SE448543B - / 3H / -IMIDAZO / 5,1-D / -1,2,3,5-TETRAZINE-4 SUBSTANCES, PROCEDURE FOR THEIR PRODUCTION AND PHARMACEUTICAL COMPOSITIONS CONTAINING THEM - Google Patents

Description

lll b! J 3 40 443 54-3 clays 3 to 8, preferably 6 carbon atoms.

Preferred tetrazine derivatives of the general formula I are those in which R 1 represents a straight-chain or branched alkyl group containing 1 to 6 carbon atoms, optionally substituted by one or two halogen atoms (preferably chlorine, fluorine or bromine) or with an alkoxy group containing 1 to 4 carbon atoms, preferably methoxy) or with a phenyl group (optionally substituted with one or two alkoxy groups containing from 1 to 4 carbon atoms, preferably methoxy), or R 1 represents an alkenyl group containing 2 to 6 carbon atoms (preferably allyl) or a cyclohexyl group. Particularly preferred tetrazine derivatives are those of the general formula I, wherein R 1 represents a straight or branched alkyl group containing from 1 to 6 carbon atoms and in particular from 1 to 3 carbon atoms, unsubstituted or substituted by a halogen atom, preferably chlorine or fluorine. R 1 in particular represents a methyl or 2-haloalkyl, for example 2-fluoroethyl or preferably 2-chloroethyl, group.

R 2 preferably represents a carbamoyl group or a monoalkylcarbamoyl group, for example methylcarbamoyl, or monoalkenylcarbamoyl group.

The present invention also includes alkali metal, for example sodium, salts of the compounds of general formula I, wherein R 1 represents a hydrogen atom and R 2 has the meaning given above, and whenever the context permits is meant by reference to the compounds of general formula I in the present preparation to include said salts. The salts are especially useful as intermediates.

According to one aspect of the present invention, the compounds of general formula I, wherein R has the above indicated are other meaning and R 1 than hydrogen, were prepared by reacting a compound 2 of the general formula: II 'R (wherein R 2 is above with ethisocyanate of the general formula: wherein R 3 represents an alkyl, alkenyl or alkynyl group containing up to 6 carbon atoms, each such group being optionally substituted with one to three substituents selected among halogen atoms, alkoxy, alkylthio, alkylsulfinyl and alkylsulfonyl groups and phenyl groups which are unsubstituted or substituted by one or more groups selected from alkoxy and alkyl groups containing up to 4 carbon atoms, and nitro groups or representing a cycloalkyl group containing from 3 to 8 carbon atoms. The reaction can be carried out in the absence or a chlorinated alkane, for example dichloromethane, or ethyl acetate, acetonitrile, Q-methylpyrrolid-2-one or preferably hexamethylphosphoramide, at a temperature of between 00 and 70 ° C, for example at ambient temperature. The reaction can continue for up to 30 days. Light should preferably be excluded from the reaction mixture.

According to a further aspect of the present invention there are prepared the compounds of general formula I, wherein R is as defined above and R1 is other than hydrogen, by reacting a compound of general formula: R2 N rr Nh / N lin YO 7 " IV (wherein R 2 is as defined above) or an alkali metal salt, for example sodium, thereof, having a compound of the general formula: R 5 x wherein R 3 is as defined above and X represents the acid residue of a reactive ester, for example a halogen atom (for example chlorine) or a sulfuric acid or sulfonic acid ester residue, for example a methoxysulfonyloxy, methanesulfonyloxy or toluene-p-sulfonyloxy group When RS meln V represents a haloalkyl, haloalkenyl or haloalkynyl group the acid residue of a reactive ester is represented by X selected from those known to be no less reactive in a compound of the general formula V than the halogen atom substituent in R general formula V represents a halogen When X in a compound of the preferably an alkali metal salt of the compound of general formula IV, and when X in a compound of the general formula V represents a halogen atom and R 3 is a haloalkyl, haloalkenyl or haloalkynyl group , wherein the halogen atom is the same as that represented by X, an excess of the dihalo compound of the general formula V is preferably used. The reaction of a compound of the general formula IV or alkali metal salt thereof with a compound of the general formula V , wherein R 3 and X are as defined above, may be carried out in a suitable anhydrous inert organic solvent, for example dichloromethane, acetonitrile or Q-methylpyrrolid-2-one or mixtures thereof, at a temperature of from 0 ° C to 120 ° C , and when a compound of general formula IV is used in the presence of an acid-binding agent, for example an alkali metal carbonate or bicarbonate, for example of sodium or potassium.

As a further aspect of the invention there are prepared compounds of general formula I wherein R1 represents a hydrogen atom and R2 has the meaning given above by reacting a compound of general formula II with a compound of general formula: Wherein R 4 represents an alkali metal atom (for example sodium) or a protecting group, such as a benzyl or p-methoxybenzyl group, followed by when R 4 represents a protecting group replacing the protecting group with a hydrogen atom in the compound thus obtained of the general formula: Rz / Äï> F'N => NN | v11 \ / N N \ Rs o. Wherein R is as defined above and R 5 represents a protecting group, such as a benzyl or p-methoxybenzyl group by IU fe.

LH 40 448 5433 methods known per se. Reaction of a compound of general formula II with a compound of general formula VI, wherein R 4 represents a protecting group, may be carried out as previously described in connection with reaction of a compound of general formula II with a compound of the general formula III. Reaction of a compound of general formula II with a compound of general formula VI, wherein R 4 is suitably inert organic solvent, for example ethanol, acetonitrile or Q-methylpyrrolidone, optionally in the presence of an acid at a temperature of from 0 to 10 ° C . The group R5 in compounds of general formula VII, wherein R 5 has above represents an alkali metal atom, can be obtained in a given sense, can be replaced by a hydrogen atom by methods known per se for forming a compound of general formula IV.

