KR100307724B1 - Saccharide - Platinum (Ⅱ) conjugate anticanser agent and preparing for process therefor - Google Patents

Abstract

The present invention is obtained by reacting the alkaline earth metal salt of the sugar of formula (VI) with the platinum (II) derivative of formula (VII) in a 1: 1 molar ratio and then removing the by-product barium sulfate. A preparation of the saccharide-platinum (II) complex complex.

The saccharide-platinum (II) complex complex represented by the following general formula (I).

In Formula (I), R is a glucosamine (C ₆ H ₁₂ NO ₅ ), a mannose amine (C ₆ H ₁₂ NO ₅ ), a β-galactose derivative, N-aminobutyl-O-β- as a sugar and a sugar derivative. D-galactopyranosyl- (1 → 4) -D-gluconamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), an α-glucose derivative, N-aminobutyl-O-α-D-glucopyranosyl- ( 1 → 4) -D-gluconamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), β-glucose derivative N-aminobutyl-O-β-D-glucopyranosyl- (1 → 4) -D-glu Is selected from conamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), n is 1 or 2.

Description

Saccharide-Platinum (II) conjugate anticanser agent and preparing for process therefor}

The present invention relates to a saccharide-platinum (II) complex complex anticancer agent having an excellent anticancer effect represented by the following general formula and a preparation method thereof.

In Formula (I), R is a glucosamine (C ₆ H ₁₂ NO ₅ ), a mannose amine (C ₆ H ₁₂ NO ₅ ), a β-galactose derivative, N-aminobutyl-O-β- as a sugar and a sugar derivative. D-galactopyranosyl- (1 → 4) -D-gluconamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), an α-glucose derivative, N-aminobutyl-O-α-D-glucopyranosyl- ( 1 → 4) -D-gluconamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), β-glucose derivative N-aminobutyl-O-β-D-glucopyranosyl- (1 → 4) -D-glu Is selected from conamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), n is 1 or 2.

Among the currently used anticancer drugs, cisplatin, cislatin (cislatin), cis- (NH ₃ ) PtCl ₂ , is known to have anticancer effects by B. Rosenburg, Nature, 205, 698 (1965) in 1965. After being approved as an anticancer agent by the US FDA in 1979, it is used as the most effective chemotherapeutic agent for a wide range of cancers such as testicular cancer, ovarian cancer, bladder cancer, bone marrow cancer, and laryngeal cancer, but side effects including kidney toxicity are pointed out as problems. Since then, carboplatin, which has significantly alleviated side effects, is cis- (NH ₃ ) Pt (CBDC) (CBDC = 1,1-cyclobutanedicarboxylate, MJ Cleare, Biochimie, 60, 835 (1978)). Although it was used as a second-generation anticancer drug after receiving a new drug license, the anticancer effect is significantly lower than that of cisplatin. Therefore, studies are being actively conducted to develop third generation anticancer drugs with excellent anticancer effects and fewer side effects. In addition to the excellent anticancer effect of cisplatin and low toxicity equivalent to carboplatin, the conditions for the third generation platinum complex anticancer agent should be excellent against cisplatin or carboplatin-resistant cancer cells, have proper solubility in water, and chemical stability. There are more than 10 compounds currently in clinical trials in the world because they have to satisfy various demanding conditions such as required, but they are not successful in commercialization.

Glycosaccharides have played an important role as energy sources, structural substances, or components of glycolipids and glycoproteins in living organisms. Recently, oligosaccharides or polysaccharides of glycoproteins and glycolipids play an important role in transferring information between cells and cancerous cells. As it became clear that it was in charge of the study, the study of sugar began to attract attention. In particular, the types of sugars that specifically bind to specific cells are known. For example, β-galactose and galactosamine are known as mammalian hepatocytes, L-fucose and Mannose (mannose) is a mammalian macrophage, N-acetylglucosamine (N-acetylglucosamine) is a hepatocyte of algae, and mannose-6-phosphate (mannose-6-phosphate) specifically binds to human fibroblast Known (Neufeld et al, The biochemistry of glycoprotein and proteoglycans, Plenum, new York, pp. 241 (1980)). Because of the specific binding of specific sugars to specific cells, studies are being actively conducted to develop targeted drugs using sugars (Juliano et al, Targeted drug delivery, Springer-Verlag, Berlin, pp. 137 ( 1991). In addition to the target orientation of these sugars, since many anticancer drugs (adriamycin, bleomycin) contain sugars, sugar is expected to play an important role in the field of anticancer drugs. Studies on the saccharide-platinum (II) complex complex have been reported in several papers. These studies have shown that the solubility of water in platinum complexes is mainly due to the use of amino sugars as amine ligands of the platinum complexes rather than the target orientation of the sugars. Height was used for the purpose (Serkas et al, Inorganica Chimica Acta, 124, 137 (1986), Tsubomura et al, Inorg. Chem. 29, 1622 (1990) .But still saccharides bind to the anion of the platinum (II) complex. There are very few cases of chemotherapy.

