DE2931079C2 - 1-Di- (ethyleneimido) -thiophosphoryl-2-trichloromethyl-Δ 2 -imidazoline, its preparation and pharmaceuticals containing this compound - Google Patents

Description

15th

CH ₂

CH ₂

CH ₂

2. A process for the preparation of l-di- (äthylenimiuo) -ihiüphu5pnory! -2-tnCn! Onncthy ¹ i- <d'-iiTiidazoline of the formula I, characterized in that one

N-dichlorothiophosphoryliminotrichloroacetic acid chloride with ethylene imine at temperatures from 0 to 5 ° C in the counterv / art of benzene, toluene, diethyl ether or carbon tetrachloride as a solvent and triethylamine as a binder for hydrogen chloride implements.

3 drugs containing l-di- (ethyleneimido) -thiophosphoryl-2-trichloromethyl-zl ² -imidazoIin.

The invention relates to l-di- (äthyIenimido) -thiophosphoryl-2-trichloromethyl- / l ² -imidazoline, a process for its production, and containing this compound, possessing a broad spectrum of lumor-inhibiting effects and for the healing of malignant neoplasms of epilhelial and mesenchymal origin as well Medicines suitable for inhibiting the growth of lymphoid cells.

The l-di- (ethyleneimido) -thiophosphoryl-2-trichloromethyl-zl ² -imidazoline according to the invention is a new substance of the following formula that has not yet been described in the literature:

50

CI ₃ CC —Ν — Ρ

(I)

CH ₂

CH ₂

55

60

This compound is a colorless crystalline substance, which fall, however, is soluble in acetone _(methanol and at a heating in ethanol, chloroform, benzene, dimethylsulfoxide and the ether insoluble in water and hexane. The melting temperature is in a range VOFI 137-138 ⁰ C under Decomposition: When heated above the melting temperature, it gradually changes color and disintegrates explosively at a temperature of 165 to 170 ° C.

In the IR spectrum the connection becomes intense Absorptions at 942 cm - 'and from 1272 cm -' marked, which are to be assigned to the asymmetrical oscillations of ethyleneimine rings, as well as by Absorptions at 515 cm- '(P = S) and 1623 cm-' (C = N).

In the NMR spectrum, the chemical shift of ³¹ P + 96.6 ppm in the weak field with respect to 85% H ₃ PO ₂ . In the proton resonance spectrum, the multiplet of the ethyleneimine protons appears with a chemical shift of 0 = 2.04 ppm (8 protons).

The protons of the imidazoline ring appear as 2 triplets with a chemical shift of <5 = 4.22 ppm (2 protons) and <5 = 3.78 ppm (2 protons) with a coupling constant of ³ J (HJCH) 8.5 Hz The protons of the imidazoline ring are also ^{split by phospho 'with the constants 3} J (PNCH) = ^ Oh and - ¹ J (PNCCH) = 3 Hz.

The anti-tumor activity and the toxic properties of l-di- (äthylenimido) -thiophosphoryl-2-trichloromethyl-4 ^J -i.Tiidazolins were examined in experiments in mice and rats.

The anti-tumor activity was also demonstrated on four strains of inoculated ascites tumors from mice namely Ehrlich's carcinoma. Sarcoma 37. NK / Ly lymphadenosis. Fischer's lymphadenosis L 5178 and a strain of solid tumors in rats Guerin cariinoma checked.

Strains of inoculated ascites tumors from mice were implanted in white unbred mice with a mean weight of 20 to 23 g. The native six to seven day ascites fluid was diluted with an aseptic physiological saline solution in a ratio of 1 to 10 and injected intraperitoneally into the mice in an amount of 0.5 ml. The test animals were divided into two groups, namely the control group and the test group. On the second day after the tumor transplantation, the animals in the test group were administered 1-di- (ethyleneimido) -thiophosphoryl-2-trichloromethyl-4 ² -imidazoline in the form of an 0.1 percent suspension in saline. The preparation described was injected subcutaneously every two days at a dose corresponding to 1/14 LDw. The injection was triple. The mice were sacrificed on the 8th to 9th day after tumor implantation. The volume of ascites fluid stored in the animals of the control and test group was determined.

