH)platinum(II) and cis-bisdichloro(cis-DACH)platinum(II) were prepared in an analogous manner using stoichiometric amounts (ca. 1 mmole) of K2Ptcl4 and the respective DACIl isomers, i.e., trans-l, trans-d and cis-. EXAMPLE 2 Sulfato(DACH)platinum(II) Dichloro(DACII)platinum(II) (DACH being a mixture-, trans-l-, trans-d- or cis-isomer)(l.0 g; 2.6 mmole) was suspended in water (20 ml), and a solution of Ag2So4 (0.75g; 2.4 mmole) in water (150 ml) was added. The reaction mixture was stirred at room temperature for 24 hours in the dark. The precipitated Agcl was filtered off, and the yellow filtrate was evaporated to dryness at 45-50 C under reduced pressure using a rotary evaporator. A yellowbrown product was obtained and dried over P205 under vacuum. Yield: 90"a. EXAMPLE 3 Cis-bis-ascorbate(DACIl)platinum(II).1ll2O Sulfato(DACII)platinum(II) 0.8 g (2 mmole) was dissolved in water (50 ml) and barium ascorbate (0.974 g; 2 mmole in 30 ml of water) was added thereto. The reaction mixture was stirred ca. 20 minutes at room temperature. Barium sulfate precipitate was filtered off, and the yellow filtrate was evaporated to dryness at 45-500C under reduced pressure using a rotary evaporator. A brown yellow product was obtained, which was then washed with acetone. The product was finally dried under vacuum. Yield = 90%. The analytical data for thegproduct is set forth in Table II. Platinum analysis - calculated for C18H28N2O12Pt.lH2O: Put28.80, Found: 28.72% EXAMPLE 4 Cis-bis-ascorbato(trans-l-DACH)platinum(II).3El2O Sulfato(trans-l-DACH)platinum(II) 0.405 g (1 mmole) was dissolved in water (20 ml) and a solution of barium ascorbate (0.478g; 1 mmole in 10 ml of water) was added thereto. The reaction mixture was stirred for 20 minutes. Barium sulfate precipitate was filtered off, and the yellow filtrate was evaporated to dryness at 45-500C under reduced pressure using a rotary evaporator. A brown yellow solid was obtained, which was then washed with acetone and dried in vacuo. Yield = 85%. The analytical data for the product is set forth in Table II. EXAMPLE 5 Cis-bis-ascorbato (trans-d-DACH)platinum(II). 21120 Sulfato(trans'DACH)platinum(II) 0.405 g (1 mmole) was dissolved in water (20 ml) and a solution of barium ascorbate (0.487; 1 mmole in 10 ml of water) was added thereto. The reaction mixture was stirred at room temperature for 20 minutes. Barium sulfate precipitate was filtered off, and the brown yellow filtrate was evaporated to dryness 45-50"C under reduced pressure using a rotary evaporator. A yellow solid was obtained which was then washed with acetone and dried under vacuum. Yield = 85%. The analytical data for the product is set forth in Table II. EXAMPLE 6 Cis-bis-ascorbato(cis-DACH)platinum(II) Sulfato(cis-DACEI)platinum(II) 0.2 g (0.5 mmole) was dissolved in water (10 ml) and a solution of barium ascorbate (0.244 g; 0.5 mmole in 10 ml of water) was added thereto. The reaction mixture was stirred at room temperature for 20 minutes. Barium sulfate precipitate was filtered off, and the yellow filtrate was evaporated to dryness at 45-50 C under. reduced pressure using a rotary evaporator. A brown yellow solid was obtained, which was then washed with acetone and dried in vacuo. Yield = 808. The analytical data for the product is set forth in