Membrane mechanisms of the effect of alkylating cytostatic agents

The data bearing witness to realization of the cytostatic activity of alkylating agents by means of interaction with cell membrane receptors was obtained. Alkylating agents block receptors of the polyphosphoinositol system. It is shown that chloralkylamines inhibiting sites coincide with M-++cholinomimetics and alpha-adrenergic receptors, inhibitors of H-histamine receptors, cyclo- and lypoxigenase inhibitors. Such likeness is determined by identical structures of the molecule active part, and quantum-chemical calculations.     

On the biochemical background of cimetidine's anti-ulcerogenic effect in the rat.

It was published earlier that during gastric ulceration an elevation in the antral and fundic mucosal cAMP/cGMP ratios can be encountered in rats. This phenomenon was interpreted as a probable sign of the reparative, antiulcerogenic processes. This assumption received further evidence when the experimental animals were treated with prostacyclin. According to the present investigations the H2-receptor blocker cimetidine, in such small dose which does not interfere with the gastric acid secretion, evoked the same cAMP/cGMP ratio elevating effect. On the basis of the performed investigations the questions raised, namely: Does the cytoprotective (anti-ulcerogenic) effect of different drugs have a common molecular basis in the (rat) gastric mucosa? If so, what is the possible role of the mucosal cyclic nucleotides in this process? Further studies are being needed in this field to clarify all details.     

The antiradiation effect of the cytostatic ribamidil and the possible mechanisms of its action

Ribamidil has a cytostatic effect on bone marrow cells and a radioprotective therapeutic action. Ribamidil favors CFUs proliferation and does not interfere with the process of CFUs migration.     

Macrophage cytostatic effect on Trypanosoma musculi involves an L-arginine-dependent mechanism

Peritoneal macrophages from mice infected with an extracellular parasite, Trypanosoma musculi were effective in inhibiting parasite proliferation in vitro. This trypanostatic activity could be suppressed by NG monomethyl-L-arginine (NGMMA), a specific inhibitor of a biochemical pathway synthesizing L-citrulline and inorganic nitrogen oxides from L-arginine. Macrophages exerted this in vitro antiproliferative effect from the 10th day of infection on and this activity was maximum around 14th day of infection. Nitrite production paralleled development of macrophage trypanostatic activity. Macrophages collected from BCG-infected mice or treated with IFN-gamma in vitro also exerted a trypanostatic activity which was suppressed by NGMMA. A trypanostatic activity suppressed by NGMMA was also exerted by splenic macrophages from T. musculi-infected mice. Trypanostatic activity of IFN-gamma-treated macrophages was reduced by addition of anti-TNF-alpha showing the participation of TNF-alpha in IFN-gamma-mediated macrophage trypanostatic activity. Nitric oxide (NO) gas inhibited T. musculi proliferation. Addition of excess iron reversed the trypanostatic effect of both macrophages and NO gas. All these data showed that, as reported for a broad spectrum of microorganisms, activated macrophages displayed an antimicrobial effect on trypanosomes through the L-arginine: NO pathway that could participate in controlling infection in T. musculi-infected mice before appearance of antibody-dependent mechanisms. NO production by activated macrophages could trigger iron loss from critical target enzymes in trypanosomes.     

Activation of spinach chloroplast glyceraldehyde 3-phosphate dehydrogenase: effect of glycerate 1,3-bisphosphate