Rats of the "Wistar" line with an average weight of 80 to 125 g received 1 ml of a 30 percent Suspension of Guerin carcinoma cells implanted subcutaneously. On the 7th to 8th day after the tumor transplant the preparation was injected at a dose corresponding to 1/10 LD50. The animals in this case were in divided into four groups. The preparation was used once for the first group, for the second and third groups injected four to five times every two days. On animals the Rö / HroligrUppe the preparation was not used. On the 20th to the 25th day after the tumor transplant the rats were killed and the tumor removed and weighed.

As a feature of anti-tumor activity were the percentage of inhibition of tumor growth, the degree of effectiveness and the percentage of

Primary healing counted. statistically evaluated test results are in

The table 1 reproduced according to the Montsevichute-Eringene method.

Table 1

Tumor-inhibiting activity of l-di- {ethyleneimido) -thiophosphoryl-2-trichlonnethyl-A ² -

imidazolins

Animal species Tumor stem dose Injection Inhibition Effect percent tough! percentage Degree sentence of of the tumor Primary off lung

Mice sarcoma 37 1/14 LD ₅₀

Mice Ehrlich- 1/14 LD ₅₀

carcinoma
Mice NK / Ly lymph-1/14 LD ₅₀

adenosis
Mice Fischer lymph 1/14 LD ₅₀

adenosis L

5178
Rat Guerin- 1/10 LD ₅₀

carcinoma
Rat Guerin- 1/10 LD ₅₀

carcinoma
Rat Guerin- 1/10 LD ₅₀

carcinoma

The preparation proved to be most effective in experiments on rats with an implanted strain of Five injection Guerin carcinoma.

The rats in which the tumor was largely resorbed after injection of the 1-di ^ äthylenimidoJ-thiophosphoryl ^ -trichloromethyl-4 ² -imidazoIine proved to be resistant to repeated implantation of the Guerin carcinoma.

The toxicity was tested in white unbred mice with an average weight of 20 to 23 g and rats of the "Wistar" line with an average weight of 80 to 125 g with subcutaneous, intramuscular, intraperitoneal and peroral administration. The l-di- (äthyIenimido) -thiophosphoryl-2-trichloromethyl-4 ² -imidazohn was prepared extempore with physiological saline solution and introduced in the form of a fine suspension with a concentration of at most 4%. The dose of the preparation was 1500 to 10 mg per kg of animal weight.

Each dose of the preparation was injected into at least 5 to 6 animals. The deaths of animals in the test group were recorded on the 8th to 10th day after Introduction of the preparation.

The toxicity was calculated in mg of the active ingredient per kg of the animal weight.

The statistically according to ütchfield and Wilcoxon in the Modifications of test results evaluated by Roth are summarized in Table 2.

Table 2

Toxicity of l-di- (ethyleneimido) -thiophosphoryl-2-trichloromethyl- / l ² -imidazoline in an acute experiment on mice (mg / kg)

97.2
92.6

90.0
62.0

68.0
99.85
100

35.5
13.4

10.0
2.6

3.1
285

52.9
33.4

27.3
6.2

50
100

In an acute experiment on rats, the following results were obtained:

Introduction type

LD ₁₀ LDi ₆ LD ₅₀ LD ₈₄ LD.

Subcutaneous 200 215 285 380 410

Intramuscular 160 175 250 360 400

Peroral 165 180 260 375 410

Intraperitoiieai 40 45 64 90 100

with subcutaneous injection

from LD »200 mg / kg

for intramuscular injection
of LDm 150 mg / kg

when administered orally

of LD ₅ O 220 mg / kg

when injected intraperitoneally

from LD50 35 mg / kg

The influence of 1-di- (ethyleneimido) -thiophosphoryl-2-trich! OrmethyM ² -imidazoIins on the content of form elements of the peripheral blood and the morphological condition of bone marrow cells was determined in rats of the "Wistar" line with a mean weight of 80 to 125 g examined.