Table II. EXAMPLE 7 Cis-bis-isoascorbato(DACII)platinum(II) .3F120 Sulfato(DACII)platinum(II) 0.405 g (1 mmole) was dissolved in water (20 ml) and a solution of barium isoascorbate (0.487 g; 1 mmole in 10 ml of water) was added thereto. The reaction mixture was stirred at room temperature for 20 minutes. Barium sulfate precipitate was filtered off, and the yellow filtrate was evaporated to dryness at 45-50"C under reduced pressure using a rotary evaporator. A brown yellow product was obtained, which was then washed with acetone. The product was finally dried in vacuo. Yield = 85%. The analytical data for the product is set forth in Table II. Platinum analysis - calculated for C18H28N2012Pt 3H20: Pt 27.34%. Found: Pt 27.56%. EXAMPLE 8 Cis-bis-D-monosaccharato(DACH)platinum(II) .3H20 Sulfato(DACH)platinum(II) 1.0 g (2.47 mmole) was dissolved in water (50 ml) and a solution of barium monosaccharate (1.372 g; 2.47 mmole in 80 ml of water) was added thereto. The reaction mixture was stirred at room temperature for 20 minutes. Barium sulfate precipitate was filtered off, and the yellow filtrate was evaporated to dryness at 45-50"C under reduced pressure using a rotary evaporator. A brown yellow solid was obtained, which was then washed with acetone and dried in vacuo. Yield = 89%. The analytical data for the product is set forth in Table 11. Platinum Analysis - calculated for C18H32N2Q6Pt.3H2O: Pt 24.96. Found: Pt 24.77%. EXANII'IE 9 Cis-bis-D-saccharate-1,4-lactone(DACII)platinum(II) Sulfato(DACIl)platinum(II) 0.101 g was dissolved in 10 ml of water and barium saccharate 1,4 lactone, prepared in situ by the addition of Ba(OH)2.8H2O 0.075 g to an aqueous solution of D-saccharic acid 1,4 lactone monohydrate 0.11 g was added thereto and the reaction mixture was stirred for 30 minutes at room temperature. Barium sulfate precipitate was filtered off and the yellow filtrate was evaporated to dryness at 45-50 C under reduced pressure using a rotary evaporator. A brown yellow solid was obtained, which was then washed with methanol and acetone. The product was finally dried in vacuo. Yield = 82%. The analytical data for the product is set forth in Table II. EXAMPLE 10 Cis-bis-shikimato(DACH)platinum(II) Sulfato(DACH)platinum(II) 0.405 g (1 mmole) was dissolved in water (20 ml) and a solution of barium shikimate (0.483 g; 1 mmole in 50 ml of water) was added thereto. The reaction mixture was stirred at room temperature for 20 minutes. Barium sulfate precipitate was filtered off, and the yellow filtrate was evaporated to dryness at 45-50 C under reduced pressure using a rotary evaporator. A bron yellow solid was obtained, which was then washed with acetone and dried in vacuo. Yield = 80%. The analytical data for the product is set forth in Table II. Platinum Analysis - Calculated for C 20H32N2O10 Pt .2H2), Pt 28.20% Found: Pt 28.76%. EXAMPLE II Iminodiacotato(DACH)platinum (II) Sulfato(DACII)platinum(II) 1.0 g (2.47 mmole) was dissolved in water (SO ml) and a solution of barium iminodiacetate (0.663 g; 2.57 mmole in 50 ml of water) was added thercto. The reaction mixture was stirred at room temperature for 20 minutes. Barium sulfate precipitate was filtered off, and the yellow filtrate was evaporated to dryness at 45-500C under reduced pressure using a rotary