Hydrochloride sulfur-containing 2-aminoimidazole or 2 - aminothiazoles with cytotoxic and antitumor activity and exhibiting an inhibitory effect on certain enzymes

 

(57) Abstract:

The invention relates to organic chemistry, and more specifically to new connections - hydrochloridum 2-aminoimidazole and 2-aminothiazole General formula (1),

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aryl, the substituent in position 4 and a disulfide bridge in the position 5, where X is alkylamino, for example, methylamino, and R1-R2is hydrogen; X-methylaminopropyl, and R1-alkyl, for example methyl and R2is hydrogen; X-methylaminopropyl, and R1-alkoxygroup, for example, methoxy and R2is hydrogen; X-methylaminopropyl, and R1-ethoxypropan and R2is hydrogen; X-methylaminopropyl, and R1halogen, for example chlorine and RF2is hydrogen; X-methylaminopropyl, and R1-R2-alkoxygroup, for example, methoxy; X-atramentaria, and R1-alkoxygroup, for example, methoxy and R2is hydrogen; X is sulfur, and R1-alkoxygroup, for example, methoxy and R2is hydrogen; X is sulfur, and R1-R2-alkoxygroup, for example, methoxy; X is sulfur, and R1halogen, for example fluorine and R2-hydrogen. The new compounds possess cytotoxic and antitumor activity and exhibit an inhibitory effect on certain enzymes. 1 Il., table 4.

The invention relates Kali (1),

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aryl, the substituent in position 4 and a disulfide bridge in the position 5, where X is alkylamino, for example, methylamino, and R1-R2-hydrogen (compound 1); X-methylaminopropyl, and R1-alkyl, for example, methylamino, and R1-R2-hydrogen (compound I); X-methylaminopropyl, and R1-alkyl, for example methyl and R2-hydrogen (compound II); X-methylaminopropyl, and R1-alkoxygroup, for example, methoxy and R2-hydrogen (compound III); X-methylaminopropyl, and R1-ethoxypropan and R2-hydrogen (compound IV); X-methylaminopropyl, and R1halogen, for example chlorine, and R2-hydrogen (compound V); X-methylaminopropyl, and R1-R2-alkoxygroup, for example, methoxy (compound VI); X-atramentaria, and R1-alkoxygroup, for example, methoxy and R2-hydrogen (compound VII); X is sulfur, and R1-alkoxygroup, for example, methoxy and R2-hydrogen (compound VIII); X is sulfur, and R1-R2-alkoxygroup, for example, methoxy (compound IX); X is sulfur, and R1halogen, for example fluorine and R2-hydrogen (compound X); which have cytotoxic and antitumor activity and exhibit an inhibitory effect on certain enzymes.

Among Bo is on the amino group in position 2, the aryl substituent in position 4 and sulphurous function in position 5. From the known 2-aminoimidazole most similar in structure to the claimed connections are(2) [1] (3) [2] (4) [3] (5) [4] (6) (10) [5] (11) [6,7] (12) [3,6,7] (13) (15) [8] (16) [9 12] (17) [13] and (8) [13] As can be seen, only the connection (17) and (18) contain a substituent in position 1, that is, in structure they are most similar to the compounds of the type (1). However, in position 4 they contain a Deputy of a different nature than in compounds of the type (1), and do not contain sulfur functions in position 5.

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All connections (2) (18) are in one way or another biological activity. Most of them exhibits a pronounced antimicrobial activity whose values reach the maximum value of these compounds as origin (4), desalkylation (5), sceptrin (12), nomen (17) and ionamin (18).

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It was reported about the manifestation of compounds (12A), (12B), (12B), (13), (14) and (15) antiviral activity against Herpes simplex virus, type I virus and Vesicular stomatitis [6]

Some of the discussed 2-aminoimidazole are antagonists of a number of receptors. So, pimenidis (3) antagonist of serotonergic receptors, and claritin (2) has a strong blocking action against Holiness the>It was noted activity 2-aminoimidazole in respect of a number of enzymes. So, purelin (6) is an activator of myosin K+, EDTA-ATPase [5] and oxazepam (11b), ageliferin (13), bromelain (14) and dibromothiophene (15) have a powerful activating effect on actomyosin ATPase of myofibril from skeletal muscle of the rabbit [7, 8] Compound (6) (9) exhibit a pronounced inhibitory activity against Na+, K+-ATPase, and (10) weakly inhibits it (22% inhibition at a concentration of 10-4M).

There is very little information on the cytotoxicity and antitumor activity of 2-aminoimidazole. For Perelygina (10) showed significant cytotoxicity against cells L1210murine leukemia in vitro (EID501.1 µg/ml [5] ), and for cladribine (2) in relation to cells Cho-K1 (ID50of 1.33 mg/ml [1]).