The substance was in the form of a 0.1 percent

Suspension in physiological saline solution administered four times every two days at a dose of 1/10 LD50 In the peripheral blood of the animals of the test group, the content of leukocytes, erythrocytes, Platelets, reticulocytes, the leukocyte formula, the sugar, urea and hemoglobin content, the Blood clotting speed, the sedimentation reaction, the fermentation activity of the blood serum and the cholinsterase activity were determined. The values were determined before the introduction of the preparation and during the healing period After the healing period became part of the animals to study the morphological composition of the bone marrow killed while the other animals were used for further examination. At for further investigations The blood was removed from living rats 20, 30 and 60 days after the last administration of the Examined preparation. The rats of the experimental group

- Tümörträger - died on the 20th to 25th day after the Tumor transplantation.

The single administration of the preparation in an amount of 1/10 LD ₅₀ does not lead to any noticeable

Changes in the morphological composition of the blood in all of the above indicators. the Cholinsterase activity was not changed after the fourth administration of the preparation. During the In the "in vitro" process, the pseudo and "true" cholinsterase activity was not influenced by the preparation.

Changes in the morphological composition of the blood occurred after the third administration of the Preparations * and were significant after the fourth administration. B. after after the third administration by 28% and after the fourth introduction by 44.7% and was 7240 + 1140. the The white blood cell count started on the fourth day after the drug administration was discontinued and was restored to baseline in two weeks after the last injection

The morphological composition of the bone marrow cells was determined in rat tumor carriers Examined 6th day after the fourth administration of the preparation. The tests showed that the

Table 3

Red blood cell count remains unchanged. In most of the animals none were noticeable Changes also found in the morphological examination of the myeloblast cells. The mitoti-

see cell division was preserved. However, it was at 20% of the treated animals a small uniform vacuole formation in the stage of myelocytes without nuclear displacement and cell volume change observed, <vas dystrophic changes in the cytoplasm and dts

Nucleus of the cells brought about by the myeloid series. In the bone marrow of the animals in this group cells with mitotic division were observed less frequently

The toxicity, cholinesterase activity and therapeutic dose of l-di- (ethyleneimido) -thiophosphoryi-2-trichloromethyl-4 ² -imidazoline were compared with the effects of known preparations such as phosphoric acid-N-benzoyl-N ', N' , N ", N" -diethylenetriamide and N-diethylenamidophosphoryliminotrichloroacetäthylenamid investigated

links

Toxicity in cholinesterase activity
mg / kg LD ₅₀

molar cholesterase

Concentration inhibition

Therapeutic
dose

l-Di- (ethylenimido) -thiophosphoryl-2-trichloromethyl - ^ - imidazoline 235

Phosphoric acid-N-benzoin-Ν ', Ν', Ν'νΝ "-diethylenamid 35

N-Diethylenamidophosphonyliminotrichloracetat-Ethylenamid 42

From the table it can be seen that the l-di- (äthylenimido) -thiophosphoryl-2-trichIormet ^'i y - / l ² -imidazoline is characterized by low toxicity and by a wide therapeutic spectrum of action compared to the effect of the known preparations and that it does not affect the autonomic nervous system.

The 1 -di (ethyleneimido) -thiophosphorus) 'l-2-trichloromethyl- / d ² -imidazoline is prepared by reacting N-dichlorothiophoryliminotrichloroacetic acid chloride with ethyleneimine in the presence of triethylamine. The liberated hydrogen chloride is bound as a salt by the excess triethylamine. The reaction is carried out in the presence of an inert organic solvent, e.g. B. Ben ^ oI. Toluene, diethyl ether or carbon tetrachloride carried out at a temperature of 0 to 5 ° C. The yield of the end product is 31 percent by weight. To prepare the starting substance, N-dichlorophosphoryliminotrichloroacetic acid chloride is reacted with phosphorus pentasulphide at a temperature of 140 to 170 ° C. The end product is distilled off in vacuo. N-Dichlorthiophosphbryliminotrichloressigsäurechlorid is obtained in the form of a clear light yellow liquid with a specific weight of 1,7,346,

The following examples serve to illustrate the invention.

