DK165181B - HETEROCYCLIC DISULFIDES PROCEDURE FOR THEIR PREPARATION AND PHARMACEUTICAL AGENTS CONTAINING THESE COMPOUNDS. - Google Patents

Description

DK 165181 B

The present invention relates to heterocyclic disulfides, to a process for their preparation and to pharmaceutical agents containing these compounds.

It is known that the protective mechanisms of the living organism, briefly referred to as humoral immunity and cellular immunity, work together to neutralize foreign bodies that can induce pathogenetic changes and be harmful, and to eliminate them, preferably microorganisms or neoplastic 10. cells.

Immunological studies show that there is a correlation between the decrease in immunological activity due to internal or external factors and the increase in infectious or tumor diseases. In addition, other diseases occur due to changes in immune system functions. These include, for example. autoimmune diseases or diseases caused by immune complexes. Therefore, for a long time have been searched for immune stimulants, ie. substances which are capable of altering the immunological activity of the recipient, preferably increase it, and which, because of their high efficiency and good compatibility, allow a wide application to support the body's defense forces. Examples thereof which have been tested for immunity stimulation are BCG and C. Parvum, also extracts of M. tuberculosis and of brucellosis.

However, at the concentrations in which they are used, these substances give clear side effects, such as e.g. local granulomas to varying degrees. Lack of knowledge of the exact nature of the drugs makes a systematic study with good reproducibility difficult for the clinical results. It is therefore desirable, in this context, to provide new immune stimulants that constitute. chemically defined substances and have low toxicity, such as e.g. "Bestatin", which is currently an intensively investigated low-molecular-weight immune stimulus and generally constitutes

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2 a scientific reference substance.

U.S. Patent No. 4,378,364 discloses certain bis (htero) disulfides whose heterocyclic ring may also be substituted by carboxyl groups. The administration of these 5 compounds to cancer patients after surgery should lead to an improvement in wellbeing, activity, appetite, weight gain and a reduction in pain. This is shown in various examples with 6,61-dithionicotinic acid (CPDS).

It has now surprisingly been found that the compounds of the present invention have high immunostimulatory and immune-enhancing effects such as this one, e.g. is expressed in the DTH response to sheep erythrocytes, in the activation of mononuclear phagocytes, and in marked CSF activity. These immunostimulatory effects may e.g. also be observed in. an increase in the resistance to infections. Furthermore, the compounds of the invention have surprisingly cytostatic effect, e.g. against B16 melanoma in mice.

Thus, the present invention relates to a class of immunopharmacologically and cytostatically effective substances which are chemically defined, have low toxicity and, per se or in combination with other active substances, constitute valuable drugs. The compounds of the invention have an LD 2 value of more than 1000 mg / kg by intravenous injection in mice. The effective immunomodulatory and cytostatic amount is in vertebrates, preferably warm-blooded mammals, in the range of from ca. 0.5 to approx. 100 mg / kg body weight by parenteral or oral administration, without causing toxic side effects, and thus 30 are very suitable for treating diseases of the immune system.

The invention thus relates to disulfides of the general formula I '

Het '-S-S-Het' (I ·) wherein Het 'means 1,3-thiazolyl which is substituted with C

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C1-6 alkyl, carboxy or C2-5 acylamino, where acyl means a residue of an aliphatic dicarboxylic acid, 1,2,4-triazolyl which is substituted by hydroxy and phenyl, 1,3,4-thiadiazolyl which is substituted by carboxylic acid C 1-4 alkylthio, dihydro-1,2,4-triazinyl substituted with C 1-6 alkyl, halo-C 2-4 alkenyl, oxo or hydroxy, or benzimidazolyl substituted with carboxy, Het ' is substituted by at least one directly or to 10 a substituent bonded carboxy group or a directly bonded hydroxy group, these groups also being in the form of a physiologically acceptable salt.

Considering that C 1-6 alkyl substituents, straight or branched alkyl groups have 1-6, preferably 1-3 carbon atoms, e.g. methyl, ethyl, n- or isopropyl, n- or tert-butyl, preferably methyl, which may be optionally substituted by carboxy.

As C2-5 acylamino groups, wherein acyl represents a residue of an aliphatic dicarboxylic acid, e.g. are mentioned 20 3-carboxy-propionyl and 4-carboxy-butyryl.

As C 1-4 alkylthio groups substituted with carboxy, e.g. are mentioned methylthio and ethylthio and as halogeno-C2-4 alkenyl groups e.g. vinyl or allyl, e.g. is substituted by chlorine or bromine.