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Among the known 2-aminoimidazole only girallon (16) shows antitumor activity against leukemia cells P388. Intraperitoneal injection of mice EID50was 1.0 mg/kg [9] In the literature are no longer reported on the expression of antitumor activity of some 2-aminoimidazole.

Known also substituted in position 1, the imidazoles, the soda is CLASS="ptx2">

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These include derivatives of 4-mercaptopyridine as isolated from marine invertebrates ovariole(19) (21) [14 18] adenochrome And (22) [19, 20] and imbrication (23) [21] Assumes that bodily can act as biological antioxidants, being in the early stages of development, embryos of marine invertebrates to trap intermediates having active oxygen [22 24]

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As can be seen, compound (19) (23) are significantly different in structure from the compounds of the type (1): first, they do not have aminophenol in position 2, and secondly, sulfur function is in position 4, not 5, and has a different character and, thirdly, the substituent in the heterocyclic nucleus is positioned at 5 instead of 4, and is alkyl, and not aryl.

Oxidative destruction imbrication (23) gives ovation And (19), further oxidation which iodine in a weak solution of hydrochloric acid leads to disulfide (24), having, like all the above-mentioned compounds, significant structural differences from the claimed compounds of the type (1). On the biological activity of (24) have been reported.

Comparison of the structures of the compounds of the type (1) and (2) (24) shows that compounds (1) are new, not previously known structural t the practical origin.

In this regard, among the known imidazoles impossible to choose a structural prototype for compounds (1). Among 2-aminoimidazole closest to (1) the structure of compounds (17) and (18), and among the sulfur-containing imidazoles connection (24), but they all have, at the same time, a very significant structural differences from compounds of the type (1).

If you find the prototypes on the biological effects, none of the above compounds (2) (24) shows the literature data of such a wide spectrum of biological activity, as the connection type (1), although the connection (10) and (16) and are cytotoxic and antitumor effects, the most characteristic of the claimed compounds.

Among the many described in the literature of thiazolo known just one example compounds containing in the molecule both the amino group in position 2, the aryl substituent in position 4 and sulphurous function in position 5, the connection (25) [25]

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Oxidation of compound (25) can be obtained disulfide, which is a close analogue of the compounds (VII) to(X). This conversion, however, are not described in literature.

From the other close to the claimed compounds described compounds (26) [25, 26] and (27) [26] As can be seen, characteristie 4.

Among the known compounds of this series described even thiazole containing disulfide bridge in position 5 (compounds (28) and (29)) [27] but the nature of the substituents in positions 2 and 4 of these compounds, other than claimed. However, it should be noted that evidence of the structure of the compounds (28) and (29) the authors did not give.

On the biological activity of compounds (25) (29) no information no. In the literature there are no reports of manifestation 2-aminothiazole cytotoxic and antitumor activity. Only for derivatives of 2-aminothiazolo for NH2the group found ratingaverage activity against cells of leukemia L1210. Thus, methyl 4-(isothiocyanate)thiazol-2-carbamate (30) inhibits the growth of leukemia cells L1210at a concentration of EID503,2 mg/ml [28] the Primary mechanism of action of this compound is a mitotic block. In addition, the connection (30) showed significant in vivo antifilarial activity against adult worms Acanthocheilonema viteae.

Some 2-N-substituted 2-aminothiazole, for example, compounds of the type (31) and (32), show anti-inflammatory and diuretic activity, and also affect the Central nervous system [29]

Drucilla Heliothis virescens [30]

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The simplest 4-phenylsilane 2-aminothiazolo connection (34) has an anesthetic effect in relation to fish [31]

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However, most often 2-aminothiazole and their derivatives are used as antimicrobial agents, since many of them have exhibited high antifungal and antibacterial activity. So, for example, compounds of the type (35) exhibit high fungistatic activity against Candida albicans, Aspergillus fumigatus and Epidermophyton floccosum [32]

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2-Aminothiazoline fragment is part of the structure of effective modern cephalosporins, for example, used in the clinic ceftizoxime (36) [33] cefdinir (37) [34] and other natural and synthetic cephalosporins [35 38] they All possess a wide spectrum of antibacterial activity and is very effective against common aerobic gram-positive bacteria (streptococci and methicillin sensitive staphylococci) and gram-negative bacteria (family Enterobacteriaceae, Hemophilus, meningococci, gonoccocal). Most of these shows stability of plasmid and chromosomal beta-lactamase.

