JPH01125325A - Antitumor agent - Google Patents

Abstract

PURPOSE:To obtain an antitumor agent having excellent antitumor activity and low side effect, by using a platinum green complex wherein the number of platinum atoms. in the molecule falls within a specific range. CONSTITUTION:The objective agent contains, as an active component, a platinum green complex containing 3-18, preferably 6-15 platinum atoms. in 1mol., obtained by reacting (A) a compound of formula [X is SO4<2->, (NO3<->)2 or (ClO4<->)2] with (B) uridine, thymidine, uracil, thymine, 2'-deoxyuridine, uridine-5'- monophosphate or 5-fluorouracil in a proper solvent preferably at 3-6pH and 1-75 deg.C in the presence of hydrogen peroxide, and having a structure consisting of cis-diaquodiamine platinum (II) and the above base or base derivative coordinated to the platinum compound.

Description

[Detailed description of the invention] [Industrial application field] The present invention relates to a novel antitumor agent.

[Conventional technology]

Cisplatin, which was proven to have antitumor activity by Rosenberg et al. (Nature, Vol. 222, p. 385 (1969)), is effective in the treatment of prostate cancer, ovarian cancer, gastric cancer, etc. due to its excellent action. However, side effects such as nephrotoxicity, nausea, and vomiting are a major problem. In addition to improving administration methods to reduce these side effects, Synthetic research on platinum complex compounds with excellent tumor activity and reduced side effects has been actively conducted.

U.S. Pat. No. 4,419,351 states that 2,4-
A platinum-[2,4-dioxopyrimidine compound] blue or green complex, or a mixture thereof, which has antitumor, antibacterial, and antiviral effects by reacting a dioxopyrimidine compound with cis-cyacocyanmineplatinum (II) It is stated that this can be obtained.

Also, the Journal of the American Chemical
Society (J, Am, Chem, Soc,) 10th
6, pp. 2049-2054 (1984) reports on the structure of a green complex of cis-diaminedichloroplatinum (n) and α-pyrrolidone.

By the way, according to the above-mentioned US patent specification, platinum complexes are obtained with bases such as uracil, thymine, pyrimidine, and polyuracil, and with uridine derivatives or thymidine derivatives. Regarding complexes with bases, a platinum blue complex was obtained as the main component, and a platinum green complex was obtained as a secondary precipitate. The platinum blue complex has been identified by elemental analysis, visible absorption spectrum, etc., and its biological activity has been investigated. However, the physicochemical properties of platinum green complexes are unclear, and in particular, the purification methods and methods for water-soluble platinum green complexes that do not precipitate are unclear.
There are no reports regarding physicochemical properties, antitumor activity, etc. On the other hand, regarding complexes with uridine derivatives or thymidine derivatives, only blue complexes have been obtained, and only some antitumor activity has been reported.

Regarding the green complexes of uridine derivatives or thymidine derivatives, neither the production method nor the antitumor activity is described.

[Problems to be Solved by the Invention] An object of the present invention is to obtain an antitumor agent with excellent antitumor activity and fewer side effects.

[Means for solving problems]

The present inventors found that the main antitumor activity of the complex of platinum and nucleoside is due to the green complex, that a pure blue complex does not have such an effect, and that among the green complexes, especially those with a platinum number It was discovered that the activity is high when the molecular size is 3 to 18, and the present invention was completed.

That is, the present invention provides the general formula (wherein, X is SOa'', (NO3)z, (ClO2
−) 2) and uridine, thymidine, uracil,
Uridine, thymidine, Uracil,
Thymine, 2'-deoxyuridine, uridine-5'-monophosphate or 5-fluorouracil is coordinated,
It is an antitumor agent whose active ingredient is a platinum green complex having 3 to 18 platinum atoms in the molecule.

The reaction is carried out using an appropriate solvent (water, methanol, ethanol,
Alcohol solvents such as butanol, ether solvents such as dimethoxyethane, tetrahydrofuran, and dioxane, dimethylformamide, dimethyl sulfoxide,
Hexamethylenephosphonoamide, acetic acid, pyridine, etc.
or a mixed solvent thereof) at pH 2-8, preferably pH 3-6, 0-100'C, preferably 1-75°C.
at a temperature of 30 minutes to 2 months, preferably 45 minutes.
It will progress in one month.