The heterocydic group Het 'may be substituted one or more times, preferably 1 to 3 times. However, such compounds are preferred which carry in the heterocyclic ring or in the condensed carbocyclic ring 1 or 2 substituents. Of these substituents, at least 30 must have an acidic function. Thus, a substituent can either carry a carboxy group, or it can also, as a carboxy or hydroxy group, be directly linked to the heterocyclic group Het '.

According to the invention, such compounds are particularly preferred where Het · is a thiazole residue which is substituted by at least one carboxyl or one carboxymethyl group.

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Since the compounds of general formula 1 'carry acidic functions, especially a carboxy group, they may also be present in phonies of their physiologically acceptable salts, e.g. as alkali or alkaline earth metal salts, preferably Na, K, 5 Ca, Mg salts or e.g. ammonium salts or substituted ammonium salts, e.g. NH4 +, ethanolammonium,. diethanol ammonium, trialkylammonium, e.g. triethylammonium, tetracylammonium, salts with basic amino acids, e.g. lysine and arginine.

The invention further relates to a process for preparing the compounds of the invention of formula I ·.

The process according to the invention is characterized in that a compound of the general formula II 15

Het * -SH (II) wherein Het 'has the meaning given above, by reaction with an oxidizing agent is converted into the compounds of the invention.

As oxidizing agents, e.g. mention is made of oxygen, hydrogen peroxide, sometimes with the addition of iron salts, e.g. Mohr's salt or by the addition of bases, e.g. sodium hydroxide, potassium hydroxide, sodium hydrogen carbonate or potassium hydrogen carbonate, organic peracids such as e.g. peracetic acid, perbenzoic acid or m-chlorobenzoic acid, iodine or bromine in the form of element, sometimes with the addition of bases, such as e.g. sodium hydroxide or potassium hydroxide, FeCl 2 or K 2 tFeiCNjg].

As oxidizing agents, there may preferably be mentioned oxygen, hydrogen peroxide, organic peracids, e.g. peracetic acid, perbenzoic acid and m-chloroperbenzoic acid and iodine in elemental form. The oxidation is preferably carried out in water or an organic solvent, e.g. methanol, ethanol, isopropanol, acetic acid ester and halogenated hydrocarbons, e.g. dichloromethane and chloroform. ·

A mixture of these solvents may also be used.

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agents. When working with oxygen, hydrogen peroxide and peracids, the use of solvents which are known to form explosive peroxides, e.g. ether, tetrahydrofuran, dioxane, acetone, methyl ethyl ketone, isopropanol, etc. are avoided.

The compounds of the invention may also be prepared by a compound of the general formula III

Het'-X (III) wherein Het 'is as defined above and X is a reactive leaving group such as halogen, preferably chlorine or bromine, such as -SO 2 R, where R is preferably methyl, trifluoromethyl, phenyl, tolyl or naphthyl, Such as -OPO (OR'2) / where R 'is preferably phenyl, is reacted with Me2S2 where Me is an alkali metal, preferably sodium.

In addition, the compounds of the invention may be prepared by a compound of the general formula 20

Het'SSO2Me (IV) wherein Het 'and Me have the above meanings are reacted with iodine in an aqueous medium.

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The reaction can take place at a temperature between approx. -40 ° C and the boiling point of the solvent or solvent mixture, preferably between ca. -10 and approx.

+ 40 ° C.

The active substance may be administered alone or in combination with one or more, preferably another drug, which favorably affects infections, such as e.g. is caused by bacteria, fungi or viruses, as well as tinnor disorders. The active substances of the invention can be administered both parenterally and orally. For parenteral administration, solutions or suspensions of the active substance are used in a pharmaceutically acceptable carrier, preferably

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6 certain plant oil, such as peanut or sesame oil, as well as alcoholic solutions of the active substance, e.g. ethanol, pyropanediol or glycerol or in mixtures of the aforementioned solvents. For the preparation of aqueous solutions, the active substance is preferably used in the form of water-soluble, physiologically acceptable salts. The compositions may contain the usual adjuvants and carriers. These are e.g. fillers, emulsifiers, slides and buffers, and flavoring agents.

The invention therefore also relates to pharmaceutical agents which are characterized in that they contain a compound of the general formula I 'and which are useful for immune stimulation, immune reconstruction and cytostatic treatment.

Below, the effect of the compounds on the immune response of mice and their immune stimulatory effects by various in vivo standard methods will be elucidated, for example. As is well known, the various assay methods presented are particularly suitable for assessing 20 immunostimulants and their efficacy.