Comparison of the structures of the inventive 2-aminothiazole (compound (VIII)-(X)) son case, the compound (25) can serve as a close structural prototype for the compounds (VIII)-(X), although the nature of the substituent in position 5 (Tolna group) other than in the claimed compounds (disulfide group). This structural difference is still significant. At the same time, information about the biological activity of the compound (25) in the literature. Moreover, in the literature there are no reports on the expression of 2-aminothiazole cytotoxic and antitumor activity, which is characteristic of the claimed compounds. Thus, if a structural prototype for the claimed compounds (VIII) to(X) can be chosen (with some reservations), the prototype on the physiological action could not be found.

Declare sulfur-containing 2-aminoimidazole and 2-aminothiazole are new compounds and are not described in literature.

The basis of the invention is the creation of new substances with cytotoxic and antitumor activity, showing an inhibitory effect on certain enzymes and operating in low doses, which can serve as a basis for development of new drugs that extend the range of means of influence on pathogens of humans and animals.

The problem is solved by the fact that claimed new X-alkylamino, for example, methylamino, and R1-R2-hydrogen (compound I); X-methylaminopropyl, and R1-alkyl, for example methyl and R2-hydrogen (compound II); X-methylaminopropyl, and R1-alkoxygroup, for example, methoxy and R2-hydrogen (compound III); X-methylaminopropyl, and R1-ethoxypropan and R2-hydrogen (compound IV); X-methylaminopropyl, and R1halogen, for example chlorine, and R2-hydrogen (compound V); X-methylaminopropyl, and R1-R2-alkoxygroup, for example, methoxy (compound VI); X-atramentaria, and R1-alkoxygroup, for example, methoxy and R2-hydrogen (compound VII); X is sulfur, and R1-alkoxygroup, for example, methoxy and R2-hydrogen (compound VIII); X is sulfur, and R1-R2-alkoxygroup, for example, methoxy (compound IX); X is sulfur, and R1halogen, for example fluorine and R2-hydrogen (compound X), as drugs that extend the range of means of influence on pathogens of humans and animals.

Declare sulfur-containing 2-aminoimidazole and 2-aminothiazole show cytotoxic activity against a broad set of cell lines of different types of tumors (studied 75 cell lines 10 types of tumors), antitumor action is the relation K+, Na+-ATPase from rat brain and reverse transcriptase from rous sarcoma virus and avian myeloblastoma.

The goal can be achieved either by selection of compounds of the type (1), including compound III).

It should be emphasized that the study of the structure and physiological activity of compound III (it is called polycarpon), selected by us for the first time from tropical ascidians Polycarpa aurata, gave impetus to the development of studies on the synthesis of this compound, and as a consequence, the synthesis of its imidazole analogs I, II, IV-VII, and thiazole analogues VIII-X. the Study of the physiological activity of all these compounds and was eventually the subject of the present invention. Other compounds of the type (1) natural sources we have not identified.

The best option of carrying out the invention

The inventive compounds of sulfur-containing 2-aminoimidazole and 2-aminothiazole are crystalline or amorphous material, the color changes from yellow to orange. They are all stable under normal storage conditions. The claimed compounds are well soluble in water, sparingly in ethanol and dimethyl sulfoxide.

The structure and composition of the inventive compounds of General formula (1) , illustrate the formation of compounds of type (1).

Example 1. The selection of compound III from a natural source.

Lyophilized ascidia Polycarpa aurata (100 g), collected off the coast of Australia, were extracted during the week, 800 ml of ethyl alcohol. The extract was evaporated to dryness and the residue (1.5 g) was chromatographically sequentially by columns with polychrome-1 (solvent system of water:ethanol 2:1), silica gel (100 to 150 mesh.) (the solvent system chloroform:ethanol 6:1) and Sephadex LH-20 (solvent system chloroform:ethanol 4:1), collecting colored faction. Last decimal by the method of preparative HPLC on a column of Phenyl-Si100-Polyol in the solvent system acetonitrile:water 4:1. The obtained colored faction. Removal of solvents and double crystallization of the residue from a mixture of chloroform:ethanol 1:4 gave 13 mg (0,013%) of the dihydrochloride of bis[2-amino-1-methyl-4-(4-methoxyphenyl)-5-imidazolyl] disulfide III, yellow-orange powder, so pl. 209 211oC (with decomp. ). UV spectrum (C2H5OH,max, ): 210 (17900), 263 (23500), 362 (5000) nm. IR spectrum (CHCl3,max): 1558, 1611, 1650, 3100 3400 cm-1. PMR-spectrum (CD3OD, , M. D. TMS): 3,25 (c, 6H, 2xNMe), 3,93 (c, 6H, 2xOMe), 7,07 (m, 4Harene), 7,49 (m, 4Harene). An NMR spectrum13C (CD3OD): 30,0 (K) 56,7 (K) 112,0 (C), 115,0 (d), 119,0 (C) of 129.6 (d), 139,2, 149,1 (C), 163,3 (C). Mass spectrum (m/z is e polycarpia III confirmed also by the data of x-ray analysis.