It is an object of the present invention to provide a sugar-platinum (II) complex complex having excellent anticancer activity and a method for producing the same. More specifically, an object of the present invention is to bind the platinum (II) complex to the carbohydrate, thereby allowing the platinum (II) complex to act on cancer cells easily through the cell affinity of the carbohydrate, thereby providing excellent anticancer activity and good solubility in water. To provide a platinum (II) complex complex.

In order to achieve the above object, the present inventors use a glutamic acid or aspartic acid, which is a dicarboxylic acid anion that can be chelated to +2 platinum (II) ions as a spacer for introducing platinum (II) complexes to saccharides and sugar derivatives. Was first introduced into the sugar and then reacted with the platinum (II) complex to synthesize a sugar-platinum (II) complex complex that is chemically stable, has good solubility in water, and has excellent anticancer activity.

In order to achieve the above object, the present invention will be described in more detail with respect to the preparation method of the saccharide-platinum (II) complex complex represented by the general formula (I) as follows. That is, disaccharides were prepared by first preparing disaccharides by reacting disaccharide with diamine alkanes using bisaccharides (Kobayashi et al, Polymer J. 17, 567 (1985)). For example, lactose is oxidized and then condensed to form lactonolactone of formula (II), followed by excess 1,4-diaminobutane at 60 ° C. with dimethyl sulfoxide (DMSO) as a solvent. React for 2 hours. The reaction solution is concentrated by evaporation under vacuum and then precipitated in excess of chloroform. The precipitate is filtered and dried to afford N-aminobutyl-O-β-D-galactopyranosyl- (1 → 4) -D-gluconamide of general formula (III).

On the other hand, glutamate or aspartate, which is a dicarboxylic acid amino acid, was used as a spacer for introducing the platinum (II) complex. In order to bind these to the sugars, a carboxylic acid group was introduced into the amine group using the method of the literature (S. Y. Jeong, J. Controlled Release 1, 57 (1984)) as follows. For example, an amino acid derivative of the general formula (IV) is obtained by reacting an equivalent succinic anhydride such as benzyl ester of dicarboxylic amino acid with tetrahydrofuran at room temperature for 8 hours with a solvent.

The synthesized amino acid derivative was dissolved in dimethylformamido (DMF), the solution was kept at 5 ° C, and the same equivalent isobutylchloroformate (IBCF) and tributylamine were added. After the reaction solution was reacted at 5 ° C. for 15 minutes, 0.8 equivalent of saccharide derivative was added thereto, and the reaction solution was reacted at 5 ° C. for 45 minutes at room temperature for 2 hours. The reaction solution was concentrated in vacuo and precipitated in excess ether. The precipitate is filtered and then dried to obtain a saccharide derivative of formula (V). Benzyl groups are removed by reacting this derivative at room temperature with a catalyst of palladium carbon in a methanol solvent containing 4.4% formic acid for 4 hours. The reaction solution is concentrated, precipitated in excess ether, purified, and the product is dissolved in methanol again. A solution of the same amount of barium hydroxide dissolved in methanol is added dropwise to obtain a precipitate. This precipitate is filtered and dried to obtain a saccharide derivative of formula (VI). Meanwhile, dicyclohexaneamine platinum sulfate of formula (VII) is prepared by the method of literature (Harrison et al, Inorg Chim Acta, 46, 15 (1980)). After dissolving the saccharide derivative of the general formula (VI) in distilled water, the same equivalent solution of dicyclohexaneamine sulfate is added and reacted at room temperature for 4 hours. The precipitate formed after the reaction (BaSO ₄ ) is filtered off and the filtrate is lyophilized to obtain a light yellowish saccharide-platinum (II) complex complex of general formula (I). In the scheme, R and n are as defined above.