Experiment 1

Effect on Delayed Type Hyper-Sensitivity DTH Cellular Immunological Response

Groups of 5 NMRI female mice weighing 18-20 g of 6 9 receive either 10 or 10 red blood cells from sheep per intravenously. animals. Sheep erythrocytes are considered within the immunology of a standard test substance (antigen) to trigger cellular and humoral immune responses. In particular, this sample provides information on the function of T cell-dependent components (T helper cells) in the immune system. The test substance [bis- (5-carboxymethyl-4-methyl-thiazol-2-yl) disulfide] obtained according to Example 8 is administered twice daily intraperitoneally on days -3, -2, -1 and 0 at concentrations of 20, 30 and 40 mg / kg i

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7 physiological saline solution. After 5 days, all animals are each injected with 2 x 10 8 sheep erythrocytes into the sole and 24 tips later the swelling of the foot is measured. The foot swelling is triggered by a delayed type hypersensitivity (DTH) skin reaction and, as one skilled in the art will know, is a target of the cellular immune response [F.M. Collins, and G.B. Mackaness, J. Immunol.

101, 830-845 (1968)]. The results in Table I show that when administering the g 10 substance obtained according to the invention, e.g. after immunization with 10 sheep erythrocytes comes an increase in the cellular immune response.

A maximum of the stimulation can be observed in this test series by administration of 30 mg / kg of test substance.

Table I

Immunization of mice with sheep erythrocyte effect on the cellular immune response (DTH reaction) 2 x daily i.p. administration% foot swelling at on day -3, -2, -1, 0 of_10 ** erythrocytes 20 pbs1 24.3 + 1.3 20 mg / kg 28.3 + 6.7

Test substance 30 mg / kg 36.5 + 6.5 40 mg / kg 31.4 + 8.7 25 1 PBS = phosphate buffered saline (NaCl: 8000 mg / 1, KCl: 200 mg / 1, Na2HPC> 4.2H20: 1440 mg / l, KH 2 PO 4: 200 mg / l.

Experiment 2 30 Influence on the stimulation of nonspecific immunity -activation of mononuclear phagocytes

Here, the influence of the test compound obtained according to Example 8 is investigated on the stimulation of peritoneal macrophages in 6-8 week-old NMRI-35 mice. Young female mice are given the test compound by parenteral or oral route at a dose of 25, 50, 100 and 200 mg / kg.

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The control group receives buffered saline. Three days after the injections, the mice are sacrificed and the animals' peritoneal macrophages are examined for their state of activation. As the target of macrophage activation, the secretion of the lyosomal enzymes (β-galactosidase, β-glucuronidase, N-acetyl-3-D-glucosaminidase) is determined.

On the other hand, in comparable macrophage cultures, pinocytosis can be investigated by the uptake of carbon-198 solid gold (Au) - as will be known to those skilled in the art.

The size of the oxidative metabolism at the macrophages is a further measure of their activation state. This activity is measured by biolumines in determining the chemoluminescence.

To this end, either in petri dishes with a diameter of 30 mm is grown. 3 x 10 6 macrophages with 1 ml TC 199 culture medium or 10 macrophages with 100 µl in round-bent polyethylene glass (to determine chemoluminescence) at 5% CO 2 and 37 ° C.

After 1 hour of incubation, the cultures are washed to remove swimming cells. The chemoluminescence (test tube culture) is then determined directly, while the petri dishes are again incubated for 24 hours at 37 ° C, and then the enzyme and pinocytosis activity in the cultures is determined.

The results below are obtained.

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Table II

Effect on oxidative metabolism by mouse peritoneal macrophages (chemoluminescence in RLE / 15 minutes).

1 x administration 30 of _intraperitoneal_orally_ PBS 2.97 + 0.28 x 105 3.65 + 0.81 x 105

Sample 25 mg / kg 7.87 + 0.28 x 105 6.41 + 0.42 x 105 compound-50 mg / kg 9.72 + 0.82 x 105 8.99 + 0.39 x 105 score 100 mg / kg 12.85 + 2.37 x 105 11.85 + 0.92 x 105 35 cc 200 mg / kg 24.40 + 3.39 x 10 ° 15.60 + 1.98 x 10 ° 1 RLE = relative light units 9

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The parenteral and oral treatment of NMRI mice with the test compound prepared according to Example 8 stimulates macrophage activity and thus * has an immune-stimulating effect. Thus, the oxidative metabolism of macrophages is significantly enhanced with the formation of oxygen radicals and the associated measurable light. At doses of 25 mg / kg and up, a dose-dependent increase in macrophage activity occurs both at parenteral and oral administration.

Table IH shows that corn macrophages from control mice release only small amounts of lysosomal enzymes (β-glucuronidase, β-galactosidase, N-acetyl-3-D-glucosaminidase) in the culture superphase. Mononuclear phagocytes from mice treated parenterally or orally with the test compound for 72 hours clearly secrete more of the above acidic hydrolases (β-Glu, β-Gal, N-Ac-Glu) and exhibit a dose-effect curve, which in all measured enzymes are the controls superior. It is seen that the test compound has a stimulatory effect on macrophage activity and contributes to an increase in enzyme release.