evaporator. A brown yellow solid was obtained, which was then washed with acetone and dried in vacuo. Yield = 79%. The analytical data for the product is set forth in Table II. EXAMPLE 12 DL-Isocitratolactone(DACH)platinum (II) Sulfato(DACH)platinum(II) 0.1 g (0.25 mmole) was dissolved in water (10 ml) and a solution of barium isocitratolactone (0.766 g; 0.25 mmole in 20 ml of water) was added thereto. The reaction mixture was stirred at room temperature for 20 minutes. Barium sulfate precipitate was filtered off, and the yellow filtrate was evaporated to dryness at 45-50 C under reduced pressure using a rotary evaporator. A brown yellow solid was obtained, which was then washed with acetone and dried in vacuo. Yield = 75%. The analytical data for the product is set forth in Table II. EXAMPLE 13 The other complexes of the invention, i.e., bisisethionato, bis- 2-aminoethylsulfato, bis-azetidinecarboxylato, bis-prolinato, bis-hydroxyprolinato, bispipecolinato, furanedicarboxylato, cyclopropane-l,ldicarboxylato- and isocitratomonoethyl ester were prepared in an analogous manner to the methods of Example 8, using stoichiometric amounts (ca. 1 mmole) sulfato(DACI1)- platinum(II) and the respective barium salts, i.e., isethionate, 2-aminoethylsulfate, azetidinecarboxylate, proline, hydroxyproline, pipecolinate, furanedicarboxylate, cyclopropane dicarboxylate and isocitratemonoethyl ester. It will be understood by those skilled in the art that the various isomeric DACH (i.e., trans-d-, trans-l-, cis-) can be prepared according to the methods of the above examples employing the appropriate DACH isomer. The analytical data for the compounds prepared in Examples 1-13 are set forth in Table II. TABLE II ANALYTICAL DATA EXAMPLE FOUND (%) CALCULATED (%) NO. C Ij N C H N 3 32.15 4.32 3.85 31.90 4.43 4.13 4 30.01 4.53 3.70 30.28 4.76 3.92 5 31.28 4.38 3.93 31.07 4.60 4.03 6 28.94 4.01 3.73 28.83 5.07 3.74 7 30.25 4.19 3.93 . 30.28 4.76 3.92 8 27.16 4.23 3.23 27.64 4.86 3.58 9 29.66 4.15 3.53 29.70 3.85 3.85 10 34.60 5.11 3.75 34.72 5.20 4.05 11 25.35 4.83 8.57 25.20 4.83 8.82 17 28.65 4.13 5.42 28.85 4.00 5.61 13 19.62 4.16 4.19 20.16 4.70 4.70 14 20.28 4.62 8.99 20.36 4.41 9.50 15 31.98 5.20 10.48 31.86 4.31 10.62 16 35.63 5.77 9.87 35.73 5.58 10.42 17 32.75 5.57 9.39 32.69 5.-11 9.53 18 37.30 6.21 9.46 37.03 6.17 9.70 19 28.84 3.53 5.33 28.80 4.00 5.60 20 29.37 4.44 5.50 28.94 4.60 6.14 21 30.10 4.46 5.32 30.32 4.87 5.05 The in vitro anti-tumor activities of the complexes of the invention are illustrated by the following examples: EXAMPLE 14 Wild type 11210 leukemic cells were grown as a suspension culture in McCoy's 5 medium supplemented with 10% horse serum, glutamine, streptomycin and penicillin at 370C, 95 relative humidity and 5% CO2. Four ml of cell suspension (105 cells/ml) are added to culture tubes and the appropriate concentration (0.01, 0.1, 1 or 10 ug/ml final concentration) of drug added to the culture tubes. After 72 hours of incubation, the cell concentration of control and experimental cultures are determined with the aid of a Coulter CounterR Model ZBf and the percent inhibition calculated. COMPOUND NUMBER ID50 ( g/ml) 3 5.0 4 1.9 5 4.4 6 4.0 7 4.1 8 2,8 9 3.0 10 2.5 11 4.2 12 1.0 13 0.43 14 4,3 15 3.6 15 4,0 17 2.7 18 0.44 19 1.0 20 0.41 