Compounds of general formula II may be prepared by the application or adaptation of methods known per se, for example procedures described by Shealy Y.F., Struck R.F., Holum L.B. and hbntgomeryxJ.A., J. Org.Chem. (1961), gâ, 2396.

Compounds of general formula III, V and VI may be prepared by the application or application of methods known per se.

By the term "methods known per se" as used in the present invention is meant methods which have hitherto been used or described in the literature.

The new tetrazine derivatives of the general formula I possess valuable antineoplastic activity, for example against carcinoma, melanoma, sarcoma, lymphoma and leukemias. They have been shown to be particularly active in mice at daily doses of between 0.5 and 16 mg / kg body weight administered intraperitoneally to TLX5 (S) lymphoma according to the procedure of Gescher et al, Biochem.

Pharmacol. (1981), åg, 89, and ADJ / PC6A and M5076 (reticulum cell sarcoma). For leukemia L1210 inoculated intraperitoneally, intracerebrally and intravenously and P388 according to the procedure described in "Methods of Development of New Anticancer Drugs" (NCI Monograph 45, March 1977. pp. L47 ~ l49, National Cancer Institute, Bethesda, United States) was the compounds are active both intraperitoneally and orally at doses of between 2.5 and mg / kg animal body weight. Inhibition of both primary tumor and: (1 40 448 545 metastasis was obtained against Lewis lung carcinoma at equal doses. Against B16 melanoma and C38 smoker in mice (NC1 Monograph 45, 32 gig.) The compounds were active interperitoneally at doses of between 6.25 and 25 mg / kg animal body weight.

The tetrazine derivatives also possess valuable immunodulatory activity and are useful in the treatment of organ and skin transplantation and in the treatment of immunological diseases. Essential individual compounds of general formula I include the following: 8-carbamoyl-3-methyl- [ 3] Imidazo [3,1d] -1,2,3,5-tetrazin-4-one A 8-carbamoyl-3-n-propyl-11β-imidazole, 1-d] -1,2,3 , 5-tetrazin-4-one 7 B-carbamoylozycretchlorethyl) - [μg] -imidazo [3,1-a] -1,2,3,3, S-tetrazin-4-one, C 3- (Z- chloroethyl) -8-methylcarbamoyl- [3§] -imidazo- [S, 1-d] -1,2,3,5-tetrazin-4-one D 8 -carbamoyl-3- (3-chloropropyl) - [3 §] -imidazin- [B, 1-di-1,2,3,5-tetrazin-4-one E 8-carbamoyl-3- (2,3-dichloropropyl) - [3,7-imidazo [5,1-4 ] -1,2,5,5-tetrazin-4-one F 3-allyl-8-carbamoyl- [3§] -imidazo [1,1] -1,2,3,5-tetrazin-4-one G 3- (2-chloroethyl) -8-dimethylcarbamoyl- [6-imidazophosphinyl] -β, sβ-terrazin-fl-one H 3- (Z-bromoethyl) -8-carbamoyl- [3§] -imidazo- [S, 10 ] -1,2,3,5-tetrazin-4-one in 3-benzyl-8-carbamoyl- [3§] -imidazo [5,1-d] -1,2,3,5-tetrazin-4-one J 8 -carbamoyl-3- (2-methoxyethyl) - [3fl] -imidazo-13,1-d] -1,2,3,5-tetrazin-4-one K 8 -carbamoyl-3-cyclohexyl- [3§] -imidazo [3,1-d] -1,1, Z, 3, S-tetrazin-4-one and L 8-carbamoyl-3- (2-methoxybenzyl) - [3§] -imidazo- (3,1-d ] -1,2,3,5-tetrazin-4-one M Compounds A and D and in particular C are of particular importance.

ZÛ 40 7 of 448 543 The letters A tí1l.M are assigned to the compounds to facilitate reference thereto later in the presentation.

The following examples illustrate the preparation of compounds of the general formula I according to the present invention and the reference example thereafter illustrates the preparation of intermediates.

EXAMPLE ~ 1 Compound A g 4 [E] -diazoimidazole-511] -carboxamide (500 mg) was suspended in methyl isocyanate (3.0 ml) and stirred in the dark at ambient temperature for 21 days. The reaction mixture was then diluted with anhydrous diethyl ether and filtered. The residue was washed quickly with anhydrous methanol, then with anhydrous diethyl ether and dried in air in the dark at ambient temperature to give 8-carbamoyl-3-methyl- [Bg] -imidazo [3,1-d] -1,2,3 , 5-tetrazin-4-one in the form of a light brown microcrystalline solid (198 mg), m.p. 20 ° C (with effervescence and darkening from 160 °: iii 20 ° C). - Elemental analysis obtained C, 36.8; H, 3.10; 44.2%; CÖHÖNÖOZ corresponds to: C, 37, 1; H, §09; N, 43, sa7; EXAMPLE 2 'Compound B 4 [S] -diazoimidazole-S [α] -carboxamide (300 mg) was suspended in anhydrous dichloromethane (10 ml) and treated with an excess of n-propyl isocyanate. The reaction mixture was then stirred in the dark at ambient temperature for 30 days. The reaction mixture was then filtered and the residue was washed quickly with anhydrous diethyl ether and dried in air in the dark at ambient temperature to give 8-carbamoyl-5-n-propyl- [3 §7-imidazo [B, 1-d] -1.2 , 3,5-tetrazin-4-one (102 mg), as a pale pink powder, m.p. 167 ° C (with effervescence) - [Elemental analysis obtained C, 43.4; H, 4.57; N, 38.0%; C 8 H 10 N 6 O 2 corresponds to: C, 43.2; H, 4.53, N, 37.8a]. Example 3 Compound C 4 []] diazoimidazole-5 [α] -carboxamide (300 mg) was suspended in anhydrous dichloromethane (10 ml) and Z-chloroethyl isocyanate (1.0 ml) was added. at ambient temperature for 30 days.