Table III

Influence of the test compound on enzyme release of lysosomal hydrolases from mouse peritoneal macrophages 25 1 x i »p./p» o. β-Glu β-Gal N-Ac-Glu administration _._ mU / ml_mU / ml_mU / ml PBS 755/484 1306/1702 1238/1168

Sample 25 mg / kg 1001/897 2584/4917 2786/1947 for 50 mg / kg 1370/1133 11058/9179 3315/2862 30 bin 100 / mg / kg 1791/1593 17596/13195 4305/3676 200 mg / kg 2136/1886 22351/17357 6548/5621

The quantitative determination of pinocytosis activity by mononuclear phagocytes is by means of

Davies et al's method [P. Davies, A.C. Allison and A.D.

Haswell, Biochem. Biophys. Res. Com. 52, 627 (1973)].

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For this, radioactive colloidal gold (Au) with a particle size of 20 nm and a specific activity of 4-12 mCi / mg Au is used. The results in Table IV show the effect of the test compound obtained in Example 8 on the endocytosis performance. The colloidal gold (Au) pinocytosis by mouse peritoneal macrophages from animals treated with the compound of the invention is significantly and dose-dependently increased compared to macrophages from untreated animals.

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Table IV

Effect of the test compound on mouse macrophages' pinocytosis performance 1 x administration of the intrapsritoneal_orally_ 15 PBS 0/286 x 10 ^ cpm 0.198 x 10 ^ cpm

Sample 25 mg / kg 0.341 "" 0.272 vs. 50 mg / kg 0.396 "" 0.358 bin 100 mg / kg 0.462 "* 0.416" "200 mg / kg 0.587" "0.506 20

Experiment 3

Increasing the resistance of Balb / c mice to infection by Candida albicans ). Twenty-four hours after the last administration of the test compound, these animals and the control animals, which have received physiological saline solution in equal amounts and at the same interval, are infused intravenously with Candida albicans (5 x 10 5 CFU / mouse). Based on the mortality after infection, the average survival time can be calculated. Of the animals in the control group, 50% die after 9.7 days and the group treated with the test compound has an average o-survival time of 16.1 days. Within the selected item 11

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gift scheme with the corresponding dosages (prophylactic administration) results in a significant increase in Balb / c mice 'resistance to Candida albicans.

Table V

Average survival times after infection with 5 x 10J CFU C. albicans Compound Gen. over 2x60 mg / kg / level time Confidence range 10 days i.p ._ (days) _95% _99%.

PBS 9.7 8.4-10.6 7.8-10.9

Test compound 16.1 14.8-17.4 14.4-17.8 (b) Therapeutic treatment 15 In the therapeutic treatment of a chronic infection with Candia albicans, Balb / c female mice (15 / group) on day 0 are intravenously infected. with Candida albicans 5 (1 x 10 CFU / mouse). After infection is completed, the animals are treated 8 consecutive days (days 3-10) every 20 with 60 mg / kg of the test compound (the compound of Example 8). The control animals are injected with physiological saline solution. On days 8, 14 and 21, urine samples are taken from the animals and the seed count is determined.

On day 30, the animals are sacrificed and the germination and necrosis formation in the kidneys is determined. Table VI shows that the test compound at therapeutic administration clearly reduces all of the chronic Candida albicans infection parameters (germ count in urine and kidneys and necrosis) and thus affects the disease therapeutically. While a high percentage of bacteria can be detected in the control animals in the control animals, the number of bacteria is significantly lowered in the treated animals.

Also, the bacterial colony formation in the kidneys is reduced by the therapy from 63 to 27%, and at the same time the necrosis formation is reduced from 87 to 10%.

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Table VI

Therapeutic treatment of a chronic Candida albicans infection (1 x 10 CFU) *

Compound Kidneys with necro- 60 mg / kg Animal pos. bacterial i.p. with a positive result. kidneys

Day 3-10 Day 8 Day 14 Day 21_Day * 30 Day 30 PBS 4/15 8/15 10/15 19/30 26/30 (27%) (53%) (67%) (63%) (87%) 10 Test Form 1/15 1/15 4/15 8/30 3/30 Binding Ice (7%) (7%) (27%) (27%) (10%)

Experiment 4

Stimulation of the DTH reaction with the compounds of the invention NMRI-Mouse is treated with the compounds of the invention as in Experiment 1.

As a test to determine the immune stimulation, the DTH reaction is tested.

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Table VII shows the relative function of the test compound relative to the compound of Example 8f whose maximum activation equals 100% (difference between control and stimulation). The table shows that the DTH reaction in the pre-treated animals with the test compounds is clearly stronger than in the corresponding control animals.