The reaction molar ratio between the compound of general formula (1) and the base or base derivative may be in the range of 1:1 to 1:4, but is generally optimally around 1:1.

The hydrogen peroxide used is preferably 0.1-10% hydrogen peroxide.

The platinum green complex thus obtained can be separated and purified by conventional methods. That is, the reaction mixture can be separated by various chromatography methods such as solvent extraction, precipitation, ion exchange, partitioning, gel filtration, and affinity.

In the method of the present invention, gel filtration is particularly suitable, and any of Sephadex, Sephaacryl, Sepharose, Biogel, Toyopearl, Fractogel, Cellulofine, etc. can be used as the carrier. These operations are preferably performed under anaerobic conditions, such as under a nitrogen stream.

The platinum complex thus separated is identified using its own specific visible absorption spectrum.

For example, a platinum green complex using uridine as a ligand has characteristic absorption in the 730 nm region.

In the ultraviolet region, strong π→π1 absorption is exhibited near 260 to 27 Onm.

The C-0 stretching vibration is 164 in the infrared absorption spectrum (IR).
5 cm-' (16 for uridine)
90 cm-'), and the carbons corresponding to C-4 and C-2 of uridine are very large in the I3Cnmr (nuclear magnetic resonance) spectrum (11 to 8 pf'm in the former).
+6 to 4 ppm in the latter), which strongly suggests that the C-4 or C-2 position of uridine is coordinated with platinum (bivalent or trivalent). In this green complex, a paramagnetic resonance spectrum (ESR) of Pt(III) was observed. This spectrum shows a typical axisymmetric pattern with g = 1.991.
gl=2.413. Examination of the hyperfine structure revealed that unpaired electrons are not localized in one platinum, but delocalized in several platinum.

Table 1 shows the synthesis results of the platinum green complex under various conditions.

(1) 2 8 0.
1 3 Pass 49.5 (2) 2 Δ
0.1 4wk 29.5 (3) 25
A O, 148h 12.6 (4)
40 Δ 0.1 3.7h 4
0(5) 50 A O, 12h
36.6 (6) 75 A O
,013h 17.9(7) 75
A 0.1 45m1n 41.9 (8)
75 A O, 145m1n 3
2 (9) 75 8
0.1 90m1n 35.800) (bl
ue) 60 B O, 141h 4
0a) A: 1% hydrogen peroxide solution B: Oxidation with air b) Substrate concentration C) Platinum tetranuclear complex, calculated assuming an average platinum valence of 2.25 The elemental analysis values for each of these compounds are as follows. be.

Compound (1) Calculated value: (pt+.(C9HIINZO6)6(NH
3) 2゜(OH) Z (n2o) b ) (S04)
As s/7nzo: C, 13,55; H, 3,2
4;N, 9.36. S, 5.36. Pt, 40.7
5; Measured value: C, 13, 84; H, 3, 21, N,
9.29. S, 5.35; Pt, 40.72 Compound (2) Calculated value: [Pt++(CJ++Nz06)9(NH:
+)zo(OH1+(HzO)3)(504)? /8
As H20: C, 17,38; H, 3,31; N,
9.51; S, 4.01. Pt, 38.34; Measured value: C, 17,21, H, 3,19, N, 9.50.
S, 3.94°Pt, 38.47 Compound (3) Calculated value: (pt,.(C9HII N206) II
(NHI) + q (OH) (H□0)4] (S
O4)? /10820 and 7:C, 16,75; H
, 3,40; N, 9.50; S, 4.35; P
t, 37.80; Measured value: C, 16,96; H, 3,
19; N, 9.37; S, 4.32°Pt, 38.
22 Compound (4) Calculated value: [Pt, (C7HIIN206)4(NHl)
, 6(oH)3(n2o), ](SO4)? /2H2
As 0: C, 11,02; H, 3,01; N, 8.
5'7; S, 5.72; Pt, 44.76; Measured value: C, 11,39; H, 2,91; N, 8.61;
S, 5.77; Pt, 44.57 Compound (5) Calculated value: (Pt+o(C7HzNzO6)s(NHs
)+e(OH')z(HzO)+o)(SO4) II
/2H20: C, 12,03; H, 3,03;
N, 8.73; S, 5.71; Pt, 43.43;
Measured values: C, 11,98; H, 2,95; N, 8
．． 79; S, 5.71; Pt, 43.34 Compound (6) Calculated value: (Ptll(CJ++Nz06)s(NH3
)+5(OH)z(HzO)s)(S04)6/3■2
As 0: C, 14, 27; H, 3, 14; N, 9.
25; S, 5.08; Pt, 41.22; Measured value:
C, 13,94; u, a, to; N, 9.24;
S, 4.92: Pt, 41.42 Compound (7) Calculated value: fPt+:+(CqH++Nz06)+□(
Nni) z:l (OH) (H2O) a ) (5
04) As 9/18820: C, 1B, 21; H
,3,4B.