Spinach (Spinacia oleracea L.) chloroplast NAD(P)-dependent glyceraldehyde 3-phosphate dehydrogenase (NAD(P)-GAPDH; EC 1.2.1.13) was purified. The association state of the protein was monitored by fast protein liquid chromatography-Superose 12 gel filtration. Protein chromatographed in the presence of NADP⁺ and dithiothreitol consisted of highly NADPH-active protomers of 160 kDa; otherwise, it always consisted of a 600-kDa oligomer (regulatory form) favoured by the addition of NAD⁺ in buffers and with low NADPH-dependent activity (ratio of activities with NADPH versus NADH of 0.2–0.4). Glycerate 1,3-bisphosphate (BPGA) was prepared enzymatically using rabbit-muscle NAD-GAPDH, and purified. Among known modulators of spinach NAD(P)-GAPDH, BPGA is the most effective on a molar basis in stimulating NADPH-activity of dark chloroplast extracts and purified NAD(P)-GAPDH (activation constant, K _a= 12 M). It also causes the enzyme to dissociate into 160-kDa protomers. The K _m of BPGA both with NADPH or NADH as coenzyme is 4–7 M. NAD⁺ and NADH are inhibitory to the activation process induced by BPGA. This compound, together with NADP(H) and ATP belongs to a group of substrate-modifiers of the NADPH-activity and conformational state of spinach NAD(P)-GAPDH, all characterized by K _a values three- to tenfold higher than the K _m. Since NADP(H) is largely converted to NAD(H) in darkened chloroplasts Heineke et al. 1991, Plant Physiol. 95, 1131–1137, it is proposed that NAD⁺ promotes NAD(P)-GAPDH association into a regulatory conformer with low NADPH-activity during dark deactivation. The process is reversed in the light by BPGA and other substrate-modifiers whose concentration increases during photosynthesis, in addition to reduced thioredoxin.Abbreviations BPGA glycerate 1,3-bisphosphate - Chl chlorophyll - DTT dithiothreitol - FPLC fast protein liquid chromatography - NAD(P)-GAPDH glyceraldehyde 3-phosphate dehydrogenase, NAD(P)-dependent - 3-PGA glyerate 3-phosphate - PGK phosphoglycerate kinase - Prt protein - Tricine N-tris (hydroxymethyl) methyl-glycine This work was supported by grants from the Ministero dell'Università e della Ricerca Scientifica e Technologica in years 1990–1991. We are grateful to Dr. G. Branlant (Laboratoire d'Enzymologie et de Génie Génétique, Vandoeuvre les Nancy, France) for introducing us to the BPGA purification procedure.     

Cytotoxic and cytostatic effect of avermectines on tumor cells in vitro]

Effect of natural avermectin complex (aversectin C) and separate avermectins A1, A2, B1 and B2 in the cell culture of murine myeloma Ns/o, Erlich carcinoma ascites and human larynx carcinoma Hep-2 was investigated. It was shown that aversectin C within the concentrations of 0.1 to 1.0 mcg/ml inhibited proliferation of tumor cells and induced their death. Proliferation inhibition was due to the delay of the cells cycle start (lag-phase prolongation) and blocking of mitotic cycle. Ns/o cells death had apoptosis signs: chromatin condensation and fragmentation, DNA fragmentation. It was demonstrated that only avermectin A1 has cytotoxic activity within the concentrations used, avermectins A2 and B2 had cytostatic activity, avermectin B1 showed no activity under the experimental conditions.     

The effect of glutathione depletion on sister-chromatid exchange induction by cytostatic drugs

Using sister-chromatid exchanges (SCEs) as an indicator for DNA damage, we investigated the role of glutathione (GSH) as a determinant of cellular sensitivity to the DNA-damaging effects of the cytostatic drugs adriamycin (AM) and cyclophosphamide (CP). Exposure of V79 cells to buthionine sulfoximine (BSO) resulted in a complete depletion of cellular GSH content without toxicity and without increasing the SCE frequency. Subsequent 3-h treatment of GSH-depleted cells with AM or S9-mix-activated CP caused a potentiation of SCE induction. In Chinese hamster ovary (CHO) cells, which showed a higher GSH level compared to V79 cells, BSO treatment led to a depletion of GSH to about 5% of the control and increased SCE induction by AM and CP. Compared to V79 cells, the effect of AM on SCE frequencies was less distinct in CHO cells, while CP exerted a similar effect in both cell lines. Pretreatment of V79 cells with GSH increased the cellular GSH content, but had no effect on the induction of SCEs by AM, and pretreatment with cysteine influenced neither GSH levels nor SCE induction by AM. The study shows that SCEs are a suitable indicator for testing the modulation of of drug genotoxicity by GSH. The importance of different GSH contents of cell lines for their response to mutagens is discussed.