Example 2. Synthesis polycarpia III.

Compound III was synthesized from commodity n-methoxyacetophenone (38) in accordance with scheme 1:

Bromination of p-methoxyacetophenone (38). Bromination of (38) was carried out by the action of CuBr2in a mixture of acetic acid and chloroform according to the method of [39] which gave pencilvania (39) in the form of needles with so pl. 65 66oC (yield 62% ). Range of TMR (d, M. D. CDCl3): 3,90 (c, 3H, OMe), to 4.41 (c, 2H, CH2Br), 6,98 (m, 2Harene), 7,98 (m, 2Harene).

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Circuit 1

Turning pencilvania (39) in hydrochloride aminoketone (40). To a solution of 343 mg (1.5 mmol) (39) in 2 ml of abs. C2H5OH was added 35 ml of a solution of an excess of methylamine in abs. C2H5OH. The mixture was stirred for a short time when 45 60oC, was poured into ice water, extracted with ether, the extract dried, evaporated to a volume of 25 ml and was barbotirovany through him dry HCl. The ether was removed, the residue was added benzene and the solvent was evaporated. To the residue was added a small amount of acetone, the residue was filtered, washed with acetone and dried. Retrieved 250 mg (83%) salt aminoketone (40) in the form of colorless fine crystals with so pl. 165 168oC. Range of the MRP, M. D. CDCl3); 2,82 (c, 3H, NHMe), 3,91 (c, 3H, OMe), 2-amino-1-methyl-4- (4-methoxyphenyl)imidazole (41). It chilled in an ice bath, a solution of 526 mg (2,84 mmol) to the hydrobromide S-ethylthiophene in 5 ml of H2O under stirring was added a chilled solution of 236 mg (of 5.89 mmol) of NaOH in 15 ml of H2O. To the resulting mixture dropwise added a solution of 615 mg (2,84 mmol) of the compound (40). The mixture is left on the ducks at room temperature. The precipitation was filtered, washed with water and dried in a vacuum desiccator. To the obtained product is added a small amount of chloroform, and boil for 3 to 5 minutes and filtered nerastvorimaya substance. Received 433 mg (75%) of imidazole (41), so pl. 208 211oC (from dioxane). Range of TMR (d, M. D. CD3OD): 3,42 (c, 3H, NMe), 3,78 (c, 3H, OMe), 6,78 (c, 1H, H-5), 6,86 (m, 2Harene), to 7.50 (m, 2Harene). Mass spectrum (m/z, Rel. int. 70 eV): 204 (M++1, 20), 203 (M+, 33), 188 (100), 160 (11), 147 (12), 146 (13), 119 (11), 118 (12), 77 (14).

The transformation of 2-aminoimidazole (41) bis[2-amino-1-methyl-4- (4-methoxyphenyl)-5-imidazolyl]the disulfide dihydrochloride III (polycarpon). To a solution of 102 mg (0.5 mmol) of the compound (41) in 4 ml of ice. The SPLA was added dropwise with stirring 102 mg (0.75 mmol) of S2Cl2. The mixture was stirred at room temperature for 5 h, diluted with 25 ml of glima deposited precipitate was separated, washed with glimm and dried under vacuum. On the economic characteristics of the compound obtained was identical natural policarpio III, obtained in example 1.

Example 3. Getting dihydrochloride of bis[2-amino-1-methyl-4 - phenyl-5-imidazolyl]disulfide 1.

Compound I was obtained from commodity acetophenone in accordance with the scheme set forth in example 2. Orange-yellow powder with so pl. 239 - 242oC (with decomp.). Range PMR (CD3OD): 3,21 (c, 6H, 2xMe), 7,39 - 7,58 (m, 10Harene.).

Example 4. Getting dihydrochloride of bis[2-amino-1-methyl-4-(4 - were)-5-imidazolyl]disulfide II.

Compound II was obtained from commodity n-methylacetophenone in accordance with the scheme set forth in example 2. Orange-yellow powder with so pl. 209 212oC (with decomp. ). Range PMR (CD3OD): 2,47 (c, 6H, 2xMe), 3,24 (c, 6H, 2xNMe), 7,34 (m, 4Harene.), 7,42 (m, 4Harene.).