21 2.0 The lack of cross-resistance of complexes of the invention to cisdiamminedichloroplatinum(lI) is illustrated below: : EXAMPLE 15 L1210 leukemia cells (L1210/PDD) which are more than 50 old resistant to cisdiamminedichloroplatinum(II) were grown as suspension cultures in McCoy's SA supplemented with 10t horse serum, glutamine, penicillin and streptomycin at 37 C, 95% relative humidity and 5% CO2, Four ml of cell suspcnsion were added to culture tubes and the appropriate concentration (0.01, 0.1, 1 or 10 ug/ml final drug concentration) was added. After 96 hours the cell concentration of control and experimental cultures were calculated with the R aid of a Coulter Counter Model ZBf and the percent inhibition calculated. ID50(g/ml) COMPOUND NUMBER L1210/0 L1210/PDD 3 5.0 2.6 4 1.9 0.4 5 4.4 2.3 8 2.8 0.9 9 3.0 0.6 10 2.5 1.2 11 4.2 1.6 cisdiamminedichloro- latinum(II) 0.1 5.5 The anti-tumor activitics of the complexes of the invention are illustratcd by the following example: EXAMPLE 16 BD mice were inoculated intraperitoneally with 106 L1210 cells. About 24 hours after inoculation of the cells, the mice were injected intraperitoneally with varying dosages of complexes of the present invention. Six mice were used for each dosage level in each experiment with an equal number of control mice inoculated with 106 L1210 cells and left untreated with a given complex. The results [ET/C = (survival time of treated animals/survival time of control animals) x 100] are set forth below. Long term survival signifies that animals were alive 30 days after inoculation with L1210 cells. COMPOUND NUMBER DOSE (mg/kg) %T/C Long Term Survivors 3 50 125 -- 60 163 -- 75 143 90 145 100 Toxic -- 4 25 128 50" 184 1/6 5 25 136 --- 50 Toxic -- 6 50 Toxic 100 Toxic -- 7 25 132 - -- 50 136 -- 100 196* 1/6 * 2/6 animals died of apparent drug toxicity COMPOUND NUMBER DOSE (mg/kg) %T/C Long Term Survivors 8 12.5 120 25 129 50 156 100 151 200 Toxic --9 12.5 127 25 138 50 156 100 Toxic --10 50 140 100 217 2/6 11 50 163 100 109* 12 25 133 50 145 100 Toxic * Apparent Toxicity 13 5 153 10 180 20 140* 1/6 50 Toxic * 2/6 animals died of apprent drug toxicity 14 6.25 12.5 In 25 Progress 50 15 12.5 157 25 148 50 161 100 Toxic 16 12.5 144 25 165 50 165 100 162* * 1 animal died from apparent toxicity COMPOUND NUMBER DOSE (mg/kg) %T/C Long Term Survivors 17 25 160 -- 50 170 100 Toxic 18 12.5 145 25 180 50 Toxic -- 19 6.25 143 12.5 136 25 136 50 Toxic -- EXAMPLE 17 The procedure of Example 16 was repeated except that the treated animals were injected with multiple doses of the complex as indicated below: COMTOUND IBER DOSE(mg/kg) Day of %T/C Long Term ------------- ----------- Administration ---------- Survivors 3 25 1,5,9 176 10 1,5,9 123 20 1,5,9 196 1/6 30 1,5,9 257 1/6 30 1,5,9,13 273 1/6 2.5 1-9 113 5 1-9 142 10 1-9 184 -- 20 1-9 239 1/6 4 25 1,5,9 218 50 1,5,9 144 7 12.5 1,5,9 151 25 1,5,9 179 50 1,5; ;9 212 8 25 1,5,9 206 50 1,5,9 225 1/6 10 12.5 1,5,9 139 25 1,5,9 149 50 1,5,9 222 1/6 11 25 1,5,9 199 50 1,5,9 220 2/6 12 25 1,5,9 219 50 1,5,9 165 - - EXAMPLE 18 The anti-tumor activity of compound number 3 was evaluated against cells sensitive to (L1210/0) and resistant to (L1210/PDD) the standard platinum complex cis-diamminedichloroplatinum(II). L1210/PDD DOSE (mg/kg) SCHEDULE %T/C $SURVIVORS 6.25 1,5,9 115 12.5 1,5,9 124 - --- 25 1,5,9 135 50 1,5,9 223 3/10 100 1,5,9 172* * Toxicity apparent as excessive loss of weight L1210/0 6.25 1,5,9 117 12.5 1,5,9 135 25 1,5,9 181 2/10 50 1,5,9 247 1/10 100 1,5,9 104 The acute toxicity data obtained with selected complexes of the invention are set forth below: EXAMPLE 19 Male albino mice were administered a single intraperitoncal injection of the appropriate test compound and were observed daily for signs of toxicity and survival. Fourteen days after treatment all surviving mice were sacrificed and the LD LD50 and LD90 were calculated. COMPOUND NUMBER DOSE (mg/kg) DEATHS/TREATED 3 110 1/6 125 2/6 140 4/6 155 6/6 175 6/6 Calculated LD10 108 mg/kg Calculated LD50 125 mg/kg Calculated LD90 150 mg/kg 5 60 0/6 80 2/6 100 5/6 125 6/6 160 5/6 Calculated LD 70 mg/kg Calculated LD50 90 mg/kg Calculated LD90 125 mg/kg 8 100 1/6 125 1/6 158 4/6 200 3/6 250 6/6 Calculated LD10 115 mg/kg Calculated LD50 155 mg/kg Calculated LD90 220 mg/kg COMPOUND NUMBER DOSE (mg/kg) DEATllS/TREATED 9 85 0/6 100 0/6 125 3/6 158 5/6 200 4/6 Calculated LD10 100 mg/kg Calculated LD50 140 mg/kg Calculated LD90 185 mg/kg 10 50 0/6 100 1/6 125 3/6 160 5/6 200 3/6 Calculated LD10 85 mg/kg Calculated LD50 130 mg/kg Calculated LD90 205 mg/kg 11 85 1/6 100 3/6 125 5/6 158 4/6 200 6/6 Calculated <RTI ID=36.3> LD10 80 mg/kg Calculated LD50 115 mg/kg Calculated LD90 155 mg/kg The potential nephrotoxicity of selected complexes of the invention are set forth below: EXAMPLE 20 Male albino mice were administered a single intraperitoncal injection of cither the calculated LD10 or LD50 of the test compound. Blood was then obtained by retroorbital puncture 96 hours after treatment for determination of blood urea nitroben (BUN) levels ( a standard method of scrcening compounds for potential renal toxicity). RELATIVE BUN COMPOUND NUMBER DOSE (mg/100 ml) 3 LD10 25+6 LD50 23+6 5 LD10 31+3 LD50 38+9 8 LD10 31#8 LD50 35+12 9 LD10 38#8 LD50 50+15 10 LD10 40+7 LD50 39+5 11 LD10 36+12 LD50 38#10 0.09% NaCl 32#2 cis diamminedichloro LD10 53+19 platinum(II) LD50 67+10 EXAMPLE 21 Cis-bis-2-aminoethanesulfonato(DACH)platinum(II) Sulfato (DACH)platinum(II), 0.202 g, was dissolved in 10 ml of water and barium-2-amino-ethanesulfonate, prepared in situ by the addition of Ba (OH)2. 8H2O, 0.15 g, to an aqueous solution of 2-aminoethanesulfonic acid, 0.125 g, was added thereto and the reaction mixture was stirred for 30 minutes at room temperature. Barium sulfate precipitate was filtered off and the yellow filtrate was evaporated to dryness at 45-500C under reduced pressure using a rotary evaporator. A brown yellow solid was obtained which was then washed with methanol and acetone. The product was obtained which was then washed with methanol and acetone. The product was finally dried in vacuo. Yield = 80% The analytical data for the product are set forth in Table III. EXAMPLE 22 Cis-bis-L-prolinato(trans-1-DACH)platinum(II) Sulfato (trans-l-DACH).platinum(II) (0.202 g) was dissolved in 20 ml of water and the barium salt of L-proline, prepared in situ by the addition of Ba (OH)2 8H2O (0.150 g) to an aqueous solution of L-proline (0.115 g) was added thereto and the reaction mixture was stirred for 20-30 minutes at room temperature. Barium sulfate