Peter Rauko

Effects of flavonoids on cisplatin-induced apoptosis of HL-60 and L1210 leukemia cells

Effects of three flavonoids, quercetin (QU), galangin (GA), and chrysin (ChR) on cisplatin (cis-Pt)-induced apoptosis of human promyelocytic leukemia HL-60 cells and murine leukemia L1210 cells were investigated. The quantitative analysis of apoptotic DNA fragmentation was used to show that preincubation of cells with flavonoids can influence cis-Pt-induced apoptosis in different way. ChR had no effect, QU enhanced, and GA reduced apoptotic DNA fragmentation. It is also shown that combined treatment with QU and cis-Pt showed synergistic effect, however, GA combined with cis-Pt exhibited antagonism on cytotoxicity in L1210 murine leukemia cells. We assume that tested flavonoids affect the important biological activities connected with cancer chemotherapy and chemoprevention as they differently modulated the sensitivity of cells to cis-Pt treatment. QU is presented as pro-apoptotic agent and GA as agent with anti-apoptotic potential.

Dual activity of triterpenoids: apoptotic versus antidifferentiation effects

Triterpenoids are natural, biologically active compounds extracted from many plants. They possess antiinflammatory, anticancer, and antioxidant properties. In the report presented, antiproliferative effects and leukemia cell growth and apoptosis modulating activities of ursolic acid (UA) and oleanolic acid (OA) were investigated. Both triterpenoids are inhibitors of leukemia cell growth and inductors of apoptosis. However, when applied in combination with anthracycline antitumor antibiotic doxorubicin (Dox), UA and OA diversely modulate therapeutic efficacy of Dox, due to different antioxidant activities. Compare to OA showing synergism/additive effect with Dox, UA (stronger antioxidant) acts antagonistically and reduces leukemia cell growth inhibiting and differentiation effects induced by Dox. In conclusion, these findings suggest that although triterpenoids UA and OA can induce apoptosis, their antioxidant activities can interfere with the therapeutic effect of antitumor antibiotic Dox which mechanism of action is attributed to the production of reactive oxygen species.

Enhanced effects of adriamycin by combination with a new ribonucleotide reductase inhibitor, trimidox, in murine leukemia

Ribonucleotide reductase is the rate limiting enzyme of de novo DNA synthesis; its activity is significantly increased in tumor cells related to the proliferation rate. Therefore the enzyme is considered to be an excellent target for cancer chemotherapy. In the present study we tested the in vitro and in vivo antitumor effects of a drug combination using trimidox (3,4,5-trihydroxybenzamidoxime), a novel inhibitor of ribonucleotide reductase with adriamycin, a widely used anticancer drug. This combination was selected because adriamycin generates free radicals being responsible for cardiotoxic side effects; trimidox has been shown to be a good free radical scavenger. The in vitro cytotoxic effect of the drug combination was examined in L1210 mouse leukemia cells employing a MTT chemosensitivity assay. Incubation of these cells with adriamycin and trimidox together yielded less than additive cytotoxic effects compared to either drug alone. These effects were not caused by the involvement of p-glycoprotein mediated drug efflux. However, when the effect of trimidox and adriamycin in combination was examined in L1210 leukemia bearing mice antitumor effects of adriamycin could be enhanced by the presence of trimidox. Our data indicate, that the in vivo combination of adriamycin together with trimidox might be beneficial for the treatment of malignancies.

Differential modulation of cisplatin and doxorubicin efficacies in leukemia cells by flavonoids

We studied the ability of five structurally related flavonoids (quercetin (QU), luteolin (LU), apigenin (AP), galangin (GA) and chrysin (ChR)) to modulate chemotherapeutic efficacy and affect chemotherapy-induced apoptosis and cell cycle arrest of murine leukemia L1210 cells. We found that QU and LU potentiated cytotoxicity of cisplatin, however the other flavonoids (AP, GA and ChR) decreased its cytotoxic effect. We proved that QU and LU positively modulated efficacy of cisplatin due to increasing of cisplatin-induced apoptotic cell death. In combination with doxorubicin all tested flavonoids decreased doxorubicin cytotoxicity and caused cells to shift from G2/M to S phase of the cell cycle. The findings that flavonoids can differentially modulate the efficacy of chemotherapeutics through interfering with chemotherapy-induced apoptosis or cell cycle arrest further improve our understanding of possible impacts of some dietary agents on chemotherapeutic intervention.

Antitumor activity of benzamide riboside and its combination with cisplatin and staurosporine

Benzamide riboside (BR), a new synthetic nucleoside analogue, has demonstrated a potent cytotoxic activity in murine leukemia in vitro. The purpose of the present investigation was to examine the antitumor activity of BR in mice bearing leukemia L1210. The results revealed that BR possesses a potent antitumor activity in vivo. It increases life-span of mice with leukemia. Synergistic cytotoxicity of BR with select DNA damaging agents, cisplatin (cis-Pt) and staurosporine (STP) was examined in MTT chemosensitivity assay, FACS analyses and apoptotic DNA fragmentation on L1210 cells in culture. A simultaneous treatment of leukemia L1210 cells with the combination of BR and STP resulted in synergistic cytotoxicity that correlated with increased apoptotic activity in those cells. On the other hand, treatment of L1210 cells with combination of BR and cis-Pt resulted in antagonistic cytotoxic effect. Finally, to elucidate the synergistic effect of BR and STP in inducing apoptosis, the attention was directed to the activation of cell death processes through various cell cycle signals. This is the first report describing in vivo antitumor activity of BR and its utilization in combination chemotherapy.

