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Novel 2,3-seco-triterpenic amides were prepared by the interaction of the chloride of 2,3-seco-l-cyano-19beta,28-epoxy-18alpha-oleane-3-oic acid with primary amines and synthetic and biogenic amino acids. A cytotoxic triterpenic conjugate with a residue of the ethyl ester of beta-alanine was found among the synthesized nitrogen-containing derivatives. Treatment with this conjugate in a concentration of 100 muM resulted in the 45.5% survival of melanoma cells in the medium.

We studied the ability of lymphokine-activated killer cells to lyse A549 human non-small cell lung cancer cells after preincubation with cisplatin. Lymphokine-activated killer cells obtained after incubation of human blood lymphocytes with interleukin-2 were characterized by high expression of natural killer cell antigens and activation molecules. Lymphokine-activated killer cells produced potent cytotoxic effect on intact A549 cells and lysed tumor cells survived after treatment with cisplatin in concentrations of IC50 and IC30. Cisplatin in noncytotoxic concentrations did not increase lytic activity of lymphokine-activated killer cells.

We compared cytotoxic activity of blood mononuclear leukocytes from healthy donors and lymphokine-activated killer cells generated from them towards tumor and normal cells. Lymphokine-activated killer cells exhibited higher (in comparison with blood mononuclear leukocytes) killer activity towards tumor cells. Lymphokine-activated cells and mononuclear leukocytes had no lytic effect on non-transformed eukaryotic cells. Hence, we demonstrated selective cytotoxic activity of effector cells (lymphokine-activated killers) towards tumor cells of different origin (but not normal cells).

The authors discuss the present-day state of search for antitumoral compounds at Blokhin Russian Oncological Research Center and promising approaches including computer technologies as means of search for new anticancerous targets and drugs.

New cyclopropane derivatives of betulin were synthesized by attachment of dichlorocarbenes or dibromocarbenes to the double bond of betulin diacetate, followed by the deprotection of hydroxyl groups. The betulin cyclopropane derivative was obtained from 20,29-dihydro-20,29-dichloromethylenebetulin by treatment with lithium in tert-butanol. These compounds were converted into the corresponding derivatives of betulonic acid by oxidation with chromium trioxide. The reduction of oxo group with sodium borohydride led to the corresponding derivatives of betulinic acid. 20,29-Dihydro-20,29-dichloromethylenebetulinic acid proved to be the most cytotoxic toward human melanoma of the Colo 38 and Bro lines and human ovarian carcinoma of the CaOv line (IC50 10 microM). 20,29-Dihydro-20,29-dibromomethylenebetulinic acid inhibited the growth of the Bro melanoma cell line and the CaOv carcinoma cell line practically by 50% at a concentration of 10 microM. The other derivatives of betulinic and betulonic acids were active toward all of the three cancer cell lines at concentrations higher than 10 microM. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2005, vol. 31, no. 3; see also http://www.maik.ru.

Two methods of obtaining of 3 alpha-betulinic acid and related compounds from their 3 beta-epimers were studied: the reaction of bimolecular substitution and the stereoselective reduction of 3-ketoderivatives. The substitution of acyloxy by formyloxy group in 3-O-tosyllupeol or of the betulin hydroxyl by benzoyloxy group resulted only in delta 2, 3-elimination products, with none of the expected products of bimolecular substitution being found. The catalytic hydrogenation of betulonic acid over Raney nickel resulted only in reduction of the isopropenyl double bond, whereas the use of 5% Ru/C gave a 60:40 mixture of epimers of dihydrobetulinic acid. Practically the same mixture of betulinic acid epimers was obtained when reducing betulonic acid with L-Selectride. The cytotoxic activity of 3 alpha-betulinic acid increased toward melanoma Bro cells and decreased toward melanoma MS cells.

Milestones of the Russian antitumor chemotherapy development inseparably associated with the names of N.N. Blokhin, L.F. Larionov and V.I. Astrakhan are reviewed. Since the early 50s of the last century, more than 7,000 generic and synthetic compounds have been studied by the three generations of chemotherapy researchers. 17 antitumor drugs have been developed and passed for clinical trials, 11 of them have been or still are being used in clinic.