precipitate was filtered off and the yellow filtrate was evaporated to dryness at 45-500C under reduced pressure using a rotary evaporator. A yellowish-white solid was obtained which was then washed with methanol and acetone. The product was finally dried in vacuo. Yield = 75% The analytical data for the product are set forth in Table III. EXAMPLE 23 Iminodiacetato(trans-1-DACH)platinum(II)2H2O Sulfato(trans-l-DACH)platinum(II), 0.405 g, was dissolved in 50 ml of water and barium iminodiacetate prepared in situ by the addition of Ba(OH)2 8H2O, 0.3 g, to an aqueous solution of iminodiacetic acid, 0.133 g in 100 ml water was added thereto and the reaction mixture was stirred for 20-30 minutes at room temperature. Barium sulfate precipitate was filtered off and the yellow filtrate was evaporated to dryness at 45-500C under reduced pressure using a rotary evaporator. A yellowish-white solid was obtained which was then washed with acetone. The product was then dried in vacuo. Yield = 70%. The analytical data for the product are set forth in Table III. EXAMPLE 24 Cis-bis-cyclopropanecarboxylato(DACH)platinum(II) Sulfato(DACH)platinum(II), 0.405 g (1 mmole), was dissolved in water (20 ml) and barium cyclopropanecarboxylate, prepared in situ bythe addition of Ba (OH) 2'8H20 (0.30 g) to an aqueous solution of cyclopropanecarboxylic acid (0.172 g) (2 mmole), was added thereto and the reaction mixture was stirred for 20-30 minutes at room temperature. Barium sulfate precipitate was filtered off and the yellow filtrate was evaporated to dryness at 45-500C under reduced pressure using a rotary evaporator. A brown yellow solid was obtained, which was purified from methanol. The product was finally dried in vacuo. Yield = 70t. The analytical data for the products are set forth in Table III. Other complexes of the invention, i.e. bis-cyclobutanecarboxylate, bis-cyclopentanecarboxylato, bis-cyclopentenecarboxylato, bis-cyclohexanecarboxylato, bis-cyclohexenecarboxylato, cyclopropane-l,l-dicarboxylato, trans DL-1,2-cyclopentanecarboxylato, cyclohexane-l,l-diacetato, bicinato, chloroethanesulfonato, N- (2-hydroxyethyl)-imino-. diacetato, N-(2-acetamido)-iminodiacetato, 3,4-furanedicarboxylato, trans-1,2-cyclopropane dicarboxylato, trans1,2-cyclobutanedicarboxylato, trans-1,2-cyclopentane dicarboxylato and trans-1,2-cyclohexanedicarboxylato were prepared in an analogous manner to the above mentioned method using stoichiometric amounts (ca/lmmole) sulfato (DACH) platinum (II), and the respective barium salts (prepared in situ) i.e. cyclobutanecarboxylate, cyclopentanecarboxylate, cyclopentenecarboxylate, cyclohexanecarboxylate, cyclohexenecarboxylate, cyclopropane-l,l-dicarboxylate, trans-DL-l, 2-cyclopentanedicarboxylate, cyclohexane-l,l- diacetate, bicinate, chloroethanesulfonate, N-(2-hydroxyethyl) iminodiacetate, N- (2-acetamido) -iminodiacetate, 3, 4-furanediacarboxylate, trans-1,2-cyclopropanedicarboxy- late, trans-1,2-cycidbutanedicarboxylate, trans-1,2-cyclopentanedicarboxylate and trans-l, 2-cyclohexanedicarboxylate. TABLE III Analytical Data Found (%) Calculated (%) COMPOUND NO. C H N C H N 22 41.23 6.93 5.4Ï 41.30 6.54 4.82 23 36.58 5.75 5.03 36.55 5.71 5.33 24 41.20 5.47 4.64 41.56 5.89 4.85 25 38.82 5.29 5.08 39.06 6.14 5.06 26 37.77 5.52 4.97 37.95 