The present invention will be described in more detail with reference to the following Examples, but the scope of the present invention is not limited to these Examples unless the scope of the claims is claimed. Elemental Analysis of Carbon, Hydrogen, Nitrogen and Platinum for Compounds of the Invention Was performed by a Perkin-Elmer C, H, N analyzer and Polyscan 61E ICP. Meanwhile, hydrogen and platinum nuclear magnetic resonance spectra were measured using a Varian Gemini-300. Example 1 Preparation of β-galactose-platinum (II) Complex Complex C ₆ H ₁₀ (NH ₂ ) ₂ Pt [(OOC) ₂ C ₃₃ H ₄₃ N ₃ O ₁₃ ] (R = C ₁₆ H ₃₃ N ₂ O ₁₁ , n = 2) 20 g of lactonolactone was dissolved in 100 ml of dimethylsulfoxide (DMSO), and then reacted with an excess (70 g) of 1,4-diaminobutane for 2 hours at 60 ° C. for N-aminobutyl- O-β-D-galactopyranosyl- (1 → 4) -D-gluconamide was prepared. 2.5 g of 2-carboxyamidobenzylglutamate (5.99 mmol) was dissolved in 20 ml of DMF, the solution was kept at 5 ° C and 0.62 ml of isobutylchloroformate (IBCF) and 1.5 ml of tributylamine were added. After the reaction solution was reacted at 5 ° C. for 15 minutes, 1.75 g of N-aminobutyl-O-β-D-galactopyranosyl- (1 → 4) -D-gluconamide (4.08 mmol) was added thereto at 5 ° C. After reacting for 45 minutes at room temperature for 2 hours, the reaction solution was concentrated under vacuum and then poured into 500 ml of diethyl ether to precipitate. The precipitate was filtered off and dried to obtain 2.95 g of galactosyl derivative of the general formula (V). 1.2 g of this derivative was dissolved in 50 ml of methanol containing 4.4% formic acid, 0.7 g of palladium carbon was added, and the mixture was reacted at room temperature for 4 hours to remove the benzyl group. The reaction solution was concentrated, precipitated with excess ether, filtered, and dried to obtain 0.7 g of product. The product was dissolved in methanol again, and a solution of 0.42 g of barium hydroxide dissolved in methanol was added dropwise to obtain a precipitate. This precipitate was filtered and dried to obtain 0.82 g of galactosyl derivative of the general formula (VI). 0.2 g (0.24 mmol) of the galactosyl derivative of Formula (VI) was dissolved in distilled water, and an aqueous solution of 0.1 g (0.24 mmol) of dicyclohexaneamine sulfate was added thereto, followed by reaction at room temperature for 4 hours. The precipitate formed after the reaction (BaSO ₄ ) was filtered off and the filtrate was lyophilized to form a β-galactose-platinum (II) complex, C ₆ H ₁₀ (NH ₂ ) ₂ Pt [(OOC) ₂ C ₃₃ H ₄₃ N ₃ O ₁₃ ] 0.21 g (total yield 71.7%). Formula: C ₄₀ H ₅₅ N ₅ O ₁₅ Pt · 3H ₂ O Elemental Analysis: C, 34.35; H, 5.95; N, 6. 15; Pt, 18.56 Theory: C, 36.54; H, 6.03; N, 6.87; Pt, 19.15 hydrogen nuclear magnetic resonance spectrum (D ₂ O, ppm): δ 1.1-1.3 (b, 4H, diaminocyclohexane C-4, C-5 protons), δ 1.5-1.7 (m, 6H, diaminocyclohexane C-3, C-6 protons, -NHCH ₂ CH ₂ CH ₂ CH ₂ NH-), δ 1.8-2.4 (b, 8H, diaminocyclohexane C-1, C-2, C-3, C-6 protons, L-glutamate C- 2, C-3 protons), δ 2.4-2.7 (m, 4H, -C (O) CH ₂ CH ₂ CO-), δ 3.1-3.4 (m, 4H, -NHC H ₂ CH ₂ CH ₂ CH ₂ NH δ 3.4-4.3 (b, 13H, O-β-D-galactopyranosyl- (1 → 4) -D-gluconamide protons, L-glutamate C-1 proton), δ 4.6 (m, 1H, O-β- D-galactopyranosyl C-1 proton Example 2 Preparation of α-glucose-platinum (II) complex complex [C ₆ H ₁₀ (NH ₂ ) ₂ Pt (OOC) ₂ C ₃₃ H ₄₃ N ₃ O ₁₃ ] (R = C ₁₆ H ₃₃ N ₂ O ₁₁ , n = 2) 20 g of maltonolactone instead of lactonolactone, and N-aminobutyl-O-β-D-galactopyranosyl- (1 → 4) -D-gluconamide Instead, the final α-glucose-platinum (II) complex complex in the same manner as in Example 1 using 0.43 g of N-aminobutyl-O-α-D-glucopyranosyl- (1 → 4) -D-gluconamide 0.68 g (total number) . To give a 67.4%) Composition _{formula: C 40 H 55 N 5 O} 15 Pt · 4H 2 O Elemental analysis: C, 35.21; H, 5. 