JPH0523276B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION (Industrial Field of Application) The present invention relates to a platinum complex having antitumor activity.

(Prior Art) Cisplatin is already commercially available as a platinum complex exhibiting an antitumor effect, and has been applied to many cases due to its remarkable effects. In addition to cisplatin, several reports have been made on platinum complexes that exhibit antitumor effects, and among them, those with linear alkyldiamine as a ligand have the general formula H ₂ N-CnR ₂ n- NH ₂ () (In the formula, R represents either a hydrogen atom or a substituent such as an alkyl group or a hydroxyl group. Also, n represents an integer of 1-3.) limited. (For example, JP-A-57-156416,
(or JP-A No. 56-103192) (Problems to be solved by the invention) As mentioned above, cisplatin is commercially available as a platinum complex cancer drug, but cisplatin has strong renal toxicity, which is a limiting factor for administration. . For this reason, it has the disadvantage that a large amount of water must be administered before and during administration, and a diuretic must be used in conjunction with the administration over a long period of time. Furthermore, cisplatin has low solubility in water and has a slow dissolution rate, so it is supplied at extremely low concentrations. Furthermore, cisplatin has extremely strong emetic toxicity, which poses a problem in treatment. For the above reasons,
Many studies have been carried out to find antitumor platinum complexes that have high solubility in water and low renal toxicity and emetic toxicity, but to date, no one has been found that has been put to practical use.

(Means for solving the problem) As shown in the formula () below, 2-methyl-
When 1,4-butanediamine coordinates with a platinum atom through two nitrogen atoms, a cyclic structure (7-membered ring structure) is formed by seven atoms including the platinum atom. Generally, it is extremely difficult to synthesize a complex having such a 7-membered ring structure, but as a result of intensive research, the present inventors synthesized a platinum () complex of the formula (), and this complex has an antitumor effect. It was found that the drug had significantly lower renal toxicity and emetic toxicity than cisplatin.

The present invention was completed based on these findings.

That is, the present invention provides the formula () This invention relates to a diamine platinum () complex represented by:

The compound of formula () of the present invention can be prepared by applying a known method, for example, the method described in Indian Journal of Chemistry (Indian J.Chem. 8 , 193, 1970), and partially modifying the reaction method. Obtainable.

The compound of the present invention is (In the above formula, M is an atom that can become a monovalent cation, for example, Na, K is a Cs, etc., and Hsl is a halogen atom such as Cl, Br, I, etc.) In water, as shown in the reaction formula, Tetrahalogenate platinum salt and amine are reacted to obtain dihalogenate diamine platinum. Preferably 5 to 160% water, particularly preferably 20% water per mole of tetrahalogenate platinum salt
Perform the reaction using 80 from Diamine is
For tetrahalogenate platinum salts, preferably
It is used in a range of 0.5 times mole to 4 times mole, particularly preferably 0.9 times mole to 1.2 times mole, and the reaction is carried out at 0°C to 100°C, preferably 50°C to 70°C, with stirring. It is preferable that the tetrahalogenate platinum salt aqueous solution and the diamine aqueous solution are mixed dropwise into distilled water and reacted at the same time at the same speed. It is preferable to drop the solution in small amounts, but it is usually carried out over a period of 1 to 4 hours. Although the reaction can be carried out in air, it is generally preferable to carry out the reaction under an inert gas stream, such as nitrogen. The dicarboxylate complex is then expressed as Suspend () in water, add a silver nitrate aqueous solution, and remove the resulting silver halide precipitate by filtration to obtain an aqueous solution of diacocomplex (). An appropriate amount of water can be used to suspend the dihalogenate-diamine complex (). Further, the amount of silver nitrate to be used is not particularly limited, but from an economic point of view, it is preferable to use 1 to 3 times the mole of the dihalogenate-diamine complex, and especially 1.9 times to 2 times the mole to avoid an excessive amount. It is preferable to use
The reaction is carried out at 0-100°C, preferably 60-80°C, with stirring. Add cyclobutane-1,1-dicarboxylic acid salt, cyclobutane-1,1-dicarboxylic acid monohydrogen salt, or cyclobutane-1,1-dicarboxylic acid to an aqueous solution of diaco complex () in an appropriate amount to (). Although it is allowed to react, it is generally preferable to add it in a range of 0.9 to 6 times the mole. The reaction can be carried out at 0 to 100°C, preferably 40 to 90°C, to obtain the compound ().