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Table VII

Compound Dose DTH reaction according to ex_ (mq / kq) Entry_ (SRBC) _ 2 20 1 x i.p. day 0 88% 5 5 200 "61% 6 100" 100% 8 100 "* 100% 10 10" 112% 12 100 "96% 10 13 200" 147% 14 100 "118%

Experiment 5

Stimulation of Macrophage Activity with Compounds of the Invention NMRI-Mouse is treated with the compounds of the invention as described in Experiment 2.

To test the immune stimulation, the function of macrophages (chemoluminescence and enzyme activity) is tested.

Table VIII shows the relative activity of the test compounds relative to the compound of Example 8, whose maximum activation (difference between control and stimulation) corresponds to 100%. The table shows that, compared to macrophages from untreated animals, these cells are also strongly stimulated with all test compounds for chemoluminescence reaction and their lysosomal enzyme content.

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Table VIII

Connection - According eg Macrophage activity Λ (100 mg / kq ip) - Chemoluminescence Exocytosis 5 1 30% 48% 2 72% 53% 5 37% * 45% 6 26% 33% 8 100% 100% 10 10 66% 21% 12 39% 54 % 13 81% 77% 14 97% 93% 15 Experiment 6

Stimulation of infection defense against Candida albicans infections by prophylactic administration of the compounds of the invention

Balb / c Mice are treated with the compounds of the invention as described in Experiment 3a.

As shown in Table IX, the mean survival time of the mice after C. albicans infection increases significantly when compared to an untreated control group.

Table IX

Dosage

Ex. (mg / kg) Relative extension of average

& No. 1 x i.p. day_ survival times after infection 1 1 144 30 2 1 123 5 10 142 6 10 145 8 1 130 10 10 135 35 12 10 205 14 10 107

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15 * = Mean survival time of the control group = 100.

Experiment 7 5 Influence of stimulation of tumor defense against B16 melanoma In C57Bl / 6 mice (10 animals / group) weighing 5 18-20 g, 2 x 10 6 live B16 melanoma cells are induced by a primary tumor. After designing a specific tumor size (0.65 cm in diameter), the primary tumor is removed.

10 The untreated animals then die from metastases in the lungs.

Following tumor induction, animals are treated intraperitoneally on days 3, 5, 7, 9, 11 and 13 after amputation of the primary tumor with 50, 100 and 200 mg / kg of the test compound obtained in Example 8. Based on the mortality after 15 amputation of the primary tumor due to the development of the lungs, taser is taken, correspondingly the average survival time can be calculated. Then the animals in the control group die by 50% after 26 days. The groups treated with the test compound (cf. Table X) with a corresponding 20 doses exhibited an increase in the average survival time extension of 43, 40 and 35 days.

Table X

Therapeutic Tumor Treatment of B16 Melanoma 25 Administration 6 x i.p. (mg / kg) Average survival time days 3, 5, 7, 9, 11 and 13_ (days) _ PBS 26

Sample 50 43 Connection 100 40 30 Solution 200 35

Experiment 8

Influence on bone marrow scolonia formation

Here, the influence of the test compound obtained in Example 8 on the stimulation of bone marrow colonies in 6-8 weeks old B2D2F1 mice is examined.

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Small female mice receive the test compound intraperitoneally at doses of 2.5 and 5 mg / kg. A day later, the animals / bone marrow cells are isolated and cultured according to generally known methods [Metcalf, Immunology 21, 427 (1971), and Stanley et al., J. Exp. With. 143, 631 (1979)].

To develop the bone marrow colonies, the usual colony stimulating factor (MCSF) source is used as the usual L-cell fluid phase (15%). As shown in Table XI, a single administration of 2.5 or 5 mg of the test compound leads to a marked increase in colony formation in bone marrow cells both with and without the addition of Colony stimulating factor (CSF) in vitro.

Table XI

In vivo effect on bone marrow colony formation

Test compound Number of bone marrow colonies (day 8) 1 x i.p. (mg / kg) _with CSF (15%) _without CSF (15%) PBS 41 + 6 0

Test Form 2.5 94 + 11 24 + 2 20 Bond 5.0 163 + 8 44 + 5

The invention will now be explained in more detail by way of example.

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Example 1

Bis- (3-Hydroxy-1-phenyl-1,2,4-triazol-5-yl) disulfide 2.9 g (15 mmol) of 3-hydroxy-1-phenyl-5-mercapto - 1.2 4-Triazole is added to 100 ml of methanol and dropwise at room temperature 3.2 ml (33 mmol) of 35% hydrogen peroxide in methanol. The solution first turns yellow and after 20 minutes the compound slowly precipitates.