84; N, 6. 28; Pt, 18.20 Theory: C, 35.90; H, 6. 12; N, 6.75; Pt, 18.81 hydrogen nuclear magnetic resonance spectra (D ₂ O, ppm): δ 1.1-1.3 (b, 4H, diaminocyclohexane C-4, C-5 protons), δ 1.5-1.7 (m, 6H, diaminocyclohexane C-3, C-6 protons, -NHCH ₂ CH ₂ CH ₂ CH ₂ NH-), δ 1.8-2.4 (b, 8H, diaminocyclohexane C-1, C-2, C-3, C-6 protons, L-glutamate C- 2, C-3 protons), δ 2.4-2.7 (m, 4H, -C (O) CH ₂ CH ₂ CO-), δ 3.1-3.4 (m, 4H, -NHC H ₂ CH ₂ CH ₂ CH ₂ NH δ 3.4-4.3 (b, 13H, O-α-D-glucopyranosyl- (1 → 4) -D-gluconamideprotons, L-glutamate C-1 proton), δ 5.2 (m, 1H, O-α- D-glucopyranosyl C-1 proton Example 3 Preparation of β-glucose-platinum (II) complex complex [C ₆ H ₁₀ (NH ₂ ) ₂ Pt (OOC) ₂ C ₃₃ H ₄₃ N ₃ O ₁₃ ] (R = C ₁₆ H ₃₃ N ₂ O ₁₁ , n = 2) 15 g of cellobiolactone instead of lactonolactone, and N-aminobutyl-O-β-D-galactopyranosyl- (1 → 4) -D-glucon Final β-glucose-platinum (II) complex in the same manner as in Example 1 using 0.40 g of N-aminobutyl-O-β-D-glucopyranosyl- (1 → 4) -D-gluconamide instead of amide 0.32 g of composite (total . Rate was obtained 35.5%) Composition _{formula: C 40 H 55 N 5 O} 15 Pt · 3H 2 O Elemental analysis: C, 34.35; H, 6. 36; N, 7.28; Pt, 18.51 Theory: C, 36.54; H, 6.03; N, 6.87; Pt, 19.15 hydrogen nuclear magnetic resonance spectrum (D ₂ O, ppm): δ 1.1-1.3 (b, 4H, diaminocyclohexane C-4, C-5 protons), δ 1.5-1.7 (m, 6H, diaminocyclohexane C-3, C-6 protons, -NHCH ₂ CH ₂ CH ₂ CH ₂ NH-), δ 1.8-2.4 (b, 8H, diaminocyclohexane C-1, C-2, C-3, C-6 protons, L-glutamate C- 2, C-3 protons), δ 2.4-2.7 (m, 4H, -C (O) CH ₂ CH ₂ CO-), δ 3.1-3.4 (m, 4H, -NH CH ₂ CH ₂ CH ₂ CH ₂ NH δ 3.4-4.3 (b, 13H, O-β-D-glucopyranosyl- (1 → 4) -D-gluconamideprotons, L-glutamate C-1 proton), δ 4.5 (m, 1H, O-β- D-glucopyranosyl C-1 proton Example 4 Preparation of α-glucose-platinum (II) complex complex [C ₆ H ₁₀ (NH ₂ ) ₂ Pt (OOC) ₂ C ₃₂ H ₄₁ N ₃ O ₁₃ ] (R = C ₁₆ H ₃₃ N ₂ O ₁₁ , n = 1) Aspartate derivative instead of glutamate derivative as spacer for introducing platinum (II) and N-aminobutyl-O-α-D-glucopyranosyl- (1 4) in the same manner as in Example 1, using 0.38 g of D-gluconamide and 0.54 g of 2-carboxyamidobenzyl aspartate. Longitudinally α- glucose-platinum (Ⅱ) complex conjugate 0.33g was obtained (total yield 38.9%) Composition _{formula: C 39 H 53 N 5 O} 15 Pt · 4H 2 O Elemental analysis: C, 34.35; H, 6. 36; N, 7.28; Pt, 18.51 Theory: C, 35.23; H, 6.01; N, 6.85; Pt, 19.07 Hydrogen nuclear magnetic resonance spectrum (D ₂ O, ppm): δ 1.1-1.3 (b, 4H, diaminocyclohexane C-4, C-5 protons), δ 1.5-1.7 (m, 6H, diaminocyclohexane C-3, C-6 protons, -NHCH ₂ CH ₂ CH ₂ CH ₂ NH-), δ 2.1 (b, 2H, diaminocyclohexane C-3, C-6 protons), δ 2.3-2.7 (m, 6H, diaminocyclohexane C-1, C-2 protons, -C (O) CH ₂ CH ₂ CO-), δ 3.1-3.4 (m, 4H, -NH CH ₂ CH ₂ CH ₂ CH ₂ NH-), δ 3.4-4.3 (b, 13H, O-α-D-glucopyranosyl- (1 → 4) -D-gluconamide protons, L-glutamate C-1 proton, δ 5.2 (m, 1H, O-α-D-glucopyranosyl C-1 proton) Example 5 Glucose Preparation of the amine-platinum (II) complex complex [C ₆ H ₁₀ (NH ₂ ) ₂ Pt (OOC) ₂ C ₂₃ H ₂₅ N ₂ O ₇ ] (R = C ₆ H ₁₂ NO ₅ , n = 2) N- Final glucoseamine was prepared in the same manner as in Example 1, using 0.5 g of glucoseamine hydrochloride and 0.78 ml of triethylamine instead of aminobutyl-O-β-D-galactopyranosyl- (1 → 4) -D-gluconamide. - platinum (ⅱ) complex conjugate 0.78g (total yield 46.7%) Composition _{formula: C 30 H 34 N 4 O} 9 Pt · 3H 2 O elemental analysis : C, 34.12; H, 5.0 4; N, 6.78; Pt, 24.04 theory: C, 32.99; H, 5. 