The reaction mixture was then stirred in the dark in The cream-colored suspension thus obtained was filtered and the residue was washed quickly with anhydrous diethyl ether and dried in air in the dark to give 8-carbamoyl-3- (2-chloroethyl) - [3§] -imidazo [É , ld] -1,2,3,5-tetrazin-4-one (483 mg) in the form of a cream powder, m.p. ls8 ° C (with strong decomposition). [Elemental analysis: found: C, 34.7; H, 3.0l; N, 34.9%; C 7 H 7 ClN 6 O 2 corresponds to: C, 34.7; H, 2.91; N, 34.7a7.

Repeating the above procedure has also given 8-carbamoyl-3- (Z-chloroethyl) - [[fl] -imidazo [B, ld] -1, Z, 3,5-tetrazin-4-one in another polymorphic form , m.p. 164-165 ° C (during decomposition).

EXAMPLE 4 Compound A A suspension of 413] -diamidoazaP5 [α] -carboxamide (1.37 g) in ethyl acetate (20 ml) was treated with methyl isocyanate (7.0 g) and stirred in a closed vessel in the dark at room temperature for 3 weeks. . The solid formed was filtered off and washed with diethyl ether to give 8-carbamoyl-3-methyl '[3fl] -imidazo [E, 1-d] -1,2,3,5-tetrazin-4-one (1 .9 g) in the form of a cream solid, m.p. (with effervescence).

This material was recrystallized from three different solvent systems to give three different products, each of which had a slightly different IR spectrum. in fact, all polymorphs of 8-carbamoyl-3-methyl- [QQI-imidazo [ε, 1-Q] -1,2,3,5-tetrazin-4-one. 7 (i) Colorless needles were obtained from a 3: 1 v / v mixture of acetone and water, Q max 3410, 3205, 1758, 1730 and 1678 cm -1, m.p. Z12 ° C (with effervescence). (ii) White microcrystals were obtained from a 1: 3 v / v mixture 3430, 3200, 1740 and 1615 cm * The three products were other acetone and water, Qnmx m.p. 200 ° C (with effervescence). (iii) A granular solid was obtained from hot water, ianax 3450, 3380, 3200, 1742, 1688 and 1640 cm -1, m.p. 215 ° C (with effervescence) (darkening from 200 ° C).

A suspension of 413] -diazoimidazole-S14] -carboxamide (1.3 g) in acetonitrile (20 ml) was treated with n-propyl isocyanate (6.5 g) and stirred in a closed vessel in darkness at room temperature for three weeks. The resulting pink solid was filtered off, washed with diethyl ether and recrystallized from a mixture of water and acetone (1: 4 v / v) to give 8-carbamoyl-3-n-propyl- [3 fi] -imidazo [3 , ld] -1,2,3,3-tetrazin-4-one (1.6 g), m.p. 170-177 ° C (with effervescence). By concentrating the mother liquor from the recrystallization, an additional quantity (0.2 g) of the same product was obtained.

EXAMPLE 6 Compound C A suspension of 4 [S] -diazoimidazole-514] -carboxamide (1.0 g) in ethyl acetate (30 ml) was treated with 2-chloroethyl isocyanate (3.3 ml) and the mixture was stirred in the dark at ambient temperature for 6 hours. day. The reaction mixture was then diluted with diethyl ether, and the resulting solid was filtered off to give s-carbamoyl-z-z-chloroethyl) - [s_n7-imidazo [s, 1-d_7-1,2,3, s-tetrazine- 4-one (1.6 g) as a colorless solid, m.p. 164-165 ° C (during decomposition). Basic analysis found: C, 34.5; H, 2.88; N, 34.5; Cl, 14.6%; C7H7ClN6O2 corresponds to: c, z4.6s; H, z, 91; N, 34.6s; c1.14.61% ._ 7.

EXAMPLE 7 Compound C A suspension of 4 [Q7-diazoimidazole-5 [47-carboxamide (5.0 g) in a mixture of dichloromethane (158 ml) and §-methylpyrrolid-2-one (8.3 ml) was treated with 2-chloroethyl isocyanate (16.7 ml), and the mixture was stirred in the dark at ambient temperature for 14 days. The reaction mixture was then diluted with anhydrous diethyl ether, and the resulting solid was filtered off and washed with diethyl ether to give 8-carbamoyl-3- (2-chloroethyl) - [Bg] -imidazo [S, ld] -1,2,3 , 5-tetrazin-4-one (6.3 g), as a purple solid, m.p. 164-165 ° C (during decomposition). [Elemental analysis: found: C, 34.7; H, 2.95; N, 34.5; C1, 14.4%; C 7 H 7 ClN 6 O 2 corresponds to: C, 34.65; 40 w 10 448 543 H, 2.91; N, 34.65; C1, 14.6l7. EXAMPLE 8 Compound C A suspension of 4 [S] -diazoimidazole-5β-carboxamide (145 g) in ethyl acetate (2175 ml) was treated with 2-chloroethyl isocyanate [478.5 ml) and stirred at SODC for two days. . The mixture was then filtered to give 8-carbamoyl-3- (2-chloroethyl) - [3§7-imidazo [B, 1-d] -1,2,3,5-tetrazin-4-one (250 g). in the form of a peach-colored phase: substance, m.p. 166 ° C. IN EXAMPLE 9 Compound A A stirred suspension of 413] -diazoimidazole-5 / 4β-carboxamide (2.2 g) in a mixture of dichloromethane (70 ml] and §-methylpyrrolid-2-one (3 g). 5 ml) was treated with methyl isocyanate (7.0 ml) and stirred at ambient temperature for 4 weeks The mixture was diluted with diethyl ether and the resulting solid was filtered off to give 8-carbamoyl-3-methyl- [3§] -imidazo- [B, 1 ~ d] -1,2,3,5-tetrazin-4-one (2.38 g) in the form of a pale purple solid, mp 202-20306 (with decomposition). Found: C, 36.8; H, 2.94; N, 43.1%; C 6 H 6 N 6 O 2 corresponds to: C, 37, 11; H, 3, 14; N, 43.3%]. 8-Carbamoyl-3-methyl- [QQI-imidazoles, 1- d] -1,2,3,5-tetrazin-4-one was obtained by dissolving the same in acetonitrile, filtering, concentrating the filtrate in dryness and decomposition of the residue obtained with diethyl ether.