After 4 hours, the precipitate is aspirated, washed with methanol and dried.

Yield: 2.1 g = 72% of theory.

Melting point: 232 ° C (decomposition).

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NMR (d 6 -DMSO) δ: 11.58 ppm, s, 2H, OH

7.38 ppm, m, 10H aromat. H.

*

Example 2 Bis- (5-carboxy-benzimidazol-2-yl) disulfide

Preparation analogous to Example 1 from 2 - mercaptobenzimidazole-5-carboxylic acid.

Yield: 70% of theory.

Melting point: 235-238 ° C.

NMR (d 6 -DMSO) έ: 9.4 ppm, d, NH

7.4-8.4 ppm, m, aromatic H.

Example 3 Bis- [4- (2-chloroallyl) -6-hydroxy-5-oxo-4,5-dihydro-1,2,4-triazin-3-yl] disulfide 2.2 g 4- ( Dissolve 2-chloroallyl) -6-hydroxy-3-mercapto-5-oxo-4,5-dihydro-1,2,4-triazine in 25 ml of methanol, dropwise at room temperature, 0.86 ml of 35% H -20 after stirring for an hour to crystallize the product. It is sucked off, washed with methanol and air dried. Yield: 1.2 g.

Melting point: 204-205 ° C (decomposition).

NMR (d 1 -DMSO) b: 12.7 ppm (br s, OH)

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5.5 ppm (dd, = CH 2) 4.8 ppm (s, CH 2 N).

Example 4

Bis- (6-hydroxy-2-methyl-5-OXO-2,5-dihydro-1,2,4-triazin-3-yl) disulfide 1.59 g of 6-hydroxy-3-mercapto-2 -Methyl-5-oxo-2,5-dihydro-1,2,4-triazine is dissolved in 30 ml of methanol, which is dropped at room temperature 0.43 ml of 35% H 2 O 2 and then stirred for 30 minutes. The solution is clarified with charcoal, evaporated and the residue is triturated with ice water. The precipitate is filtered off, washed with ice water, air dried, suspended.

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18 in methanol, filtered off, washed with a little methanol and dried in vacuo.

Yield: 0.8 g-

Melting point: 220 ° C (decomposition).

Δ NMR (d 6 -DMSO) δ: 3.87 ppm (s, CH.).

Example 5

Bis- (6-hydroxy-4-methyl-5-oxo-4,5-dihydro-1,2,4-triazin-3-yl) disulfide 3.7 g of 6-hydroxy-3-mercapto-4 -Methyl-5-oxo-4,5- -dihydro-1,2,4-triazine is dissolved in 400 ml of methanol and 38 ml of water. At room temperature, 2.5 ml of 35% H 2 O 2 is dropped.

After stirring for 30 minutes, evaporate to 100 ml, the precipitate is sucked off and dried.

Yield: 1.5 g.

Melting point: 237 ° C.

NMR (CF 3 CO 2 D) δ 3.67 ppm s, 2H, OH

3.8 ppm s, 6H, CH 2 Example 6

Bis- (5-carboxymethyl-4-methyl-1,3-thiazol-2-yl) disulfide 23.62 g of 5-carboxymethyl-4-methyl-2-mercapto-1,3-thiazole are dissolved in 250 ml of methanol, filter, then slowly, while cooling in water bath, add 12.5 ml of 35% H2 O2. Then stir for 30 minutes in a water bath and 2 hours at room temperature. After filtration and washing with methanol, 22.8 g of the title compound are isolated.

Melting point: 16 2 ° C.

NMR (CF3 <X> 2D) S: 2.6 ppm, s, 6H, CH3 4.2 ppm, s, 4H, CH2

Example 7

Bis- (5-carboxymethyl-1,3-thiazol-2-yl) disulfide 0.88 g of 2-mercapto-1,3-thiazol-5-yl acetic acid is dissolved in ca. 10 ml of methanol and added under stirring.

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19 ring at room temperature 0.5 ml of 33% E ^ C ^ solution. After half an hour, the crystal suspension is cooled, filtered and the filtration residue dried. Yield: 0.6 g with a melting point of 150 ° C. Comparison by thin layer chromatography on Merck silica gel plates shows complete turnover. (Rf: 0.3, eluent: acetic acid ester / ethanol / water / formic acid = 65: 25: 10: 1).

Example 8 Bis- (4-carboxymethyl-1,3-thiazol-2-yl) disulfide When 1.75 g of 4-carboxymethyl-2-mercapto-1,3-thiazole is reacted analogously to Example 7, 0.95 is obtained. g of the compound mentioned in the title.

Melting point: 163-165 ° C.