Example 5. Getting dihydrochloride of bis[2-amino-1-methyl-4- (4-ethoxyphenyl)-5-imidazolyl]disulfide IV.

Compound IV was obtained from commodity n-ethoxyacetophenone in accordance with the scheme set forth in example 2. Orange-yellow powder with so pl. 202 205oC (with decomp.). Range PMR (CD3OD): 1,45 (t, J 7,1 Hz, 6N, 2xCH2CH3), 3,26 (s, 6N, 2xNMe), 4,16 (K, J 7,1 Hz, 4H, 2xCH2CH3),? 7.04 baby mortality ( m, 4Harene.), of 7.48 (m, 4Harene.).

Example 6. Getting dihydrochloride of bis[2-amino is of tienna in accordance with the scheme, described in example 2. Orange-yellow powder with so pl. 222 - 225oC (with decomp.). Range PMR (CD3OD): 3,31 (c, 6H, 2xNMe), 7,51 (m, 4Harene.), 7,56 (m, 4Harene.).

Example 7. Getting dihydrochloride of bis[2-amino-1-methyl-4- (3,4-acid)-5-imidazolyl]disulfide VI.

Compound VI was obtained from commodity 4,5-dimethoxyacetophenone in accordance with the scheme set forth in example 2. Orange-yellow powder with so pl. 247 251oC (with decomp.). Range PMR (CD3OD): 3,25 (c, 6H, 2xNMe), 3,92 (c, 6H, 2xOMe), 3,97 (c, 6H, 2xOMe), 7,12 (m, 6Narene.).

Example 8. Getting dihydrochloride of bis[2-amino-4-(4 - methoxyphenyl)-1-ethyl-5-imidazolyl]disulfide VII.

Compound VII was obtained from commodity n-methoxyacetophenone in accordance with the scheme set forth in example 2, except that at the stage of conversion pencilvania (39) in the substituted a-aminoketone on intermediate (39) operated with a solution of an excess of ethylamine in abs. C2H5OH that gave homologous connection (40) aminoketone with N-ethyl group. Orange-yellow powder with so pl. 205 209oC (with decomp.). Range PMR (CD3OD): a 1.25 (t, J 7,1 Hz, 6N, 2xCH2CH3, 3,67 (K, J 7,1 Hz, 4H, 2xCH2CH3), 3,92 (c, 6H, 2xOMe), 7,07 (m, 4Harene.), to 7.50 (m, 4Harene.).

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Scheme 2

Conversion of thiazole (42) bis[2-amino-4-(4-methoxyphenyl)-5 - thiazolyl] the disulfide dihydrochloride VIII. To a solution of 206 mg (1.0 mmol) of thiazole (42) in 2 ml of SPLA added dropwise with stirring to 210 mg (1.5 mmol) of S2Cl2. The mixture was stirred for 30 min at room temperature, and then added 3 ml of dioxane. The precipitated oil with grinding partially secretaryshall. The mixture was filtered, the filtrate was discarded and the residue was dissolved in 3 ml of methanol. The solution was filtered and the product was planted by dilution of the filtrate with 40 ml of ether. Received 229 mg (42%) salt 2-aminotetraline VII in the form of bright yellow fine powder with so pl. 213 217oC (with decomp.). Range PMR (CD3OD, ,, M. D. TMS): 3,93 (c, 6H, 2xOMe), 7,13 (m, enyl)-5-thiazolyl]disulfide IX.

Compound IX was obtained from commodity 3,4-dimethoxyacetophenone in accordance with the scheme set forth in example 9. Bright yellow powder with so pl. 238 241oC (with decomp.). Range PMR (CD3OD): 3,92 (c, 6H, 2xOMe), 3,94 (c, 6H, 2xOMe), 7,08 7,22 (m, 6Narene.).

Example 11. Getting dihydrochloride of bis[2-amino-4-(4 - forefeel)-5-thiazolyl/disulfide X.

Compound X was obtained from commodity 4-fortetienne in accordance with the scheme set forth in example 9. Bright yellow powder with so pl. 223 227oC (with decomp. ). Range PMR (CD3OD): to 7.32 (t, J 8,4 Hz, 4H, HNarene.), 7,66 (DD, J 8.4 and 5.0 Hz, 4H, HNarene.).

Study of the biological activity of compounds I-X.

The biological activity of the claimed compounds was investigated in different experimental models.

The study of cytostatic and cytotoxic activity in vitro.