27; N, 7.33; Pt, 25.51 Hydrogen Nuclear Magnetic Resonance Spectrum (D ₂ O, ppm): δ 1.1-1.3 (b, 4H, diaminocyclohexane C-4, C-5 protons), δ 1.5-1.7 (m, 2H, diaminocyclohexane C-3, C-6 protons), δ 1.8-2.4 (b, 8H, diaminocyclohexane C-1, C-2, C-3, C-6 protons, L-glutamate C-2, C-3 protons), δ 2.4-2.7 (m, 4H, -C (O) CH ₂ CH ₂ CO-), δ 3.4-4.1 (b, 7H, D-glucosamine protons, L-glutamate C-1 proton), δ 5.2 (m, 1H, D- glucosamine C-1 proton Example 6 Preparation of Mannosamine-Platinum (II) Complex Complex [C ₆ H ₁₀ (NH ₂ ) ₂ Pt (OOC) ₂ C ₂₃ H ₂₅ N ₂ O ₇ ] (R = C ₆ H ₁₂ NO ₅ , n = 2) 0.43 g of mannoseamine hydrochloride and 0.78 ml of triethylamine instead of N-aminobutyl-O-β-D-galactopyranosyl- (1 → 4) -D-gluconamide 0.56 g (total yield 42.1%) of the final mannoseamine-platinum (II) complex complex was obtained in the same manner as in Example 1. Formula: C ₃₀ H ₃₄ N ₄ O ₉ Pt.4H ₂ O Elemental Analysis: C, 33.47; H, 5. 12; N, 6.34; Pt, 22.18 Theory: C, 32.15; H, 5.40; N, 7.14; Pt, 24.86 Hydrogen Nuclear Magnetic Resonance Spectrum (D ₂ O, ppm): δ 1.1-1.3 (b, 4H, diaminocyclohexane C-4, C-5 protons), δ 1.5-1.7 (m, 2H, diaminocyclohexane C-3, C-6 protons), δ 1.8-2.4 (b, 8H, diaminocyclohexane C-1, C-2, C-3, C-6 protons, L-glutamate C-2, C-3 protons), δ 2.4-2.7 (m, 4H, -C (O) CH ₂ CH ₂ CO-), δ 3.4-4.1 (b, 7H, D-mannosamine protons, L-glutamate C-1 proton), δ 4.7 (m, 1H, D- mannosamine C-1 proton) Example of anticancer activity test The anticancer activity test for the saccharide-platinum (II) complex complexes of the present invention was performed for lung cancer (A549), ovarian cancer (SK-OV-3) and skin cancer (SK-Mel- 2), human cancer cell lines such as central nervous system cancer (XF-498), and colon cancer (HCT-15) and leukemia cell lines of rodents (L1210), and in vitro tests using mice. In vitro, each cancer cell line was cultured in RPMI 1640 culture solution, and then the concentration of the compound (ED ₅₀ ) which inhibited the growth of each cancer cell by 50% was obtained by adding a solution with different anticancer drug concentration to the cell solution. In vivo test, 8 mice of 6 to 8 weeks of age (BDFI mouse) were used as a group, and 10 ⁶ mouse leukemia cells L1210 per mouse were implanted in the abdominal cavity, and the carbohydrate-platinum (II) complex complex was 0.9%. After dissolving in physiological saline, 10-30mg / kg was administered by intraperitoneal injection for 1, 5, 9 days, and the average life extension effect (T / C,%) was observed. The results are shown in Table 1 and Table 2. It can be seen that the anticancer activity of the saccharide-platinum complexes of the present invention is much superior to conventional commercialized cisplatin or carboplatin. Table 1 In vitro anticancer activity against various cancer cells of the saccharide-platinum (II) complex complex Table 2 In vivo anticancer activity of leukemia cell line (L1210) of saccharide-platinum (II) complex complex ^{* If} T / C (%) = 125 or above, it is considered to have anticancer effect.