This material is in the form of an orange-stained solid, m.p. about 200 ° C (during decomposition). Additional analysis C, 37.4, H, 3, Z6; N, 43, S1]. Its NMR spectrum in dimethylsulfoxide-D6 was identical to that of the above-mentioned pale pupar solid, but its IR spectrum (KBr disc) showed some differences.

EXAMPLE 10 - Compound D * A stirred solution of sodium nitrite (0.64 g) in water (4.6 ml) was cooled to 50-10 ° C and treated dropwise at this temperature with a solution of 5-amino-4- methylcarbamoylimidazole (1.00 g) g_________ ___ ___, _ ..... ~ ._...._,., ,, ....-_._ i. .._, _. 44 '548 543 in an aqueous solution of acetic acid (1M; 14.3 ml) for 5 minutes.

Stirring was maintained at So-10 ° C for 5 minutes. The dark red solution was extracted with ethyl acetate (4 x 35 ml) and the combined extracts were dried over magnesium sulphate. The resulting solution contained crude 4 [1] -diazo-5 []] -methylcarbamoylimidazole, which was unstable and immediately was used for the next step without further purification.

The solution of 4 [5] -diazo-SÅÄJ-methylcarbamoylimidazole in ethyl acetate prepared as described above was treated with 2-chloroethyl isocyanate (4.3 ml) and allowed to stand in the dark for 1 day.

The solution was then evaporated at 40 ° C / 10 mm Hg and the residue was triturated with petroleum ether (b.p. 400-60 ° C) to give an orange gum (4.23 g). This gum was then treated with ethyl acetate (50 ml) and filtered, and the filtrate was evaporated at 40 ° C / 10 mm Hg to give an organ gum (2.94 g). average pressure on silica gel eluting with a mixture of ethyl acetate and acetonitrile (4: 1 v / v) to give 3- (2-chloroethyl) -8-methylcarbamoyl- [BH] -imidazo [5,10] -1, 2,3,5-tetrazin-4-one (0.8l g) as a purple solid, m.p. 120 DEG-120 DEG C. (during decomposition). Application Analysis Found: C, 37.3; H, 3.5E; N, 3.1.9%; C 8 H 9 ClN 6 O 2 corresponds to: C, 37.4, H, 3.53; N, 32.7a]. This gum was purified by column chromatography under EXAMPLE 11 Compound E A suspension of 4 [&] - diazoimidazole-51%] - carboxamide (1.0 g) in ethyl acetate (SO ml; dried over anhydrous potassium carbonate) was treated with 3-chloropropyl isocyanate (4 , 86 g), and the mixture was stirred at ambient temperature for 3 days. The reaction mixture was then diluted with anhydrous diethyl ether, and the solid formed was filtered off and washed with anhydrous diethyl ether to give 8-carbamoyl-3- (3-chloropropyl) - [3H] -imidazo [5,1-di -1,3,3,5-S-tetrazin-4-one (1.05 g) as a pink solid, m.p. 153-154 ° C (with decomposition). [Elemental analysis: found: C, 37.1; H, 3.42; N, 32.7; Cl, 13.8%; C H ClN 0 8 9 6 2 corresponds to: C, 37.4; H, 3.53; N, 32.8; Cl, 13.8å]. EXAMPLE 12 Compound F By proceeding in the same manner as previously described in Example 11 but substituting the 3-chloropropylisocyanate used as a starting material for the appropriate quantity of 2,3-dichloropropylisophanate, 8-carbamoyl-3- (2,3 -dichloropropyl- [3§] - imidazo [5,1d] -1,2,3,5-tetrazin-4-one in the form of a off-white solid, mp 153-155 ° C (with decomposition). Analysis: Found: C, 32.7; H, Z, 51; N, Z8.7; Cl, 24.1%; C8 H8 Cl2 N6 O2 corresponds to: C, 33.0; H, 2.77; N, 28.9 ; Cl, 24.4a].

EXAMPLE 13 Compound G Stirred allyl isocyanate (4.5 ml, redistilled immediately before use) was treated with 415] -diazoimidazole-5 [Å] -carboxamide (1.0 g) and then with hexamethylphosphoramide (20 ml).

The mixture was stirred at ambient temperature in the dark for 18 hours, after which it was diluted with anhydrous diethyl ether and filtered. The resulting colorless solid was washed with anhydrous diethyls: iii to give s-allyl-s-carbamyl-β-imidazo [5,11d] -1,2,3,5-tetrazin-4-one (1.6 g) in the form of a colored solid, m.p. 149-150 ° C. [\) max (KBrgaisc): 1730, 1615 cm_1; NMR in DMSO-die; singlets at 8.75, 7.67 and 7.606: double-double triplet at 6.026 (J = 5.5, 8, 10Hz), double-double at 5.356 (J = 1.5, 8 Hz) and 5.20 6 (J = 1.5, 10Hz) and doubled at 4.866 (J = 5.5)].