NMR (DMSO-d 6) δ: 3.7 ppm, s, 4H, CH 2

7.6 ppm, s, 2H, CH

9.4 ppm, s, 2H, CC

Example 9 20

Bis- (2-carboxymethylthio-1,3,4-thiadiazol-5-yl) disulfide 1.4 g (2-mercapto-1,3,4-thiadiazol-5-yl) mercaptoacetic acid is dissolved in 10 ml methanol and 0.65 ml of 35% H 2 O 2 is added dropwise under ice-cooling. Stir for one hour at room temperature and then filter. The crystals are washed with a little methanol and dried in vacuo.

Yield: 1.3 g.

Melting point: 179 ° C (decomposition).

NMR (DMSO-d 6) δ: 13 ppm, broad, CO 2 4.2 ppm, s, CEL ·.

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Example 10

Bis- (5-carboxy-1,3-thiazol-2-yl) disulfide 21 g of 2-mercapto-1,3-thiazole-5-carboxylic acid is dissolved in 100 ml of methanol under gentle heating. During ice cooling.

35 ling slowly add 10.7 ml of 35% H 2 O 2 and then

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The mixture is stirred for 45 minutes at room temperature. It is extracted, washed with methanol and dried in vacuo.

4 Yield: 21.5 g

Melting point: 280 ° C (decomposition).

Δ NMR (DMSO-d -) δ: 8.48 ppm, s, thiazole-H.

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Example H

Bis- (5-carboxy-4-methyl-1H-3-thiazol-2-yl) disulfide 1.75 g (10 mmol) of 4-methyl-2-mercapto-1-3-thiazole-10-5-carboxylic acid is suspended. in 50 ml of methanol, and under ice-cooling, 0.86 ml of 35% H2 O is added, then stirred for one hour at room temperature. The crystals formed are sucked off, washed with a little methanol and dried in vacuo.

Yield: 1.52 g.

Melting point: 240-241 ° C (decomposition).

NMR (DMSO-dJ S * 2.6 ppm, s, CH

O 3

Example 12 Bis- (4-carboxymethyl-5-methyl-1,3-thiazol-2-yl) disulfide

Step 1: 4-Bromopropionylacetic acid methyl ester Dissolve 22 g of propionylacetic acid ethyl ester in 60 g of CH2 Cl2 and drop 8.7 ml of Br2 into 30 ml of CH2 Cl2, then stir for 1.25 hours at room temperature.

Then add 0.66 ml of Br 2 again in 5 ml of CH.C1-, Δ 2. Δ and stir for an additional 1.25 hours. Wash with 3 x 50 ml H 2 O, 1 x with 20 ml saturated NaHCO 3 and 2 x with 50 ml H 2 O, dry over Na 2 SO 4 and evaporate. This yields 38.6 g of oil.

Step 2: (2-Mercapto-5-methyl-1,3-thiazol-4-yl) acetic acid thyl ester in 200 ml of ethanol / 200 ml of water.

Stir for 2.5 hours, then add 7.5 ml of trifluoroacetic acid. After stirring for one hour, evaporate

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21 to dryness and 200 ml of CHCl 3 / I 2 O (= 1: 1) is added.

Wash twice more with water, dry over sodium sulfate and then concentrate. The solid residue is suspended in ether and filtered off.

Yield: 8.9 g.

Melting point: 149 ° C (decomposition) *

NMR (dg-DNSO) δ · 11/0 ppm, broad, SH

3.7 ppm, s, AND 3.5 ppm, s, CH 2 2.1 ppm, s, CH 3

Step 3: (2-Mercapto-5-methyl-1,3-thiazol-4-yl) -acetic acid 1.98 g of the compound of step 2 is dissolved with heating in 20 ml of methanol. Stir for 40 minutes at room temperature, add 50 ml of water and then acidify with concentrated hydrochloric acid to pH 1.0. After an hour of stirring under ice-cooling, the compound is suctioned off and washed chloride-free with ice water. Drying in vacuo over P205 gives 1.67 g.

NMR (dg-DMSO) δ: 13.9 ppm, broad, CC> 2H 3.5 ppm, s, CH 2 2.08 ppm, s CH 3

Step 4: Bis- (4-carboxymethyl-5-methyl-1,3-thiazol-2-yl) disulfide 430 mg of the compound of step 3 is suspended in 20 ml of methanol and dissolved under heating. At room temperature 0.25 ml of 35% H 2 O 2 is added dropwise and stirred for one hour. After evaporation to ca. The mixture is filtered for 5 ml and washed with slightly ice-cooled methanol.

Yield: 280 mg.

NMR (dg-DMSO) δ: 3.65 ppm, s, CH 2 2.37 ppm, s, CH 3

Example 13

Bis- (5-Carboxymethyl-1,3-thiazol-2-yl) -disulfide 0.8 g of 2-mercapto-5-carboxymethyl-1,3-thiazole is dissolved in 10 ml of methanol and is added dropwise with stirring.