As described in detail above, an example of an anticancer activity test was performed while the platinum (II) complex ((DACH) Pt ²⁺ ), which is an anticancer component, was bound to a saccharide by using an amino acid group as an anion as a spacer, thereby alleviating the original toxicity of the platinum (II) complex. As shown in (Table 1, Table 2), the anticancer effect was found to be much better than the existing cisplatin or carboplatin, as well as to significantly improve the solubility of the insoluble platinum complex.

Claims (2)

The saccharide-platinum (II) complex complex represented by the following general formula (I). In Formula (I), R is a glucosamine (C ₆ H ₁₂ NO ₅ ), a mannose amine (C ₆ H ₁₂ NO ₅ ), a β-galactose derivative, N-aminobutyl-O-β- as a sugar and a sugar derivative. D-galactopyranosyl- (1 → 4) -D-gluconamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), an α-glucose derivative, N-aminobutyl-O-α-D-glucopyranosyl- ( 1 → 4) -D-gluconamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), β-glucose derivative N-aminobutyl-O-β-D-glucopyranosyl- (1 → 4) -D-glu Is selected from conamide (C ₁₆ H ₃₃ N ₂ O ₁₁ ), n is 1 or 2. The complex of formula (I) is prepared by reacting the alkaline earth metal salt of the sugar of formula (VI) with the platinum (II) derivative of formula (VII) in a 1: 1 molar ratio, removing barium sulfate as a by-product. Way