Fungal chitin–glucan derivatives exert protective or damaging activity on plasmid DNA

Water-soluble derivatives of the chitin-glucan (Ch-G) complex isolated from the fungal mycelium of the industrial strain of Aspergillus niger have been previously shown to possess potent antimutagenic protective activity in vivo. Their direct action on DNA has not been yet evaluated. Using carboxymethylation, sulfoethylation and subsequent ultrasonic treatment, lower molecular weight water-soluble derivatives were obtained from the crude fungal Ch-G. The biological effects of the prepared compounds were evaluated in direct interaction on plasmid DNA in vitro. Monitoring of electrophoretic mobility of different conformers of plasmid DNA implied that carboxymethyl chitin-glucan (CM-Ch-G) induced single- and double-strand breaks into supercoiled DNA in a concentration-dependent manner. On the other hand, sulfoethyl chitin-glucan (SE-Ch-G) alone did not induce any DNA breaks in plasmid DNA. However, process of DNA damaging induced by free-radical oxidation initiated with Fe(2+) was inhibited, while the process of DNA breakage induced by H(2)O(2) was increased in the presence of SE-Ch-G.

Antigenotoxic potential of glucomannan on four model test systems

Antimutagenic, anticlastogenic, and bioprotective effect of polysaccharide glucomannan (GM) isolated fromCandida utilis was evaluated in four model test systems. The antimutagenic effect of GM against 9-aminoacridine (9-AA)- and sodium azide (NaN3)-induced mutagenicity was revealed in theSalmonella typhimurium strains TA97 and TA100, respectively. GM showed anticlastogenic effect against N-nitroso-N′-methylurea (NMU) induced chromosome aberrations in theVicia sativa assay. The bioprotective effect of GM co-treated with methyl-methane-sulphonate (MMS) was also established inChlamydomonas reinhardtii repair deficient strainsuvs10 anduvs14. The statistically significant antimutagenic potential of GM was not proved against 4-nitro-quinoline-1-oxide (4-NQO)-induced mutagenicity inSaccharomyces cerevisiae D7 assay. It may be due to bioprotectivity of α-mannan and β-glucan, which are integral part ofS. cerevisiae cell walls. Due to the good water solubility, low molecular weight (30 kDa), antimutagenic/anticlastogenic, and bioprotective activity against chemical compounds differing in mode of action, GM appears to be a promising natural protective (antimutagenic) agent.

Comparative DNA protectivity and antimutagenicity studies using DNA-topology and Ames assays

Two experimental techniques, the DNA-topology assay and the Ames assay, were proved to be suitable for monitoring compounds with a genotoxic potential and/or with an antimutagenic effect. Both procedures were used in assaying the acid-mine water (AMW) containing toxic metals and sulfoethyl chitin-glucan (SE-Ch-G), a derivative of chitin-glucan, in which bioprotective activities were detected earlier. It was shown that after toxic metal concentrations were decreased due to AMW dilution to the limits that correspond with those set by the Slovak Technical Norm (STN) for drinking water, AMW was not genotoxic in the Ames assay. As it is possible to detect any single-strand DNA (ssDNA) break in the DNA-topology assay, the SE-Ch-G protective effect against the ssDNA breaks induced by Fe(2+) in the DNA-topology assay was recorded. SE-Ch-G exhibited the antimutagenic potential after its application simultaneously with diagnostic mutagens in the Ames assay. These results demonstrate the complementarity of both experimental systems.

Main targets of tetraaza macrocyclic copper complex on L1210 murine leukemia cells

Several metal complex agents have already been introduced into clinical tumor therapy and others are subject of antitumor studies. In this study we focused on the tetraaza macrocyclic copper complex (Cu(TAAB)Cl(2)). We studied the influence of the substance on cell growth, cell cycle, membrane integrity, necrosis, apotosis and glutathione level on the leukemic cell line L1210 in 1-day (22 h) and 3-day (72 h) experiments. The metal complex shows a dose-dependent antiproliferative effect, without affecting cell cycle phases. The present results confirm that copper complex can damage plasmatic membranes and trigger apoptosis, and that after treatment of leukemic cells with the copper complex, glutathione levels were increased.

Induction of cytotoxicity and ssDNA breaks by 9-bromo-5-morpholino-tetrazolo[1,5-c]quinazoline in tumor cells cultured in vitro

9-Bromo-5-morpholino-tetrazolo[1,5-c]quinazoline (BMTQ) acted cytotoxically on murine leukemia cell line L1210 and human colon carcinoma cells Caco-2. We found the two highest concentrations of BMTQ (149.2 and 74.6 microM) induced an acute cytotoxic effect, however other tested concentrations (<74.6 microM) manifested a concentration/dependent and time/dependent cytotoxic effect. The sensitivity of murine leukemia cells L1210 and human colon carcinoma cells Caco-2 was expressed in the same order. The cytotoxicity of BMTQ was not accompanied by changes of the cell cycle profile. Following the cytotoxicity-related effects of BMTQ we observed the induction of ssDNA breaks after BMTQ treatment. All the concentrations of BMTQ increased the level of ssDNA breaks 1.3-2.9 times (after 2 h of treatment) and 1.6-2.8 times (after 4 h of treatment) in Caco-2 cells compared to the control. No apoptotic DNA fragmentation induced by BMTQ in Caco-2 cells was recorded.