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22 O.

0.5 ml of 33% H 2 O 2 solution. Stir overnight and filter out the crystallized product.

Yield: 0.6 g, m.p. 150 ° C (decomposition), thin layer chromatographically uniform (Rf = 0.3, silica gel, eluent: acetic acid ester / ethanol / water / formic acid = 60: 25: 15: 1.

IR (KBr tablet) $: 1710 cm (COOH).

1 H NMR (d 6 -DMSO) δ in 3.8 ppm (s, CH 2 »thiazole, 2H) 7.6 ppm (s, thiazole-4H, 1H).

10

Example 14

Bis- (4-Carboxy-1,3-thiazol-5-yl) disulfide 5 g of bis- (4-methoxycarbonyl-1,3-thiazol-5-yl) disulfide is suspended in 100 ml of methanol. To this is added a solution of 1.5 g of NaOH in 20 ml of H2 O and then heated for one hour under reflux. After the solution has cooled, methanol is evaporated on a rotary evaporator, the aqueous phase is extracted once with a little acetic acid ester and acidified with 2N HCl. The product 20 is filtered off, dried and recrystallized from acetic acid ester. 2 g of melting point are obtained 154 ° C.

IR (KBr tablet):: 1680 cm ”1 (COOH).

1 H-NMR (d 6 -DMSO) δ 8.6 ppm (s, thiazole-2H).

Examples 15-17 are carried out as described in Example 1.

Example 15 Bis- (5-glutaric acid monoamido-1,3-thiazol-2-yl) disulfide 1 H-NMR (dg-DMSO) δ 1.6-2.6 ppm (m, 12H, six CH 2 groups) 7.50 ppm (s, 2H, thiazole-4, 12.63 ppm (br s, 2H, -NH-), 13.05 ppm (br s, 2H, -CO2H).

ISLAND

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23

Example 16

Bis- (5-succinic acid monoamido-1,3-thiazol-2-yl) disulfide 1 H-NMR (d.-DMSO) δ:

* Oh

2.4-2.7 ppm (m, 8H, -CH2), 7.50 ppm (s, 2H, thiazole-H), 12.66 ppm (br s, 2H, -NH-), 13.15 ppm (br s, 2H, -CC> 2H).

Example 17 Bis- [5- (2-carboxyeth-1-yl) -4-methyl-1,3-thiazol-2-yl] disulfide 1 H-NMR (dg-DMSO) δ: 2.27 ppm (s, ), 2.55 ppm (d, 4H, thiazole-CH2), 2.96 ppm (t, 4H, CH2-CC> 2-).

15 20 25 30 35

Claims (4)

Disulfides, characterized in that they have the general formula 1 ' Het'-SS-Het '(I ·) 9 wherein Het' means 1,3-thiazolyl substituted with C1-6 alkyl, carboxy-C1-6 alkyl, carboxy or C2-5 acylamino, where acyl means a residue of an aliphatic dicarboxylic acid, 1,2,4-triazolyl, which is substituted by hydroxy and phenyl, 1,3,4-thiadiazolyl, which is substituted by carboxy-C 1-4 alkylthio, dihydro-1,2,4-triazinyl, which is substituted with C 1-6 alkyl, halo-C 2-4 alkenyl, oxo or hydroxy, or benzimidazolyl substituted with carboxy, wherein Het 'is substituted by at least one directly or to a substituent bonded carboxy group or a directly bonded hydroxy group, whereby these groups may also be in the form of a physiologically acceptable salt. Disulfides, characterized in that they have the general formula 1 'Het'-SS-Het' (I ') wherein Het' means 1,3-thiazolyl substituted 1 or 2 times by C1-3 alkyl, carboxy-C1_3 -alkyl, carboxy or C2-5 acylamino, where acyl is a residue of an aliphatic dicarboxylic acid, wherein Het 'is substituted by at least one directly or to a substituent bonded carboxy group which may also be in the form of a physiologically acceptable salt. Process for the preparation of disulfides of the general formula I ', characterized in that (a) a compound of the general formula II 4 has Het'-SH (II) where Het' has the above meaning, by reaction with an oxidant is converted into a compound of general formula I ', or (b) a compound of general formula III Het'-X (III) 10 wherein Het' is as defined above and X is a reactive, readily cleaved group , Me is an alkali metal, or (c) a compound of the general formula IV Het'-S-SO2Me (IV) wherein Het 'and Me have the above meanings are reacted with iodine in an aqueous medium. Pharmaceutical agent, characterized in that it contains a compound of general formula I '.