N, 9.24. S, 4.05. Pt, 35.60;
Measured value: C, 17,76; ■+3.14; N, 9.
23. S, 4.03°Pt, 35.59 Compound (8) Calculated value: (Pt14(C9H11N206)+4(N
H3)21(OH)3(H2O)2)(SO4) 8/
As 18H20: C, 19,63; N+3.48;
N, 9.27; S, 3.33; Pt, 35.44;
Measured values: C, 19, 24, H, 3, 22, N, 9.1
6. S, 3.30°Pt, 35.72 Compound (9) Calculated value: (Pt+6(CJzNzO6)+6(NH:
+)za(Oil)z(HzO)z)(S04)I. /
As 25Hz: C, 19,28; H, 3,55°N, 9.37; S, 3.57; Pt, 34.80B
Measured values: C, 19,42; H, 3,20; N, 9
．． 16; S, 3.62; Pt, 34.31 Compound 00) Calculated value: (Ptzs(CJ(zNzo6)z8(NH
:+)zq(OH)z7)(SO4)+□/9H,0 as:C,20,8'2:H,3,03;N,8.1
9; S, 2.65; Pt, 37.58° Measured value:
C, 20,55; H, 3,02; N+8.20;
S, 2.64; Pt, 37.97 Further, Table 2 summarizes the main data of the platinum green complex.

(Blank below the essence) Table 2 Main data of platinum uridine green complex (1)
268 (4,85) 300 10: 6:
20 2.40 (2) 265 (4,86)
300 11: 9:20 2.36 (3)
270 (4,81) 300 10: 8:1
9 2.30 (4) 266 (4,55)
300 9: 4:16 2.33 (5)
268 (4,66) 300 10: 5:18
2.30 (6) 265 (4,61) 30
0 8: 5:15 2.38 (7) 26
7 (5,00) 300 13:12:23 2
．． 38 (8) 267 (5,06) 300
14:14:23 2.36 (9) 265
(5,14) 300 16:16:28 2.
Synthesis conditions and elemental analysis values of 38 other compounds are shown below.