5.62 4.92 27 33.43 4.58 5 5.5 33.D0 4.72 6.01 28 27.95 5.16 8.58 27,95 4.87 8.89 29 33.40 5.31 5.48 33.79 5.23 5.63 30 36.61 5.72 5.21 36.55 5.71 5.33 31 20.77 5.05 9.51 20.36 4.86 9.73 32 25.31 5.32 11.64 25.25 5.05 11.70 33 18.26 3.91 3.78 18.46 3.69 4.30 34 33.93 5.43 8.33 34.00 5.98 35 25.14 4.37 8.59 25.20 4.83 8.82 36 33.41 5.76 9.47 33.50 5.93 9.77 37 34.65 4.94 7.99 34.77 4.89 7.60 38 38.50 5.20 7.31 38.47 4.71 7.92 39 27.96 5.15 7.62 27.69 5.19 8.07 40 27.96 4.43 5.57 27.90 4.65 5.92 41 31.64 4.83 5.85 31.93 4.43 6.21 42 28.34 5.22 10.47 28.45 4.54 11.00 43 33.86 5.62 5.40 33.12 5.12 5.52 In Vitro Cytotoxicity The following compounds were tested for in vitro cytotoxicity according to the protocol discussed in Example 14. Compound No. 1050 ( g/ml) 22 0.5 23. 0.6 24 1.0 25 0.3 26 1.3 27 0.4 28 1.6 29 0.3 30 0.4 31 3.1 32 2.5 33 0.3 34 1.2 35 3.0 36 0.8 37 1.0 38 2.3 39 3.0 40 0.4 41 0.5 42 0.7 In Vivo Efficacy Studies - Single Dose The following compounds were tested for in vivo oncolytic activity following a single i.p. injection of the drug according to the protocol described in Example 16. Compound No. Dose (n..'r':' T/G LTS 25 50 153 -- 25 178 12.5 161 27 50 183 25 159 12.5 158 28 50 Toxic 25 185 -- 12.5 161 29 50 199 1/6 25 176 12.5 252 1/6 30 50 145 25 154 12.5 149 26 50 157 25 158 12.5 185 31 50 153 25 134 12.5 109 Corpeand No. Dose (mg/kg) %T/C LIS 32 50 150 -- 25 131 -- 12.5 128 33 50 Toxic 25 144 12.5 128 34 50 160 25 151 12.5 165 36 50 149 -- 25 212 1/6 12.5 179 37 50 108 25 155 12.5 172 38- 50 158 25 170 12.5 123 22 100 170 50 140 24 100 105 50 165 25 132 In Vivo Efficacy Studies - Multiple Dose The following complexes were tested for in vivo oncolytic activity following 3 i.p. injections (days 1, 5, 9) according to the protocol described in Examples 16 and 17. Compound No. Dose (mg/kg) T/C LTS 25 25 253 -- 12.5 202 26 25 300 1/6 12.5 356 1/6 6.25 189 -- 31 100 295 2/6 50 193 1/6 37 25 242 1/6 12.5 298 5/6 6.25 246 3/6 38 25 240 2/6 12.5 227 1/6 6.25 277 T/C calculated for 30 day observation period. Long term survivor data represents survivors/treated 50 days after treatment. Tlic platinum complexes of the invention nay be compounded with suitable pharmaceutically acceptable carriers and administered orally, intramuscularly, topically, ctc. It is preferred, however, to combine the complex with suitable media, e.g., 5 dextrose, klucel, water, etc. , for intravenous administration. Care should be takcn, however, to avoid the use of saline as an i.v. medium. Those skilled in the art will be aware of suitable carriers or the complexes of the invention suitable for formulation into capsules, tablets, powders, ointments, pcllets, etc The complexes may also be administered in combination with other anti-tumor agents in a combined chemotherapeutic regimen. The amount of complex included in the pharmaceutical composition and the dosage of complex utilized in the method of treatment of the invention will vary depending in each case upon the condition of the patient, the nature of the tumor undergoing treatment, the anti-tumor activity of the complex, the toxicity and solubility characteristics thereof, etc. Generally-, however, an amount of platinum complex ranging from about 25 to about 200 mg/kg is adequate for most applications.