() + (cyclobutane-1,1-dicarboxylic acid salt or cyclobutane-1,1-dicarboxylic acid monohydrogen salt or cyclobutane-1,1-dicarboxylic acid) → () The compound () of the present invention can be obtained by elemental analysis, infrared Absorption spectra, fast atom bombardment mass spectrometry (FAB)
-MS Pt ¹⁹⁴ ) etc. to confirm the structure.

The compound of the present invention has extremely low renal toxicity and emetic toxicity, high solubility in water, and rapid dissolution rate, and has excellent antitumor effects, making it useful as an antitumor agent. Furthermore, the compounds of the present invention are stable in air at room temperature and do not require particularly low temperature storage. EXAMPLES Below, the present invention will be explained in more detail with reference to Examples.

(Example) Example 1 10 g of potassium chloroplatinate is dissolved in 350 ml of water, and 16 g of potassium iodide is dissolved in 50 ml of water and added with stirring. Stirring was continued for 5 minutes at 35°C to obtain a black aqueous solution of potassium iodoplatinate.
Also, 2-methyl-1,4-butanediamine 2.46
Dissolve g in 400ml of water. Pour 250 ml of water into a flask, and add 250 ml of water to the aqueous solution of potassium platinum iodide prepared above and 2.
-Methyl-1,4-butanediamine aqueous solution is simultaneously added dropwise at a constant rate over 2 hours. The precipitated reddish-brown crystals are collected by filtration, washed with water, and then washed with ethanol and ether. The obtained crystals were dried under vacuum to give disudiiodo-2-methyl-1,
9.94 g (yield 74.9%) of reddish brown crystals of 4-butanediamine platinum are obtained.

1 g of this cis-diiodo-2-methyl-1,4-butanediamine platinum is suspended in 20 ml of water, 604 mg of silver nitrate dissolved in 10 ml of water is added, and the mixture is reacted at 60° C. for 20 minutes with stirring. After cooling the solution to room temperature, it is filtered, and the silver iodide produced is separated and washed with water. Combine the filtrate and washing liquid, add cyclobutane-1,1-
Dicarboxylic acid 523mg in 1N sodium hydroxide aqueous solution
Dissolve in 7.29 ml, add, and stir at 60°C for 2 hours.
After concentrating the solution to 5 ml, it was cooled to 0°C and the white crystals formed were collected by filtration, washed with a small amount of water cooled to 0°C, washed with ethanol, and dried under vacuum to obtain the formula (). Obtain white crystals of the compound.

Yield 131 mg Elemental analysis calculation value (%): C, 30.07; H, 4.59; N,
6.38; Pt, 44.40 Actual value (%): C, 30.20; H, 4.31; N,
6.15; Pt, 44.50 FAB-MS: (M+H) ⁺ = 439 The solubility of the compound of the present invention in water is greater than 8 mg/ml, and in the IR absorption spectrum, 3200
-3125cm ^-1 (N-H) and 1700-1620cm ^-1 (C=
O) absorption.

The solubility of cisplatin in physiological saline is
Since the compound of the present invention is approximately 1.2 mg/ml, it is clearly highly water-soluble and has a high dissolution rate. Therefore, even when used as an injection, it is necessary to dissolve the crystals at an appropriate concentration at the time of use. Therefore, it is possible and suitable for immediate use.

Next, the antitumor effects of the compounds of the present invention will be explained by giving experimental examples.