A synthesized analog of myelopeptide HP-2-->M[symbol: see text]-2 (Leu-Val-Val-Tyr-Pro-Trp) caused a significant (60-80%) and prolonged inhibition of s.c. grafted tumors P388, Ca-755, B-16 and sarcoma 180 in isogenic mice but did not affect the growth of tumor B-16 in nude mice. Nor did it influence proliferative activity or viability of cultured human tumor cells. The best results were obtained with s.c. injections of 0.5-2 mg/kg HP-2-->M[symbol: see text]-2, twice or trice a day, at 96 hr intervals. No symptoms of severe poisoning were registered at doses of HP-2-->M[symbol: see text]-2 100 times the therapeutic one. A pharmacokinetic study in mice revealed prolonged circulation of HP-2-->M[symbol: see text]-2 in blood and a high affinity for the bone marrow (t 1/2 (130.1 hr and 431.6 hr, respectively). HP-2-->M[symbol: see text]-2 restored in vitro the ascites P388-suppressed cytotoxicity of murine T-lymphocytes. HP-2-->M[symbol: see text]-2 is regarded as a candidate for clinical studies of its potential of immunocorrection in cancer patients suffering T-lymphocyte immunity disturbances.

3,5-Cyclic phosphates and phosphoramides of 6-halogenated glucofuranoses were synthesized via interaction of 3,5,6-bicyclophosphites of 1,2-O-alkylidene-alpha-D-glucofuranoses with halogens (followed by treatment with nucleophilic reagents) and N-chloroamines. 3,5-Cyclic trans-dibutylphosphoramides of 6-chloro-6-deoxy-1,2-O-isopropylidene- and 6-chloro-6-deoxy-(R)-(2,2,2)-trichloroethylidene)-alpha-D-glucofuranoses were shown to possess antiproliferative activity against CaOv human ovarian carcinoma cells in vitro (CE50 of approximately 10(-5) M). Cyclic trans-dibutylphosphoramide of 6-chloro-6-deoxy-1,2,-O-isopropylidene-alpha-D-glucofuranose also displayed marked antitumor effect on P-388 transplantable murine leukemia in vivo (the maximum increase in life span of 100% was reached at the quintuple injection of 100 mg/kg daily).

The several conjugates of aluminium and cobalt complexes of phthalocyanines with human alpha-fetoprotein have been synthesized. Their cytotoxic activity against tumor cells and human peripheral blood lymphocytes was studied. The experimental data demonstrate that the cytotoxic activity of alpha-fetoprotein-phthalocyanine conjugates against three types of tumor cells of various origin is much higher (for aluminium and cobalt complexes more than 1000 and 50 times, respectively) in comparison with phthalocyanines themselves. The application of phthalocyanines as conjugates with alpha-fetoprotein makes it possible to markedly enhance the selective toxicity of phthalocyanines against human tumor cells.

The effect of cisplatin and the new drug cycloplatam (amine (cyclopentylamine)-S-(-)-malatoplatinum (II)) on protein kinase C (PKC) activity and Ca(2+)-dependent binding of PKC to T lymphocytes membranes was studied in vivo and in vitro. At first, the effect of the drugs on PKC activity of intact and activated lymphocytes was studied in vivo. In 48 hours after intraperitoneal injection of mice with therapeutic doses of the drugs, PKC activity of intact lymphocytes was differentially affected. Cisplatin did not practically alter the enzyme activity, whereas cycloplatam inhibited the PKC activity by 37% versus control. In lymphocytes activated by mouse P-388 leukemia cells in vivo, the drugs caused almost complete suppression of PKC activity and Ca(2+)-dependent binding of the enzyme to the membranes. The drugs were effective in intact cells. After incubation of intact lymphocytes in vitro for 24 hours with cisplatin or cycloplatam (10(-5)M), PKC activity was increased 1.62- and 1.35-fold, respectively, versus control. Ca(2+)-dependent binding of the enzyme to the membranes was also increased 1.61- and 1.36-fold by cisplatin and cycloplatam, respectively. On the contrary, at 10(-4) M concentration under similar conditions, the drugs did not affect the PKC activity of the lymphocytes. Furthermore, cycloplatam, unlike cisplatin, reduced the PKC binding to cellular membranes by 31%. The mechanisms of the drugs effects on PKC activity are suggested. The data indicate that increase or decrease of PKC activity induced by the drugs cause stimulation or depression of functional activity of T lymphocytes, respectively. Thus, the membrane-bound PKC can play the key role in initiation and development of immunomodulatory effects of cisplatin and cycloplatam.