Claims: Claims 1. A water-soluble square-planar cis-platinum(II) four-coordinate complex having the formula: EMI48.1 wherein: X is a monovalent anion selected from the group consisting of ascorbate, isoascorbate, monosaccharate, saccharate-4-lactone, shikimate, isethionate, 2-aminoethylsulfate, azetidinecarboxylate, proline, hydroxyproline, pipecolinate, cyclopropanecarboxylate, cyclobutanecarboxylate, cyclopentanecarboxylate, cyclopentenecarboxylate, cyclohexanecarboxylate, cyclohexenecarboxylate, bicine, glycine, 2-amino- ethanesulfonate, 2-chloroethanesulfonate and Y is a divalent anion selected from the group consisting of iminodiacetate, isocitratelactone, furanedicarboxylate, cyclopropane-l,l-dicarboxylate, isocitratomonoethylester, N-methyliminodiacetate, N- (2-hydroxyethyl) -iminodiacetate, N-benzylimino diacetate, N-phenyliminodiacetate, N-(2-acetamido) iminodiacetate, cyclohexane-1,1-diacetate, trans 1, 2-cyclopropanedicarboxylate, trans-1,2-cyclobu tanedicarboxylate, trans-1,2-cyclopentanedicarboxy late and trans-1,2-cyclohexanedicarboxylate. 2. A platinum complex of claim 1 which is selected from: EMI49.1 [bis-ascorbato] EMI49.2 [bis-isoascorbato] EMI49.3 [bis-monosaccharato] EMI50.1 [bis-saccharato 1,4 lactone] EMI50.2 [bis-shikimato] 3. A platinum complex of claim 1 which is selected from: EMI51.1 [bis-isethionato] EMI51.2 [bis-2-aminoethylsulfato] EMI51.3 [bis-azetidinecarboxylato] EMI52.1 [bis-prolinato] E. EMI52.2 [bis-hydroxyprolinato] EMI53.1 [bis-pipecolinato] G. EMI53.2 [isocitratolactone] 4. A platinum complex of claim 1 which is selected from: EMI54.1 [Iminodiacetato] B. EMI54.2 (N-phenyliminodiacetato) C. EMI54.3 (N-(2-acetamido)-iminodiacetato) EMI55.1 (N-methyliminodiacetato) E. EMI55.2 (N-(2-hydroxyethyl)iminodiacetato) F. EMI55.3 (N-benzyliminodiacetato) 5. A. platinum complex of claim 1 which is selected from: EMI56.1 [furanedicarboxylate] B. EMI56.2 [cyclopropanedicarboxylato-] EMI56.3 [isocitratomonoethylester] EMI57.1 (bis-cyclopropanecarboxylato) E. EMI57.2 (bis-cyclobatanecarboxylato) F. EMI57.3 (bis-cyclopentanecarboxylato) 6. A platinum complex of claim 1 which is selected from: EMI58.1 (bis-cyclopentenecarboxylato) B. EMI58.2 (bis-cyclohexanecarboxylato) C. EMI58.3 (bis-cyclohexenecarboxylato) EMI59.1 (cyclohexane-1,1-diacetato) E. EMI59.2 (1,2-cyclopropanedicarboxylato) F. EMI59.3 (trans-1,2-cyclobutanedicarboxylato) EMI60.1 (trans-1,2-cyclopentanedicarboxylato) H. EMI60.2 (trans-1,2-cyclohexanedicarboxylato) 7. A platinum complex of claim 1 which is selected from: EMI61.1 (bis- 2-chloroethanesulfcnato) B. EMI61.2 (bis-2-aminoethanesulfonato) C. EMI61.3 (bis-glycinato) EMI62.1 (bis-bicinato) 8. A pharmaceutical composition in unit dosage form suitable for administration to an animal afflicted with tumor cells sensitive to a platinum complex of any of the preceding claims comprising a therapeutically anti-tumor effective amount of said platinum complex and a pharmaceutically acceptable carrier therefor. 9 A method four the preparation of a complex of claim 1 comprising: a) reacting a water-soluble haloplatinate(II) in an aqueous medium with DACH to produce a di-halo-(DACH)- platinum (11); b) reacting said product of step a) with a soluble sulfate salt in an aqueous medium to produce sulfato(DACH)- platinum(II); c) reacting the product of step b) with a soluble salt of X or Y to produce said complex of claim 1, and d) recovering said complex.

Inventors: BROWN, DAVIS B., HACKER, MILES P., KHOKHAR, ABDUL R., MCCOMMACK, JOHN J.
Applicants: RESEARCH CORPORATION
Application year: 1984
Main classification: C07C87
IPC codes: C07D205, C07C59, A61K31, C07H23, C07C211, C07C61, C07C51, C07C55, A61P35, C07F15