EXAMPLE 14 Compound H A solution of 415] -diazo-5 [β] -dimethylcarbamoylimidazole (1.59 g; prepared as described in Reference Example 1 below) in dry ethyl acetate (57 ml) was treated with 2-chloroethyl isocyanate (6.36 g) and stirred at room temperature in the dark for 24 hours.

The solution was then evaporated in vacuo at 35 ° C, finally at 0.1 mm Hg to remove excess 2-chloroethyl isocyanate.

The residual liquid was then purified by column chromatography at medium pressure on silica gel eluting with a mixture of ethyl acetate and acetonitrile (4: 1 v / v, to give 3- (2-chloroethyl) -8-dimethylcarbamoyl] β-imidazo [S, ld] - 1,2,3,5-tetra-zin-4-one (0.82 g) in the form of colorless crystals, mp 114,166 ° C. [Blementar analysis found: C, 39.7; H, 3 , 95; N, 30.8%; CQHIICINÖOZ corresponds to: C, 39.9; H, 4, 10; N, 31, 0%].

EXAMPLE 15 Compound I A stirred suspension of 4 [[alpha] - diazoimidazole] -SAZI-carboxamide (1.0 g) in hexamethylphosphoramide (4 ml) was treated with 2-bromoethyl isocyanate (4.5 ml) and the mixture was stirred in the dark at ambient temperature. for 2 days. The reaction mixture was then diluted with anhydrous diethyl ether, and the solid formed was filtered off and washed with anhydrous diethyl ether to give 3- (2-bromoethyl) -8-carbamoyl- [3 3] -imidazo [5,1d] -1,2, 3,5-tetrazin-4-one (1,17 g) as a colorless solid, m.p. 156-1S7 ° C (with decomp.). [Elemental analysis: found: C, 29.5; N, 2.36; N, 29.1; Br, 27.3%; C 7 H 7 BrN 6 O 2 corresponds to: C, 29.3; H, 2.46; N, 29.3; Br, 27.8å]. EXAMPLE 16 By proceeding in the same manner as described above in Example 15 but substituting the 2-bromomethyl isocyanate used as starting material for the appropriate quantity of benzyl isocyanate, 3-benzyl-8-carbamoyl- [3§] -imidazo- [5 , 1-d] -1,2,3,5-tetrazin-4-one (0.83 g] as a skin-colored solid, mp 176-177 ° C (with decomposition). [Elemental analysis found: C , 53.6; H, 3.66; IN, 31.0%; CIZHIONO2Z corresponds to: C, 53.3; H, 3.73; N, 3.1, 11].

EXAMPLE 17 Compound K A suspension of 4 [S] -diazoimidazole-511] -carboxamide (0.3 g) in acetonitrile (5 ml) was treated with 2-methoxyethyl isocyanate (0.5 g) and the mixture was stirred at between 450 and 47 ° C for darkness for 24 hours. The solid formed was filtered off and washed with diethyl ether to give crude β-carbamoyl-3- (2-methoxyethyl) - [3] imidazo [3,1d] -1,1, 3,5-tetrazine-4- on (o, 45 g), m.p. 145-147 ° C (with decomposition).

The product was purified by recrystallization from water-acetone to give pink rosettes or from water-dimethyl sulfoxide to give colorless needles, m.p. 164-165 ° C (with decomposition). [Elemental Analysis C, 40.4; H, 4.20; N, 35.2%; C 8 H 10 N 6 O 3 corresponds to: C, 40.34; H, 4.0; N, 35.2%].

EXAMPLE 18 Compound LZ A suspension of 4 [E] -diazoimidazole-5 [1] -carboxamide (0.3O g) in acetonitrile (10 ml) was treated with cyclohexyl isocynate (1.0 g) and the mixture was stirred at 60 ° C in the dark. for 3 days. The solid formed was filtered off and washed with a mixture of ethanol and 0.880 aqueous ammonia (100: 0, S v / v; 20 ml) for one minute to give 8-carbamoyl-3-cyclohexyl- [3 fi] - imidazo [S, ld] -1,2,3,5-tetrazin-4-one (0.015 g), m.p. 196 ° C (with effervescence). EXAMPLE 19 Compound J A suspension of 4 [5] -diazoimidazole-S13] -carboxamide (0.4 g) in acetonitrile (10 ml) was treated with benzyl isocyanate (0.6 g), and the mixture was stirred at 60 ° C in the dark. overnight. The reaction mixture was then cooled and filtered to give 3-benzyl-8-carbamoyl- [Bg] -imidazo [5,1d] -1,2,3,5-tetrazin-4-one (0.75 g) as of a pale pink solid, m.p. 187 ~ 188 ° C (with effervescence).

EXAMPLE 20 Compound M A suspension of 413] -diazoimidazole-511] -carboxamide (0.1 g) and R-methoxybenzyl isocyanate (0.4 g) in acetonitrile (5 ml) was stirred at 60 ° C in the dark for 4 hours. . The pale pink substance formed was filtered off and washed repeatedly with cold diethyl ether to give 8-carbamoyl-3- (R-methoxybenzyl) - [3§] -imidazo [ε, 1-Q] -1,2,3,5 -tetrazin-4-one (0.23 g), m.p. 180-182 ° C (with effervescence).

EXAMPLE 21 By proceeding in the same manner as in the above example, 8- (N-allylcarbamoyl) -3- (2-chloroethyl) - (SE) -imidazo (s, 1-a) -1, z, s, s- tetrazin-4-one (LR. 1150 mfl; mm) in nMso- (IÖ:: zu 40 448 545 multiplets 3.96, 5.06 and 5.84 ppm; triplets 4.60 and 6.2 vPm = via 5 -diazo-4-allyl-carbamoylimidazole, singlet 8.78 ppm: from 5-amino-4-allylcarbamoyl-imidazole -amino-4-allylcarbamoylimidazole was prepared from 5-nitro-4-allylcarbamoylimidazole, mp 218-2-2 ° C) by reduction with titanium trichloride.