To determine prognostically unfavourable groups of acute leukemia patients, the authors studied the in vitro accumulation of 3H-vincristine (Vcr) and adriamycin (ADR) as well as inclusion of 3H-cytosar (Ara-C) into marrow blast DNA from patients showing different effects of treatment. It was found resistant to induction chemotherapy increases with verapamil addition to culture medium (Vrp+ cells). ADR inclusion into Vrp+ cells was the same as that into Vrp- cells. The inclusion of 3H-Ara-C into S-phase cell DNA in the cells Vrp+ was 3 time that in the cells Vrp-. All the responders to cytosar treatment had Vrp- blasts. It is evident that evaluation of Vrp effect on 3H-Vcr accumulation under short-term culturing is able to indicate groups of patients with low probability of achieving complete remissions in response to standard regimens of Vcr, Ara-C and anthracyclines treatment.

Antiproliferative activities of combinations of semisynthetic plasmanyl-(N-acyl)-ethanolamine [PNAE(s)], an inhibitor of protein kinase C, with two antitumor complexes of platinum (II) [cisplatin and ammine(cyclopentylamine)-S-(-)-malatoplatinum (cycloplatam)] were investigated. The exposure of human melanoma BRO cells in culture simultaneously with cisplatin (1-10 microM) and PNAE(s) (100 microM-1 mM) in a molar ratio of 1/100 for 24 h induced a considerable decrease in the ability of these cells to incorporate [3H]thymidine into DNA. A considerable antiproliferative synergism of these agents was observed. The effect of cycloplatam/PNAE(s) combination in similar experiments was significantly different from cisplatin/PNAE(s), i.e. interaction of these agents was complex and synergism was not found.

It was shown that accumulation of adriamycin (ADR) and its impact on DNA synthesis in the blast cells of patients with hemoblastosis studied under conditions of short-term incubation correlated with the antileukemic activity of anthracyclines. The combination of the two biochemical and pharmacological indices, i.e. the high levels of ADR accumulation and inhibition of DNA synthesis, is may useful in solving the problem on inclusion of the antibiotics to the schemes of combined chemotherapy of patients with hemoblastosis.

Protein kinase C was extracted from mouse brain and partially purified by ion-exchange chromatography on a DEAE-cellulose column. Its activity was determined by incorporation of phosphate from [gamma-32P]ATP into histone H2b. The semisynthetic alkyl-phospholipid plasmanyl-(N-acyl)-ethanolamine (PNAEs) with selective antitumor activity inhibited the activity of the protein kinase in a cell-free system in the presence of phosphatidylserine, a protein kinase C activator. The inhibition was competitive with respect to phosphatidylserine, the inhibition constant being 40 microM.

The action of a new anticancer agent, the semisynthetic alkyl-phospholipid plasmanyl-(N-acyl)-ethanolamine (sPNAE), namely 1-O-octadecyl-2-oleoyl-sn-glycero-3-phospho-(N-palmitoyl)-ethanolamine, on protein kinase C (PKC) was investigated, and it was found to inhibit in a dose-dependent manner PKC isolated from mouse brain. The inhibition was competitive with respect to phosphatidylserine (K(i) = 20 microM). Lyso-PNAE, a possible cell metabolite of sPNAE, also inhibited PKC. A two-site model was used to calculate the binding affinity and the number of binding sites for phorbol ester in a culture of human melanoma BRO cells. The values of Kd, the dissociation constant, were K'd = 0.5 nM and K"d = 72 nM, whereas the values of Bmax, the number of binding sites, were B'max = 4.6 x 10(4) sites cell-1, and B"max = 2.9 x 10(5) sites cell-1. sPNAE was able to reduce the affinity of BRO cells for phorbol ester with almost no changes in the number of binding sites: K'd = 1.6 nM, K"d = 557 nM, and B'max = 4 x 10(4), B"max = 1.9 x 10(5). These data suggest that sPNAE may inhibit PKC in intact cells. Since various inhibitors of PKC may enhance the antiproliferative activity of cis-diamminedichloroplatinum(II) (cis-DDP), we investigated the effect of the combination of sPNAE and cis-DDP on the proliferation of BRO cells. sPNAE synergistically enhanced the antiproliferative activity of cis-DDP.

Methylcobalamin (10 microgram/kg bw, intramuscularly, on the 3d and 5th days after tumor transplantation) or methylcobalamin chlorpalladat -- MeCbl.PdCl3 (250 mg/kg bw in 2% starch suspension per os on the 2d and 6th days) were given to C57BI mice bearing adenocarcinoma Ca--755. On the 8th day after transplantation the methionine synthetase activity and proliferative pool of tumor cells were estimated. It has been disclosed that under the effect of MeCbl the activity of methionine synthetase rises concurrently with an increase in pool of the proliferating cells. Inhibition of tumor growth (by 80--90%) after the treatment with MeCbl.PdCl3 correlated with a decrease in the holoenzyme level in tumor cells.