ΙΟ " ² - By S play 1/14 LD ₅₀ 3.9 10- ' 76 1/5 LD ₅₀ 3 · 10-3 50 1/10 LD ₅₀ 1

aJN-dichlorothiophospheryliminotrichloroacetic acid chloride

50

34.6 g of N-dichlorophosphoryliminotrichloroacetic acid chloride and 6.5 g of phosphorus pentasulfide are ^{heated to temperatures of 140 to 170 °} C. in an oil bath with gentle stirring. A low-boiling substance forms after 1 to 1/2 hours. After 4 hours the reflux condenser is replaced by a distillation condenser and the low-boiling fractions - CCI3CN and PSClj - are distilled off in the vacuum of the water jet pump.

The remainder is distilled in the vacuum of the oil pump at 75 to 76 ° C (0.13 mbar). 14.9 g are hardened accordingly (48%) of theory N-dichlorothiophosphoryliminotrichloroacetic acid chloride.

C ₂ CIcNPS:
Found;

N4,52,4,65; P9,88 ₎ 9 ₁ 59;

S 10,66,10,62; Cl 68.21.68.37;

C 8.72.7.92 weight percent MRd60.9.
Calculated:

N 4.47; P 9.81; S 10.19;

Cl 67.88, C 7.65 percent by weight MÄd60.3.

b) I -di (ethyleneimido) -thiophosphoryl-2-trichloromethyl-zl ² -imidazoline

7.5 g of ethyleneimine and 17.0 g of triethylamine are dissolved in 250 ml of benzene. In the obtained solution is a solution of 17.3 g of N-dichlorothiophosphoryliminotrichloroacetic acid chloride in 200 ml of benzene introduced dropwise with constant stirring, a temperature of 0 to 5 ° C is maintained.

After all of the above-mentioned solution has been added, the reaction mixture is kept at a temperature of 25 ° C. for 7 hours. The resulting precipitate of triethylammonium chloride is filtered off and the solution of the end product in benzene is evaporated in vacuo until the solvent has been completely removed. The residue obtained is a pale yellow clear liquid from which the end product crystallizes. To increase the purity of the end product, it is recrystallized from a mixture of hexane and acetone (1: 5). 5.6 g corresponding to 31% of theory of 1-di- (äthyIenimido) -thiophosphoryl-2-trichloromethyl-id ^J -imidazoline with a melting point of 137-138 ° C. with decomposition are obtained.

C ₈ H ₁₂ CI ₃ N ₄ PS:
Found:

N 1637,16,54; S9,71,967;

25 Calculated:

N 16.82, S 9.62 weight percent.

Example 2

The preparation of the l-di- (ethyleneimido) -thiophosphoryl-2-trichloromethyl-z4 ² -imidazoline is carried out analogously to Example 1, but toluene is used as the inert organic solvent.

Yield of the end product 5.06 g, corresponding to 28% of theory.

Example 3

l-Di-iälhylenimidoJ-thiophosphoryl ^ -trichlormelhyM ² Mmidazoline is prepared as in Example 1, with diethyl ether being used as the inert organic solvent.

Yield of the end product 4.7 g, corresponding to 26% of theory.

Bcispi? ' i

The preparation of l-di- (äthylenimido) -thiophosphoryl-2-tnchlormethyM ² -imidazolins carried out as described in Example 1, except that carbon tetrachloride is used as an unreactive organic solvent. Yield of the end product 4.3 g, corresponding to 24% of theory.

130 246/521

Claims (1)

Palent claims: 1. l-Di ^ äthylenimidoJ-thiophosphoryl ^ -trichloromethyl-imidazoline the form! I. CH ₂ Cl ₃ C-C-N-P 10