REFERENCE EXAMPLE (1) An intimate mixture of 5-nitroimidazole-4-carboxylic acid (2.0 g) and phosphorus pentachloride (2.67 g) was stirred and heated on an oil bath at 100 ° C for 1 hour. The resulting yellow slurry was evaporated at 60 ° C / 0.1 mm Hg for 30 minutes to give 1,6-dinitro-5§, 10§-diimidazofl, 5-a: 1 ', 5'-d] pyrazine - -5,10-dione (1.90 g) as a yellow solid, m.p. 249-25l ° C (during decomposition). ¿"? Max (Wu.

Windaus, Ber., 1923, §§, 1622e describe using the same method their products (KB disc) 1750 cm'1; m / e 278 684 and Gireva, Chem. Abs. yoke, such as "S-nitroimidazole-4-carbonyl chloride". (ii) An aqueous solution of dimethylamine (25% w / v; 60 ml) was cooled to between 0 ° and 5 ° C and treated portionwise with stirring with 1,6-dinitro-Sh, 10§-diimidazo [1,5- Axl ', 5'-d] pyrazine-5,10-dione (6.0 g] within this temperature range. The deep purple solution was stirred for 2 hours, the residue was evaporated at 50 ° C / 10 mm Hg and then acidified by treatment with concentrated hydrochloric acid to give an orange solution.

This solution was extracted with ethyl acetate (7 x 200 ml) and the combined extracts were dried over magnesium sulphate and evaporated to give a yellow solid (6.6 g).

This solid was decomposed with toluene (50 ml) and then recrystallized from ethyl acetate to give 51%] -nitro-4 [3] -dimethylcarbamoylimidazole (2.53 g) as yellow crystals; m.p. 193-195 ° C. [Elemental analysis found: c, 3s, 9; H, 4.2s; N, so, 4%; N, 3o, 4% _7.

C 6 H 8 N 4 O 3 corresponds to: C, 39.1; H, 4.38; (iii) A solution of 5 [N] -nitro-4 [S] -dimethylcarbamoylimidazole (1.62 g) in dry dimethylformamide (32 ml) was treated with platinum oxide (0.32 g) and shaken under hydrogen at atmospheric pressure. pressure and room temperature. complete (710 ml).

After 3 hours, the hydrogen absorbent mixture was treated with charcoal and filtered through diatomaceous earth. The dark brown filtrate was evaporated at SOOC / 0.1 mm Hg and the residue formed was triturated with diethyl ether to give crude 5 [Å] -amino-413] -dimethylcarbamoylimidazole (1.75 g) as a dark brown christian solid. m.p. 119-1s1 ° c [Qmaxuosr disc) 1595 mfl; NMR in DMSO-die: singlets at 3.2 and 7JD5], which was still contaminated with colloidal platinum and used in the next step without further purification. s (iv) A stirred solution of sodium nitrite (0.79 g) in water (5.7 ml) was cooled to between So and 10 ° C and treated dropwise within this temperature range with a solution of S [Å] -amino- -SZAI-dimethylcarbamoylimidazole (1.75 g) in water-acetic acid (1M; 17.6 ml) for 5 minutes. The resulting solution was extracted with ethyl acetate (4 x 40 ml), the combined extracts dried over magnesium sulphate and evaporated at 3000/10 mm Hg to give 413] -diazo-S11] -dimethylcarbamoylimidazole (1.59 g) as orange crystals, m.p. 10-110 ° C (with decomposition) / Elemental analysis found: C, 42.6; H, 4.17; N, 41, 4%; C6H7N5O corresponds to: C, 43¿6; H, 4.27; N, 42.4a].

Standard tests with compounds A, I, C, D and H of the present invention against TLXS lymphoma implanted in mice gave the results shown in Table I.

Standard tests with compound C of the present invention against leukemia tumors of type L1210 and P388 implanted in mice gave the results shown in Table II.

* Cl TABLE I 17 44.8 543.

Activity of the associations according to Swedish patent application no. 8204817-4 against TLX5 lymphoma implanted in mice.

Compound Optimal dose mg / kg administered 3 days after implantation Survival time, compared with controls Survival time in days * 160 157% 160 137% 40 All S-treated mice survived the 60th day after implantation-1 60 40 All S treated mice survived the 60th day after implantation 60 40 A1 1a treated mice survived the 60th day after implantation 60 * * mice, which survived for 60 days, remained alive and were not further examined _. ..-......_................, ..., ..._..., _ q ._.._., ~ _. _-_._..___ .. l 448 543 18 TABLE II.

Activity of compound C according to Swedish patent application no. 8204817-4 gene against leukemic tumors implanted in mice.

Dose in Number Survival time Tumor type mg kg-1 day_1 dose days in days * L1210 40 1 60 S ÖÛ S 60 9 60 12.5 9 60 P338 40 1 60 T ÖÛ 5 60 5 ÖÛ * mice that survived for 60 days remained alive and were not examined further. v: s _; 40 moose? 446 545 The present invention includes within its scope pharmaceutical compositions which contain as active ingredient at least one tetraane derivative of the general formula I in addition to a pharmaceutical carrier or coating. In clinical practice, the compounds of general formula I are normally administered orally, rectally, vaginally or parenterally, for example intravenously or intraperitoneally.

Methods for formulating pharmaceutically active compounds are well known in the art and a suitable carrier can be determined by the physician or pharmacist depending on such factors as the desired effect, size, age, sex and condition of the patient and the properties of the active compound. The compositions may also contain in the usual manner such materials as solid or liquid diluents, wetting agents, preservatives, flavoring and coloring agents and the like.