Experimental Example 1 Proliferation inhibition test on subcultured murine leukemia L1210 cells (test method) Mouse leukemia cells subcultured using RPMI1640 medium containing 10% fetal bovine serum were tested with and without drug (compound of the present invention). The degree of growth inhibition (%) was calculated from the number of cells at the time of addition, and the IC ₅₀ was calculated from a graph plotting the drug treatment concentration and the degree of inhibition on log probability paper.
The value (50% growth inhibition concentration) was determined. the result,
The IC ₅₀ value was 1.20 μg/ml.

Thus, the compound of the present invention exhibits an inhibitory effect on the proliferation of cancerous cells at low concentrations.

The compound of the present invention acquired resistance upon administration of cisplatin. It also exhibits an excellent growth inhibitory effect on various cisplatin-resistant tumor cells, which will be explained below using experimental examples.

Experimental Example 2 Growth inhibition test for cisplatin-resistant tumor cells (test method) Mouse leukemia L1210 cells and P388 cells 1×
10 ⁵ mice were each implanted intraperitoneally into CDF ₁ female mice, and 6 mg/Kg of cisplatin was administered intraperitoneally on the second day after implantation. After another 5 days, the tumor cells were transferred to other
Transplant CDF ₁ into female mice intraperitoneally and perform the same treatment. This is repeated to obtain cisplatin-resistant tumor cells.

Using the tumor cells thus obtained, a proliferation inhibition test is conducted in the same manner as in Experimental Example 1.

Table 1 shows the ratio of the IC ₅₀ values for cisplatin-resistant tumor cells (IC ₅₀ R) and the IC ₅₀ values for tumor cells not resistant to cisplatin (IC ₅₀ R/IC ₅₀ ) obtained in this manner.

Table 1 IC ₅₀ R/IC ₅₀ tumor cells Compound L1210 P388 Cisplatin 11.4 10.7 Compound of the present invention 3.19 3.26 As is clear from Table 1, the compound of the present invention has a growth inhibitory effect even on cisplatin-resistant tumor cells at low concentrations. show.

Experimental Example 3 Antitumor test against mouse leukemia cell L1210 (test method) 1×10 ⁵ mouse leukemia L1210 cells were intraperitoneally transplanted into 6-week-old female CDF ₁ mice, and from the next day, once a day for 5 days. The drug (the compound of the present invention) was administered intraperitoneally. Physiological saline was similarly administered to the drug-untreated group. T/C was calculated from the average survival days of the drug-treated group and the non-treated group using the following formula.

T/C = (Average survival days of the drug-treated group) / (Average survival days of the non-drug treatment group) x 100 Furthermore, if any mice die due to acute toxicity of the drug during the test, the 50% lethal dose ( _LD50 )
was calculated using a standard method. Table 2 shows the results of Experimental Example 3.

In Table 2, max(T/C) indicates the maximum value of (T/C), and opt.dose indicates the dose (optimal dose) that indicates max(T/C).

Table 2 max (T/C) opt.dose LD ₅₀ (mg/Kg) (mg/Kg) 182 32 80.0 As is clear from Table 2, the compound of the present invention has a positive effect on mice inoculated with murine leukemia (L1210) cells. It has a life-prolonging effect.

The compound of the present invention also exhibits a remarkable survival prolonging effect on tumor cells other than mouse leukemia L1210 cells, which will be explained below using experimental examples.

Experimental Example 4 Antitumor test against various tumor cells (test method) 1 x 10 ⁶ mouse leukemia P388 cells were intraperitoneally transplanted into 6-week-old female CDF ₁ mice, and the drug was administered once a day for 5 days starting from the next day. (Compound of the present invention) was administered intraperitoneally. In addition, 1×10 ⁶ mouse lung cancer Lewis Lang carcinoma (LL) cells were intraperitoneally transplanted into male BDF ₁ mice, and the drug was intraperitoneally administered once a day for 5 days starting from the next day. Also, mouse fibrosarcoma
1×10 ⁶ M5076 cells were transplanted subcutaneously into the lateral region of the body of a female C57BL/6 mouse, and the drug was intraperitoneally administered once a day for 5 days starting from the next day. Also mouse colon cancer
1×10 ⁶ colon26 cells were intraperitoneally transplanted into female CDF ₁ mice, and the drug was intraperitoneally administered once a day for 5 days starting from the next day. Physiological saline was administered in the same manner to the no-drug treatment group.