Synthesis condition (II) 23χH,O□ 0.13k 44.90
2) 26χH2O20,13 discharge 17.403) 21χ
H20□ 0.1 4wk 29.5Q4) 401χ
HzOz O, 0548hr 50. IQ5) 75
1χH20Z O, 145m1n 19 elemental analysis value Compound (11) Calculated value: (Pt+o(CJI++N206)6(NH
3) 2o(OH)z(HzO)6〕(S04)II/8
As H20: C, 13,50; H, 3,27°N,
9.33. S, 5.34. Pt, 40.60° Measured value: C, 13,42, H, 3,18; N, 9.15
．． S, 5.25゜pt, 40.48 Compound 02) Calculated value: (Ptz(CqH++Nz0Jb(NH3)
z□(OH) 3 (HzO)? ] (SO4) 9
/12H2O as: C, 12,38; H, 3,33
; N+9.09; S, 5.51; Pt, 40.
97: Measured value: C, 12, 36; 14, 3.17;
N+8.99; S+5.44; Pt, 40.70 Compound θ3) Calculated value: (Pt++(CJzNzOs)9(NH3)
z. (OH) * (H2O) 3 ) (S04)?
/8H20: C, 17, 38; H, 3, 31;
N+9.51; s, 4. Old, Pt, 38.34; Measured value: C, 17,21; H, 3,19; N, 9.
50; S, 3.94°Pt, 38.47 Compound side calculated value: (Pt+o(CJzNz06), (NH3)
+t(OH)z(HzO)7)(S04)? /3H2
As 0: C, 15,67; H, 3,13°N, 8.
99; S, 4.65; Pt, 40.40: Measured value:
C, 15,86, H, 3,08; N, 8.90:
S, 4.62°Pt, 40.47 Compound 05) Calculated value: (Pt+3(CJl++Nz06)+x(N
H3) zo(OH)3(LO):+)(504),/
As 13HzO: C, 19,93; N+3.40;
N+9-14; S, 3.18. Pt, 35.97° Measured value: C, 19,63; H, 3,25, N, 9.
15. S, 3.1B.

Pt, 36.27 The compound of general formula (1), which is a starting material, is prepared by reacting cisplatin or cis-diammine short platinum (II) with silver nitrate or silver perchlorate, preferably under heating, or by reacting cis-diammine short platinum (II) with silver nitrate or silver perchlorate, preferably under heating. It is produced by reacting platinum (n) with silver sulfate.

As is clear from the following experimental examples, the platinum green complex of the present invention exhibits strong antitumor activity and is effective as an anticancer agent.

Experimental Example: Antitumor activity against L1210 The antitumor activity of the platinum green complex of the present invention was measured by determining the growth inhibition rate using L1210 cultured cells. That is, a test sample (final concentration of 5 μg/vl or 10 μg/vl) was added to a solution containing 1 × 10 cells per cell.
n1) and cultured at 37°C for usually 4 days. The medium contained G I B C01640゜Kanamycin and 1
0% F B S (Fatal Bovin Seru
m) was used. Control (usually 1.4×
106 cells/m! The inhibition rate was calculated from the growth ratio of The results are shown in Table 3.

(Margin below essence) αυ 10:6:2078.092.8--ano
11: 6:22 59.2 93.
7 ---Q31 11: 9
:20 77.1 0.43
84.1 (3) 10: 8:19
84.8 1.34 9
1.3 (4) 9: 4:16
99.3 3.0 99.8α4
1 10: 8:18 2
91.6 2.05 97.3(
618:5:15--99.2
3.25 99.805) 13:13
:20 32.4 68.3-1 (811
4:14:23 12.3 51.9
---(7) 13:12:2
3 20.9 69.2--(9)
16:16:2B 35.5
- -11; )... b) Ratio of giant and normal L 1210 cells after 4 days at 37°C with a dose of 10 μg/d C) Growth inhibition at 10 μg/d administration measured excluding giant cells d) Pt When the blue complex (oxidized by air) was administered at 10 μg/Tn1, the green complex showed high activity, but the blue complex was completely inactive, showing a remarkable difference. Furthermore, a detailed study shows that even within the same green body, the growth inhibition rate varies widely, ranging from 35.5 to 99.3%, depending on the synthesis conditions.

This is found to be correlated with activity when considering the size of the molecule. FIG. 1 illustrates the relationship between growth inhibition rate (%) and molecular size converted to platinum number. As is clear from the figure, smaller molecules have higher activity than larger molecules.

That is, those having a platinum number of 3 to 18, particularly those having 6 to 15 platinum, are highly active. Exactly the same tendency was observed for 5 μg/ml administration (Table 3, Figure 1).