Solid compositions for oral administration include compressed tablets, pills, dispersible powders and granules.

In such solid compositions, one or more of the active compounds is mixed with at least one inert diluent, such as calcium carbonate, potato starch, alginic acid or lactose. The compositions may also normally contain additional substances other than inert diluents, for example lubricants, such as magnesium stearate. Liquid compositions for oral administration include pharmaceutically acceptable emulsions, solutions, suspensions, syrups and elixirs containing inert customary diluents, such as water and liquid paraffin. In addition to inert diluents, such compositions may also contain adjuvants, such as wetting and suspending agents, for example, polyvinylpyrrolidone, and sweetening, flavoring, perfuming and preservative agents. The compositions of the invention for oral administration also comprise capsules of absorbable material, such as gelatin, containing one or more of the active substances and with or without the addition of diluents or excipients.

Solid compositions for vaginal administration include pessaries formulated in a manner known per se and containing one or more active compounds.

Solid compositions for rectal administration include suppositories designed in a manner known per se and containing one or more of the active compounds.

Formulations of the invention for parenteral administration include sterile anhydrous or anhydrous solutions, suspensions or emulsions. Examples of anhydrous solvents or suspending media are polyethylene glycol, dimethyl sulfoxide, vegetable oils such as olive oil, and injectable organic esters such as ethyl oleate. These compositions may also include adjuvants, such as preservatives, wetting agents, emulsifying agents and dispersing agents. They may be sterilized, for example by filtration through a bacterial filter, by the incorporation of sterilizing agents into the compositions or by irradiation.

They may also be prepared in the form of sterile solid compositions which can be dissolved in sterile water or any other sterile injectable medium immediately before use.

The percentage of active ingredient in the compositions according to the invention can be varied, it being necessary that it constitutes such a proportion that a suitable dose for the desired therapeutic effect is to be achieved. Obviously, several unit dosage forms can be administered at about the same time. In general, the formulations should normally contain at least 0.025% by weight of active substance when administered by injection; for oral administration, the preparation normally contains at least 0.1% by weight of active substance. The dose used depends on the desired therapeutic effect, the mode of administration and the duration of treatment.

The tetrazine derivatives of the general formula I are useful in the treatment of malignaneoplasms, for example carcinoma, malanoma, sarcoma, lymphoma and leukemia in doses which are usually between 0.1 and 200, preferably between 1 and 20 mg / kg body weight per day. . The following composition examples illustrate pharmaceutical compositions of the present invention.

COMPOSITION EXAMPLE 1 A solution suitable for parenteral administration was prepared by the following ingredients: 8-carbamoyl-3- (2-chloroethyl) - [3§] -imidazo [B, 1-d] -1,2,3,5-tetra-D zin-4-one 1.0 g Dimethylsulfoxide 10 ml Arachis oil and 90 ml of 40 ..... .._ ~> ......., -_- <~ | -...... ~ .. -.__ _. by dissolving 8-carbamoyl-3- (2-chloroethyl) - [3§] -imidazo- [3,1-d] -1,2,3,5-tetrazin-4-one in the dimethyl sulfoxide and addition of the arachis oil: The resulting solution was divided under aseptic conditions into ampoules in an amount of 10 ml per ampoule.

The vials were sealed to give 10 vials each containing 100 mg of 8-carbamoyl-3- (2-chloroethyl) - [3 fl] -imidazo [B, ld] -1, zl, s, s-cetrazin-4-one . Similar ampoules containing solutions suitable for parenteral administration can be prepared by the same procedure but substituting 8-carbamoyl-3- (2-chloroethyl) - [3 §] -imidazo- [B, ld] - 1,2 , The 3,5-tetrazin-4-one against another compound of general formula I.

COMPOSITION EXAMPLE 2 Capsules suitable for oral administration were prepared by placing 8-carbamoyl-3- (Z-chloroethyl) -Z 2 - imidazo [ε] -1,2,3,5-tetrazin-4-one in gelatin shells of size 2 in an amount of 10 mg per capsule.

Similar capsules can be prepared using another compound of general formula I or any other capsule shell of suitable size.

Claims (15)