The median survival time was determined from the survival days of the drug-treated group and the untreated group, and T/C was calculated using the following formula.

T/C=(median survival days of drug-treated group)/(median survival days of non-drug treatment group)×100 The results of Experimental Example 4 are shown in Table 3.

Table 3 Antitumor effect of the compound of the present invention on various tumor cells Tumor cell max (T/C) opt.dose (mg/Kg) P388 260 32 LL 222 32 M5076 152 16 colon26 198 32 As is clear from Table 3, the present invention The compound has a significant survival effect on mice implanted with many types of tumor cells.

Next, the renal toxicity of the compound of the present invention will be explained by giving experimental examples.

Experimental Example 5 Renal toxicity test (test method) A drug (the compound of the present invention or cisplatin) was intraperitoneally administered once to 6-week-old male CDF ₁ mice, and 4 days later, blood was collected and blood urea nitrogen concentration was determined. (BUN value) was calculated. Table 4 shows the results of Experimental Example 5. The opt.dose of cisplatin is 4 mg/Kg according to the test method shown in Experimental Example 3 above, but according to the above renal toxicity test method, a dose 4 times the opt.dose is already a normal value (30 mg/dl or less). far exceeds.
Therefore, in this experiment, as shown in Table 4, the compound of the present invention was tested at a dose 4 times or more of the opt.dose obtained in Experimental Example 3.

The body weight ratio in the table is the ratio of the body weight on the fourth day after drug administration to the body weight on the day of drug administration.

Table 4 Compound Dosage Body weight ratio BUN value (mg/Kg) (mg/dl) Physiological saline − 1.05 22.7 Cisplatin 16 0.72 92.9 Compound of the present invention 240 0.74 19.8 As is clear from Table 4, the compound of the present invention is superior to commercially available cisplatin. Compared to this, the BUN value in the blood is extremely low and is equivalent to that of physiological saline. This shows that it has extremely low renal toxicity. Therefore, the compound of the present invention serves as an antitumor agent with extremely low nephrotoxicity. Considering these characteristics and high water solubility, when administering intravenously as an injection, it is preferable to administer for a single hour rather than continuously.

The compound of the present invention uses a diamine having an asymmetric carbon atom as a ligand. We separated the optical isomers and used them as ligands to synthesize complexes and conducted tests. Reference examples, working examples, and experimental examples will be shown and explained below.

Reference example 1 (R-2-methyl-1,4-butanediamine) 40g of R-3-methyladibic acid was mixed with 200g of concentrated sulfuric acid,
Add to a mixed solution of 320 ml of benzene and heat to 45°C using an aqueous solution to dissolve 3-methyladipic acid.
Add 56 g of sodium azide little by little to this solution and react at 45-50°C. After the addition is complete and stirring is continued for 10 minutes, a saturated solution of 200 g of sodium hydroxide is added dropwise. The generated sodium sulfate is filtered off, and the benzene phase in the filtrate is separated. After extracting the aqueous phase with 500 ml of benzene and 500 ml of ether,
Extraction is carried out four times with 500 ml of chloroform. Combine the chloroform, benzene, and ether solutions,
After dehydration by adding anhydrous sodium sulfate, the sodium sulfate is filtered off and concentrated using a rotary evaporator. After concentration, vacuum distillation is performed to obtain R-2-methyl 1,4-butanediamine.

Yield: 6.92g (yield 27.1%) b・p・: 83°C/30mmHg Purity: 99.3% Optical purity: 100% In addition, in Reference Example 1 above and Reference Example 2 below, purity and optical purity are determined by gas chromatography. Determined by method and optical rotation measurement method.