Cytostatic and cytotoxic effect of compounds I-X in vitro against different lines of tumor cells was studied in the Department of synthesis and chemistry of medicinal substances of the National cancer Institute in Bethesda, Maryland, USA). Evaluation of cytotoxic (EID50) and cytotoxic (LD50) the action of the claimed compounds was carried out in the x compounds were tested at a minimum of 5 concentrations obtained by 10-fold dilutions. Test duration was 48 hours. The results of these studies are summarized in table.1. The most active among all investigated compounds was the connection I, and the least active compound X. Abnormally high cytotoxic activity was shown in the cells of cancer of the Central nervous system (line SNB-75) compound III (EID500.02 μg/ml) and in relation to lung cancer (line NOR-92) compound VII (EID500,03 µg/ml). Very high cytotoxic activity (EID50< 0.5 μg/ml) showed: compound VI in relation to cell leukemia line (CCRF-CEM) and melanoma (line UACC-257) and the compound IX in relation to cell lung cancer (line NOR-92) and prostate cancer (line RB). High cytotoxic activity (EID501.0 microgram/ml) showed several of the investigated compounds: compound I against leukemia cells (lines CCRF-CEM, HL-60(TB), MOLT-4, RPMI-8226), lung cancer (lines NORD-92, NCI-H522, KXFL 529 and DMS 114), colorectal cancer (lines COLO 205 and SW-620), cancer of the Central nervous system (lines SNB-78 and XF 498), melanoma (line LOX IMVI, M14, M19-MEL, SK-MEL-5, UACC-257) and breast cancer lines MDA-MB-435 and MDA-N); compound II in relation to cell leukemia line (RPMI-8226), lung cancer line NCI-H522 and LXFL 529), Akamai (line CCRF-CEM), lung cancer (lines NORD-92, NCI-H522 and LXFL 529) and rectal cancer (lines HCT-116 and SW-620); compound IV in relation to cell leukemia line (CCRF-CEM) and breast cancer line MDA-MB-231/ATCC); connection V in relation to cell leukemia line HL-60(TB) and RPMI-8226), lung cancer line NCI-H23 and NCI-H522), colon cancer line HCT-15), cancer of the Central nervous system (line SF-539), cancer of the ovary (line OVCAR-4) and breast cancer lines MDA-MB-231/ATCC and MDA-N); compound VI in relation to cell leukemia line MOLT-4) and ovarian cancer (line OVCAR-8); compound VII in relation to cell leukemia line (CCRF-CEM) and the compound IX in the cells of cancer of the rectum (line KM) and prostate cancer (line DHM and WMF). The level of cytotoxic activity (EID50) of the compounds studied in relation to other lines of tumor cells ranged mostly in the range of 1.0 to 2.0 µg/ml, there was also quite high.

the 50% Cytotoxic activity (LD50) none of the investigated compounds were not evident at a dose lower than 2.5 µg/ml. the Most active cytotoxins were the compounds I, II, III and VI. Values (LD50these compounds against certain lines of tumor cells fluctuated in the range of 2.5 to 3.0 µg/ml Thus, compound I showed a high XF498), melanoma (line LOX IMVI, M14 and SK-MEL-5) and breast cancer lines MDA-MB-435 and MDA-N); compound II in relation to cell lung cancer line NCI-H522), cancer of the Central nervous system (line XF498) and ovarian cancer (line OVCAR-5); compound III in relation to cell lung cancer line NCI-H522) and compound VI in respect of the melanoma cell line UACC-257).

A high level of cytotoxic activity values (LD50fluctuated in the range of 3.0 to 5.0 µg/ml) showed the studied compounds against a large number of lines of tumor cells. So, the connection I was active against cell leukemia line MOLT-4), lung cancer (lines NORD-02, NCI-H23, LXFL529 and DMS114), colorectal cancer (line COLO205), cancer of the Central nervous system (line SNB-78) and melanoma (lines M19-MEL and SK-MEL-28); compound II in the cells of cancer of the rectum (line SW-620) and melanoma (line M14); compound III in relation to cell lung cancer (lines NORD-92 and LXFL529), colorectal cancer (lines HCT-116 and SW-620), melanoma (SK-MEL-5), kidney cancer (line RXF-393) and breast cancer line MDA-N); compound IV in relation to cell leukemia line HL-60(TB)), lung cancer line NCI-H23), colorectal cancer (lines SW-620), cancer of the Central nervous system (line SF-539), melanoma (line M14, SK-MEL-5), and UACC-257), ovarian cancer (leukemia line (line HL-60(TB) and RPMI-8226), lung cancer (lines NORD-92, NCI-H23 and NCI-H522), colorectal cancer (lines COLO205, HCT-15, KM12 and SW620), cancer of the Central nervous system (line SF-539 and U251), melanoma (lines MALME-3M, M14, SK-MEL-28, SK-MEL-5, UACC-258), ovarian cancer (lines OVCAR-3, OVCAR-4 and OVCAR-5), kidney cancer (lines 786-O, TC-10 and UO-31) and breast cancer lines MDA-MB-231/ATCC, MDA-MB-435 and MDA-N); compound VI in relation to lung cancer line NCI-H23), colorectal cancer (lines COLO205, HCT-116, HT29, and SW-620), cancer of the Central nervous system (line SF-539), melanoma (line M14, SK-MEL-28, SK-MEL-5, UACC-62), cancer of the ovary (lines OVCAR-3, OVCAR-4 and OVCAR-8), kidney cancer (line 786-O and UO-31) and breast cancer lines MDA-MB-231/ATCC, MDA-MB-435 and MDA-N); compound VII in relation to melanoma line SK-MEL-5, UACC-257); connection IX in relation to colorectal cancer (line COLO205), cancer of the Central nervous system (line SF-539), melanoma (line M14, SK-MEL-5, UACC-257) and ovarian cancer (line OVCAR-5).