4¿8 545 PArBNrKRAv 1. [ÉÉ] -Imidazo-15,1-d] -1,2,3,5-tetrazine-4-one derivative of the general formula; Wherein R 1 represents hydrogen, a straight or branched alkyl, alkenyl or alkynyl group containing up to 6 carbon atoms, each such group being unsubstituted or substituted with from a to three substituents selected from halogen atoms, straight or branched alkoxy, alkylthio, alkylsulfinyl and alkylsulfonyl groups containing up to 4 carbon atoms, and phenyl groups which are unsubstituted or substituted by one or more groups selected from alkoxy and alkyl groups containing up to 4 carbon atoms, and nitro groups, or R 1 represents a cycloalkyl group containing from 3 to 8 carbon atoms, and R 2 represents a carbamoyl group, which on the nitrogen atom may bear one or two groups selected from straight and branched alkyl and alkenyl groups each containing up to 4 carbon atoms, and cycloalkyl groups containing from 3 to 8 carbon atoms, and when R 1 represents hydrogen, alkali metal salts thereof. The tetrazine derivative according to claim 1, wherein R 1 represents an alkyl, alkenyl or alkynyl group substituted with one, two or three optionally substituted phenyl groups, the optional substituents on the phenyl group or groups being selected from alkoxy and alkyl groups containing up to 4 carbon atoms and the nitro group. A tetrazine derivative according to claim 1, wherein R 1 represents a straight or branched alkyl group containing 1 to 6 carbon atoms, optionally substituted with one or two halogen atoms or with an alkoxy group containing 1 to 4 carbon atoms or with a phenyl group optionally substituted with one or two alkoxy groups containing from 1 to 4 carbon atoms, or R 1 represents an alkenyl group containing 2 to 6 carbon atoms or a cyclohexyl The tetrazine derivative according to claim 3, wherein the halogen atom or -s are chlorine, fluorine and / or bromine, the alkoxy group or groups tZ3 is 444 meth45 and the alkenyl group is allyl.a The tetrazine derivative according to claim 1, wherein R 1 represents a straight or branched alkyl group containing 1 to 6 carbon atoms, unsubstituted or substituted by a halogen atom. The tetrazine derivative according to claim 1, wherein R 1 represents an alkyl group containing 1 to 3 carbon atoms, osubstituted or substituted by a halogen atom. The tetrazine derivative according to claim 1, wherein R 1 represents a methyl or 2-haloalkyl group. The tetrazine derivative according to claim 7, wherein the halogen atom of the alkyl group is chlorine or fluorine. The tetrazine derivative according to claim 1, wherein R 1 represents a 2-fluoroethyl or 2-chloroethyl group. The tetrazine derivative according to claim 1, wherein R 1 represents a benzyl or p-methoxybenzyl group. A tetrazine derivative according to any one of claims 1 to 10, 2 represents a carbamoyl group, a monoalkylcarbamoyl group wherein R group contains up to 4 carbon atoms in the alkyl radical or a monoalkenylcarbamoyl group containing up to 4 carbon atoms in the alkenyl radical. The tetrazine derivative according to claim 1, wherein R 1 represents a straight or branched alkyl, alkenyl or alkynyl group containing from 1 to 6 carbon atoms, wherein each such group is unsubstituted or substituted by from one to three halogen atoms and R 2 represents the carbamoyl group. 13. Process for the preparation of a derivative of the general formula: R2 \ _ “§N N | Wherein R represents vate, a straight or branched alkyl, alkenyl or alkynyl group containing up to 6 carbon atoms, each such group being unsubstituted or substituted by from one to three substituents selected from halogen atoms, straight 448 saa 24 or branched alkoxy, alkylthio, alkylsulfinyl and alkylsulfonyl groups containing up to 4 carbon atoms, and phenyl groups which are unsubstituted or substituted by one or more groups selected from alkoxy and alkyl groups containing up to 4 carbon atoms, and nitro groups, or R 1 represents a cycloalkyl group containing from 3 to 8 carbon atoms, and R 2 represents a carbamoyl group, which on the nitrogen atom may bear one or two groups selected from straight and branched alkyl E and alkenyl groups each containing up to 4 carbon atoms, and cycloalkyl groups containing from 3 to 8 carbon atoms, and when R 1 represents hydrogen, alkali metal salts thereof, which comprises (A) when R n general formula Iiej is a hydrogen atom, reacting a compound of general formula II (wherein R 2 has the meaning given above) with an isocyanate of the general formula: wherein R 3 represents a straight or branched alkyl, alkenyl or alkynyl group containing up to 6 carbon atoms, each such group being unsubstituted or substituted by one to three substituents selected from halogen atoms, straight or branched alkoxy, alkylthio, alkylsulfinyl and alkylsulfonyl groups containing up to 4 carbon atoms, and phenyl groups , which are unsubstituted or substituted by one or more groups selected from alkoxy and alkyl groups containing up to 4 carbon atoms, and nitro groups, or R 3 represents a cycloalkyl group containing 3 to 8 carbon atoms; (B) when R in the general formula I is not a hydrogen atom, reacting a compound of the general formula 25 'i44s 545 Rz _' N _ e \ än _ 'N' I lv N Kiwi / H \\ / ( wherein R 2 is as defined above) or an alkali metal salt thereof having a compound of the general formula: R 3x v. 3 e. wherein R is as defined above and X represents the acid residue of a reactive ester; or (C) - when R 1 in the derivative of the general formula I is hydrogen or an alkali metal salt thereof, reacting a compound of the general formula:. R2 NZ (5 new., _.- ._ i (wherein R2 has the meaning given above) with a compound of the general formula: 114Nco in vi - wherein R4 represents an alkali metal atom or a protecting group, such as a benzyl or Q-methoxybenzyl group , followed by when R 4 represents a protecting group exchange of the protecting group for a hydrogen atom in the compound of the general formula thus obtained: wherein R 2 has the meaning given above and R 5 represents a protecting group, such as a benzyl or β-methoxybenzyl group, by methods known per se A process according to claim 13 process (B), wherein X represents a halogen atom or a methoxysulfonyloxy, methanesulfonyloxy or toluene-p-sulfonyloxy group. Pharmaceutical compositions containing as active ingredient at least one tetrazine derivative of the general formula R 2 N crew N 1 \ \ / N \ fl / N \ R 1 wherein R 1 represents hydrogen, a straight or branched alkyl, alkenyl or alkynyl group containing up to 6 carbon atoms, each such group being unsubstituted or substituted by from one to three substituents selected from halogen atoms, straight or branched alkoxy, alkylthio, alkylsulfinyl and alkylsulfonyl groups containing up to 4 carbon atoms, and phenyl groups which are unsubstituted or substituted with one or more groups selected from alkoxy and alkyl groups containing up to 4 carbon atoms, and nitro groups, or R 1 represents a cycloalkyl group containing from 3 to 8 carbon atoms, and R 2 represents a carbamoyl group, which on the nitrogen atom may bear one or two groups selected from straight and branched chain alkyl and alkenyl groups each containing up to 4 carbon atoms, and cycloalkyl groups containing from 3 to 8 carbon atoms, and when R 1 represents hydrogen, alkali metal salts thereof, in combination with a pharmaceutical carrier or coating. “_ __.- Y _ 1.4. .,> ._....._...._....__..., _, __.._...-__.........__..._ ._..._.....