Reference Example 2 (Isolation of optical isomers by optical resolution method of 2-methyl-1,4-butanediamine) Recrystallization method using racemic form of 2-methyl-1,4-butanediamine as salt of dibenzoyltartaric acid Optical resolution was carried out using the difference in solubility.

When obtaining R-2-methyl-1,4-butanediamine, (-)-dibenzoyltartaric acid is used,
When obtaining S-2-methyl-1,4-butanediamine, (+)-dibenzoyltartaric acid was used.
Table 5 shows the respective separation yields, purity, and optical purity.

Table 5 Resolution yield (%) Purity (%) Optical purity (%) R-form 57.8 100 98.6 S-form 51.4 100 98.8 Example 2 Using each optical isomer amine obtained in Reference Example 1 and Reference Example 2 Then, in the same manner as in Example 1, cis-cyclobutane-1,1-dicarboxylate-R-2-methyl-1,4-butanediamine platinum (compound R) and cis-cyclobutane-1,1-dicarboxylate-R-2-methyl-1,4-butanediamine platinum (compound R)
-dicarboxylate-S-2-methyl-1,4
-Butanediamine platinum (compound S) was obtained. Table 6
Table 7 shows the synthesis yield and elemental analysis values of each complex from potassium chloroplatinate, and Table 7 shows the physical properties. Further, the (M+H) ⁺ values confirmed by FAB-MS were both 439.

Table 6 Elemental analysis value (%) Compound Synthesis yield (%) C H N Pt R 24.6 29.98 4.43 6.22 44.8 S 23.1 30.21 4.37 6.36 45.0 Table 7 Compound 1R absorption spectrum in water (cm ^-1 ) Solubility (mg/ml) N-H C=O R >15 3200-3125 1700-1620 S >15 3210-3130 1700-1620 Experimental Example 6 IC ₅₀ values were determined for Compound R and Compound S by the same test method as Experimental Example 1 and Experimental Example 3. etc., and the obtained results are shown in Table 8.

Table 8 Compound IC ₅₀ max opt.dose (μg/Kg) (T/C) (mg/Kg) R 0.78 189 32 S 1.08 206 32 Compound LD ₅₀ (mg/Kg) R 33.6 S 48.0 Experimental example 7 Compound R and The max (T/C) etc. of Compound S were determined in the same manner as in Experimental Example 4, and the obtained results are shown in Table 9.

Table 9 Compound Tumor cell max (T/C) opt.dose (mg/Kg) R P388 253 20 R LL 166 30 S P388 253 40 S LL 164 50 Experimental example 8 Compound R and compound S were treated in the same manner as in experimental example 5. A kidney toxicity test was conducted using the drug, and the results are shown in Table 10. The dosages are shown in Table 8. The amount was 4 times the opt.dose.

Table 10 Compound Dose Body weight ratio BUN value (mg/Kg) (mg/dl) R 128 0.71 10.6 S 128 0.90 21.4 As is clear from the experimental results for Compound R and Compound S above, both of these compounds It can be seen that it has high solubility, exhibits excellent antitumor effects in various cells, and has extremely low renal toxicity.

(Effects of the Invention) The compound of the present invention exhibits an inhibitory effect on tumor cell growth at low concentrations and has an extremely excellent antitumor effect. In addition, it has high solubility in water and a fast dissolution rate. Furthermore, it has low renal toxicity and low emetic toxicity. In addition, the hematological toxicity generally observed in platinum complex antitumor agents is mild in the compounds of the present invention, mainly due to a decrease in the number of white blood cells, and the toxicity to platelets is slight. Furthermore, recovery to normal state is extremely rapid, and control is easy when used as an antitumor agent. For these reasons, the compound of the present invention can be used as an excellent antitumor agent. Furthermore, the compounds of the present invention are stable in air at room temperature and do not particularly require low-temperature storage.

Claims (1)

[Claims] 1 formula Diamine platinum () complex represented by