A mathematical model of the folic acid cycle and of thymidylate, purines, methionine and serine biosynthetic reactions in leukemia L-1210 and Ehrlich ascite tumour cells was developed. This model was used for an analysis of biochemical criteria of different sensitivity of these tumours to methotrexate, such as differences in the rates of methotrexate transport into the cells, levels of target enzyme, dihydrofolate reductase, its affinity for the inhibitor and the capacity of "salvage" pathways of tetrahydrofolate formation. It was shown that low sensitivity of the Ehrlich ascite tumour cells to methotrexate is due to mainly a high activity of methionine synthetase, which represents a "salvage" pathway of tetrahydrofolic acid regeneration in the presence of methotrexate. The results of this analysis were used to predict a combined utilization of methotrexate and methionine synthetase inhibitor. The pretreatment of the tumour cells of the methionine synthetase inhibitor enhances the effects of methotrexate on the thymidylate and purines syntheses, thus increasing their sensitivity to this drug.

The metabolism of murine tumors sensitive and resistant to 6-theopurines and 5-fluorouracil (5-FU) was studied. A positive correlation between the tumor sensitivity to thiopurines, the level of hypoxanthine-guanine-phosphoribosyl transferase and the rates of the antimetabolite incorporation into RNA and DNA was shown. The sensitivity of tumors to 5-FU correlated with the rates of pyrimidine nucleosides and antimetabolite uptake and the time of retention of labeled 5-FU in RNA. The activites of orotate-phosphoribosyl transferase, uridine kinase, the degree of thymidylate synthetase inhibition of 5-FU and ratio of the de novo to salvage pathways of thymidylate biosynthesis are also important.

The amount of free purine and pyrimidine ribonucleotides in the spleens of mice (C57Bl and DBA/2) and in lympholeukemia cells (La and L1210), sensitive and with induced resistance to 5-fluorouracil, was determined by chromatography on a column with DEAE-cellulose. It was found that the cytidine ribonucleotide pool in the spleens of DBA/2 mice is 2 times lower as compared to C57Bl mice. The lympholeukemia cells (La and L1210) isolated from the animals also differed in their uridine nucleotide pools. The development of leukemia was accompanied by a decrease in ATP and GTP. No significant changes in the total amount of pyrimidine nucleotides under developing resistance to 5-fluororuacil were observed.

A general description of the folic acid cycle and associate reactions of thymidilate, purines, methionine and serine biosyntheses is presented as a system of differential equations. The concrete models of this reaction complex as well as methotrexate transport into the leukemia L 1210 and Ehrlich ascite tumour cells and the dynamic inhibition of the target enzyme--dihydrofolate reductase--by methotrexate have been described and investigated. The adequacy of mathematical models to the experimental data has been demonstrated.

The effect of methylcobalamin on 3H-methotrexate uptake by tumor and normal tissues of mice with mammary adenocarcinoma (Ca-755) was studied. Methylcobalamin stimulated the rate of 3H-methotrexate influx into the tumor and small intestine but did not change its influx into the spleen. The effect was dependent on the dose of methylcobalamin. Comparative analysis of the kinetic differences in 3H-methotrexate influx and efflux in tumor and susceptible host tissues revealed the optimal dose of methylcobalamin 0.01 mg/kg to improve the antitumor drug action.

The content of potassium and sodium ions in the blood and plasma of rats exposed to constant magnetic fileds (CMF) of 1000 and 4500 oersted for varying time was studied. The transport of potassium ions through membranes of red blood cells was examined in in bitro experiments using a CMF of 4500 oersted. An exposure of animals to a CMF of 1000 oersted for 1 and 24 hours did not produce significant changes in the potassium and sodium content in blood. An exposure of animals to a CMF of 4500 oersted induced certain changes in the electrolyte composition of the blood. During an hour exposure the most distinct changes were an increase in the potassium concentration in plasma and its decrease in blood. During a 3-hour exposure the most marked change was a decrease in the sodium content in blood. Repeated and prolonged chronic exposure did not bring about summation of the effects. The exposure did not influence the transport of potassium ions. The exposure did not affect the pH and hematokrit values of the blood.