More interestingly, compared to normal L 1210 cells,
The generation of giant cells with approximately twice the diameter was found. The numbers of both were counted separately and the ratio of giant cells to normal cells was determined and is shown in Table 3. In the case of highly active green bodies, it was found that the proportion of giant cells was high. For example, in the case of bull monochrome, no giant cells were observed, but in the most effective case, more than 374 of the remaining cells (only 0.7-0.8% compared to the control) It was a giant cell.

The last column of Table 3 shows the L when those giant cells are excluded.
oug/m! The inhibition rate at the time of administration is shown.

The findings we have obtained here are noteworthy as they are thought to be the first example of giant cells being generated using a platinum green complex using animal cells. On the other hand, it was reported that giant cells were formed in Escherichia coli when growth under an electric field was observed using a PT electrode about 20 years ago [Nature, No. 205]
Volume, 689 pages (1965)].

As described above, it has been revealed that platinum green is active against L 1210, but blue is inactive. Furthermore, a clear correlation between the molecular size of the green body and its activity was observed. The formation of giant cells was discovered for the first time with a platinum green complex, and its relationship with biological activity became clear. These factors are thought to be related to the permeability of the membrane.

When the compound of the present invention is used as a medicine, it can be mixed with appropriate pharmacologically acceptable excipients, carriers, diluents, and other pharmaceutical formulation additives to form tablets, granules, powders, capsules, and injections. It can be administered orally or parenterally in the form of tablets, ointments, and tablets. The dose may vary depending on the tumor type, symptoms, administration form, patient age, weight, etc., but in general, the effective dose is 0.1 to 5.000 m
g/kg, preferably 0.1-1,000mg/k
g, more preferably 1 to 500 mg/kg, which can be administered in 1 to 6 divided doses a day.

〔Example〕

The present invention will be described below with reference to Reference Examples, Production Examples, and Examples; however, the present invention is not limited to these Reference Examples and Examples.

Reference example: Synthesis of cis-cyacocyanmineplatinum (I[) sulfate 145 mg of cis-short diammine platinum and 93 mg of silver sulfate
．． 3 d of water was added to 5 mg, and the mixture was stirred for Ω~30'C1-night while shielding from light. The precipitated silver iodide was filtered off, and the filtrate was used as cis-short diammine platinum (n) sulfate in the following production examples.

Production example Cis-cyacocyamine platinum (I[) sulfate (0.3m
mol) Uridine 73.2 mg (0,3
mmol), adjusted to pH 4.3 with 0.5N sodium hydroxide solution, and diluted with 1% hydrogen peroxide solution (336μ!). added.

The mixture was stirred at 40°C under an argon stream and reacted for 3.7 hours. Gel filtration of the resulting reaction solution (carrier: Toyopearl)
Purification was performed to obtain 63.9 cm of a platinum green complex containing green uridine as a ligand. The above operations were performed under nitrogen or argon.

Visible absorption: λmax 730nm IR (KBr): 1645 (C=O), 87
0.815.585(Pt-N)cyn-' Melting point: >300 °C Circular dichroism (CD spectrum): [θ] (λmax nm); 37,800 (272
).

-37,800 (219) "C-Nmr (D20; DSS standard): p
pm63.5.72.1.76.3.86;9.92.
8°104.7.143.6.160.2 and 158.
5゜179.8 and 176.7 Patent applicant Kozo Iizuka, Director of the Agency of Industrial Science and Technology

Claims (1)

[Claims] 1. General formula ▲ There are mathematical formulas, chemical formulas, tables, etc. ▼ (In the formula, X is SO_4^2^-, (NO_3^-)_2
or (CIO_4^-_)_2) and uridine, thymidine, uracil,
Uridine, thymidine, uracil, and cis-diacodiamine platinum (II) obtained by reacting thymine, 2'-deoxyuridine, uridine-5'-monophosphate, or 5-fluorouracil in the presence of hydrogen peroxide. An antitumor agent containing as an active ingredient a platinum green complex coordinated with thymine, 2'-deoxyuridine, uridine-5-monophosphate, or 5-fluorouracil and having 3 to 18 platinum atoms in the molecule. 2. The antitumor agent according to claim 1, wherein the number of platinum in the molecule is 6 to 15.