The data table. 1 shows that cell lines of melanoma panels are extremely sensitive to the effects of the inventive compounds. This kind of unusual sensitivity for compounds exhibiting cytotoxicity through inhibition of topoisomerase II. Data processing of biospehere using an appropriate computer program showed that the model response of cell lines the drives of the working systems is the explanation of the mechanism of cytotoxic action polycarpia III and other compounds of this series is, undoubtedly, considerable scientific interest.

Thus, the study of cytostatic and cytotoxic activity in vitro of the compounds I-X showed that almost all of them are promising materials for the study of the antitumor action in vivo. With the widest range of cytostatic and cytotoxic activities are, as can be seen from the table.1, compounds I, III and V

Tests conducted in the Pacific Institute of Bioorganic chemistry, far Eastern branch of Russian Academy of Sciences.

In the laboratory biospehere tibah RAS compounds I-III were tested for cytotoxic activity against murine lymphocytes, as well as hemolytic and anti-virus (anti-Sendai virus in primary cultures of mouse lymphocytes) activity in a wide concentration range (from 0.1 to 100.0 µg/ml). The investigated compounds had weak hemolytic activity and does not inhibit binding of Sendai virus with mouse lymphocytes (cytopathic effect). At the same time, they showed pronounced cytotoxic activity against murine lymphoblasts and tumor cells (Ehrlich carcinoma, ascitic form). Weak hemolytic activity of these compounds testifies in favor of predpolagaetsya membranes.

Determination of acute toxicity polycarpia III.

Acute toxicity of compound III was determined on male mice of CBA and outbred mice by the method of intraperitoneal administration. A portion polycarpia III was dissolved in dimethyl sulfoxide and brought to the analyzed concentration of the saline solution. The concentration of DMSO in the solution did not exceed 5% of the test Results is presented in table.2.

The death of the mice was observed, mainly on the second day. A characteristic response of mice by intraperitoneal administration of high doses policarpio was immobilization in a few minutes after injection (pose with stretching of muscles). Test data show that polycarpon III has a very high acute toxicity.

Study of cytotoxic activity polycarpia III against tumor cells of Ehrlich method enable [3H]-thymidine into acid-insoluble cell fraction.

Evaluation of the cytotoxic activity polycarpia III were conducted according to the method of [42] a Suspension of tumor cells (1x105cells per well) were incubated for 18 hours at 37oC "CO2-incubator" in the wells of microplates with different doses of the substance III in 100 μl of medium 199, containing donsimoni atmosphere, consisting of 5% CO2and 95% air. 6 hours before the end of the incubation each well was made aqueous solution of [3H] -thymidine at the rate of 0.5 µci per well. Radioactivity of samples was determined in a liquid scintillation counter "Mark II" (USA). The synthesis of macromolecules in cells was assessed by the level of radioactivity and expressed in control. In this experiment it was found that the value of the 50% cytotoxic activity polycarpia III in vitro against suspension of tumor cells of Ehrlich 1.8 µg/ml.

Determination of antitumor activity of the compounds III, VI, VIII and IX in vivo.

The tests were carried out on mice BDF1inoculated intraperitoneally leukemic tumor cells L1210and P388(1x105-1x106cells per mouse), and outbred mice, which were inoculable intraperitoneally suspension of tumor cells of Ehrlich adenocarcinoma (tumor of Ehrlich's ascite form) (1x106cells per mouse). The antitumor effect was evaluated by the increase in life expectancy (UPI) compared with the control according to the formula:

UPS [(T-C)/C]x100%

where T and middle days of the death of the animals in the experimental and control groups, and to study concentration of saline. The concentration of DMSO in the solution did not exceed 5% of the test Results are summarized in table.3.

As can be seen from the data table. 3, compound III, VI, VIII and IX show moderate antitumor activity in vivo against presented here types of tumors, because UPS in excess of 25% can be considered statistically significant minimum criterion of activity. Manifestation pronounced therapeutic effect polycarpia III and related compounds is complicated in most cases, their toxicity to test animals.

Inhibitory effect polycarpia III and related compounds in respect of certain enzymes

Inhibition of reverse transcriptase

Compound III was tested as a potential inhibitor of DNA synthesis catalyzed by an RNA-dependent DNA polymerase (reverse transcriptase) virus myeloblastosis birds. As the matrix was used activated DNA, obtained by splitting Dnazol-1 by standard procedures. Acid-soluble fraction after treatment with Dnazol was 3 to 6% As can be seen from the drawing polycarpon III describes the system intensively inhibited DNA synthesis (value ID500.8 µg/ml or 3,5x10-6M).

It should be noted that the act is as primer-template RNA (oligo dT) ID50it was above 10 times. Similar effects have been noted in the literature. Thus, a known inhibitor of revertase 2', 3'-dideoxy-3'-azidothymidine in the in vitro system was EID500.3 ág/ml (10-6M) on the matrix of poly(rA)-oligo(dT) and EID5030,0 mg/ml (10-4M) on activated DNA.

Similar to the above result was obtained using reverse transcriptase isolated from rous sarcoma.

The mechanism of action polycarpia III is not yet clear. By analogy with literature data it can compete for the binding site of revertase with primer-matrix complex-type inhibitors revertase NRA polyoxomolybdate and. or to compete with substrate for binding to the enzyme, like antibiotic aphidicolin or inhibitors of the type of azidothymidine. The different sensitivity of the system to policarpio III depending on the nature of the matrix testifies in favor of the first assumption. It is also possible direct impact polycarpia III matrix-type bleomycin And (ID50regarding revertase equal to 40.0 μg/ml) or a mixed type of inhibition.

Inhibition of Na+, K+-ATPase in rat brain.

It was studied the effect of compounds I-III Na+togikagi [43] as a result, it was found, what compounds I-III are strong inhibitors of this enzyme. The values of I50(concentration of inhibitor required for 50% inhibition of enzyme activity) for compounds I-III, form 4,3x10-7M, 5,2x10-7M and 5,h-7M, respectively. Complete inhibition of ATP-Noah reaction occurs at a concentration of about 5 inhibitors,h-5M. it Should be noted that the degree of inhibition of enzyme activity depends on its concentration. Compounds I-III are irreversible inhibitors of Na+, K+-ATPase: 10 50-fold dilution of the enzyme treated by them does not diminish the inhibitory effect of compounds (table.4). Inhibition of Na+, K+-ATPase activity depends on the time of the inactivated enzyme with the inhibitor. Within the first five minutes the reaction of interaction of the enzyme with inhibitors I-III obeys pseudo-first-order constants modification (K') is equal to 0,070, to 0.055 and to 0.060 min-1μm-1respectively.

Inhibition reaches a constant value within 15 minutes Inhibitory effect of compounds I-III depends on the pH of the environment. The maximum expression inhibitory effect is observed at pH values ranging from 8.0 to 9.0. Na2ATP (substrate of Na substrate 5.0 mm.

Thus, the above results of the study of biological activity of the inventive compounds show that these compounds have, as a rule, high cytostatic and cytotoxic activity in vitro, show a pronounced antitumor activity in vivo and are effective inhibitors of enzymes such as reverse transcriptase and Na+, K+-ATPase.

The claimed compounds potentially suitable for creation on their basis of effective anticancer agents that may find application in medicine for the treatment of various human malignant tumors.

Hydrochloride sulfur-containing 2-aminoimidazole or 2-aminothiazolo General formula I

< / BR>
where X alkylamino, such as methylamino, and R1R2is hydrogen;

X methylaminopropyl, and R1alkyl, for example methyl, and R2is hydrogen;

X methylaminopropyl, and R1alkoxygroup, for example methoxy, and R2hydrogen;

X methylaminopropyl, and R1ethoxypropan and R2hydrogen;

X methylaminopropyl, and R1halogen, for example chlorine, and R2is hydrogen;

X methylaminopropyl, and R1R2alkoxygroup, for example methoxy;

X Agrippa, for example methoxy, and R2is hydrogen;

X is sulfur, and R1R2alkoxygroup, for example methoxy;

X is sulfur, and R1halogen, for example fluorine, and R2hydrogen

with cytotoxic and antitumor activity and exhibiting an inhibitory effect on certain enzymes.

 

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