Differential killing and radio-modifying effects of iodoacetate in mammalian normal and cancer cells.

To explore possible applications of iodoacetate (IA), a glycolytic inhibitor, in cancer treatment, we screened its cytotoxicity and radioprotective/sensitizing efficacy in three different mammalian cell lines; A549 (human lung carcinoma), MCF7 (human mammary cancer), a non-cancerous CHO (Chinese hamster ovary) cells and human lymphocytes. Experiments were carried out using IA concentrations ranging from 0.01 to 2.5 µg/ml, with or without 60Coγ-radiation. In the outcomes, IA was found to exhibit higher toxicity in the cancer cells, whereas it was non-toxic/marginally toxic to the non-cancerous cells. Considerably higher glucose uptake in both cancer cells lines was observed indicating higher rates of glycolysis. IA significantly inhibited glycolysis as reflected by GAPDH activity inhibition. Radiomodifying effects of IA were found to be concentration dependent in both cancerous and non-cancerous cells. The response in non-cancerous was found to be biphasic: at lower concentrations, it offered significant radioprotection; however, the protection decreased with increasing concentration. Moreover, at the highest tested concentration, marginal radiosensitization was also observed (as indicated by clonogenic assay). In both cancer cells, IA offered significant amount of radiosensitization which was considerably high at higher concentrations. Further experiments were carried out to estimate the Dose Modification Factor (DMF) to quantify and compare relative radiosensitization by IA in cancer and normal cell lines. The DMF was calculated for three different concentrations of IA, 0.5, 1, and 1.5 µg/ml, and corresponding values were found to be 1.26, 1.43, and 1.89 for A549 cancer cells, whereas for normal CHO cells, it was 1.13, 1.13, and 1.24. In conclusion, differential killing and radiosensitizing effects of IA suggest that it may have potential use as a anticancer agent and radiosensitizer in cancer therapy.

Genotoxicity of selenite in diploid yeast.

Selenite, a chemical of industrial importance and also an antimutagenic/anticarcinogenic agent, was tested for mutagenic and recombinogenic effects in 2 diploid yeast strains, Saccharomyces cerevisiae BZ 34 and D7. Selenite induced gene conversion and toxicity in BZ 34 and a variety of genetic events, viz. back-mutation, gene conversion, mitotic crossing-over, aberrant colony formation and also toxicity in the D7 strain. In both strains, the genetic effects of selenite showed a peak and a decline during 5 h of treatment while its toxicity increased marginally during 1-5 h. In the BZ 34 strain, the presence of glutathione (GSH) during selenite treatment greatly enhanced the convertogenic and toxic effects of selenite.

Combined mutagenic action of chemicals and radiation in diploid yeast.

The interaction between the mutagenic action of chemicals and radiation was studied by using a diploid yeast strain Saccharomyces cerevisiae BZ 34 with mitotic gene conversion as the end-point. The cells were treated with EMS, MMS or 4-NQO alone or in combination with gamma-radiation. The 2 alkylating agents EMS and MMS produced an additive mutagenic response, whereas 4-NQO exhibited an antagonistic effect in the combined treatment with gamma-radiation.

Effect of caffeine on the genotoxic effects of gamma radiation and 4-NQO in diploid yeast.

Caffeine is an environmental agent to which people are commonly exposed through medicines, drinks, food items, etc. It has been shown to be mutagenic in a number of test systems. In addition, it has also been shown to modify the mutagenic response of ionizing radiation, UV, and several chemical mutagens in a number of test systems. We have studied the effect of caffeine on gamma radiation and 4-Nitroquinoline 1-oxide (4-NQO)-induced gene conversion in the yeast Saccharomyces cerevisiae D7. Stationary phase cells were either exposed to 100-600 Gy of 60Co gamma radiation or treated with 0.15-0.3 microM 4-NQO (30 degrees C, 1 hour), after which they were plated on synthetic complete or minimal media with or without caffeine. Caffeine concentrations ranged from 5 to 15 mM. The results indicated that caffeine at 5 and 10 mM decreased gamma radiation-induced gene conversion frequencies significantly at 400 and 600 Gy. At 600 Gy, the decrease was about 30% and 50% with caffeine concentrations of 5 and 10 mM, respectively. In contrast, caffeine was found to increase the induced gene conversion frequency when cells treated with 0.15, 0.225, and 0.3 microM 4-NQO were plated on media containing caffeine. The increase with 5, 10, and 15 mM caffeine was approximately 1.5, 2, and 2.5, respectively, times the value of 4-NQO alone. The results indicate that the posttreatment repair processes following gamma irradiation or 4-NQO treatment are modified via different pathways.

A study of the induction of gene conversion in yeast by sodium bisulphite under radical reaction conditions.

Sodium bisulphite (NaHSO3) at low concentrations is known to undergo radical reactions in the presence of Mn2+ ions, generating sulphite anion radicals (SO-3.). The kinetics of pH variation in aqueous solutions of NaHSO3 in the presence of 0.1 mM-MnSO4 suggests that the anion radicals are generated rapidly at low bisulphite concentration (10 mM) and quenched at a higher concentration (100 mM). The ability of NaHSO3 to induce gene conversion in the diploid yeast Saccharomyces cerevisiae BZ34 under conditions suitable for radical reactions has been investigated. The results show that NaHSO3 does not induce gene conversion under these conditions.

Hyperthermic modification of radiation induced gene conversion in Saccharomyces cerevisiae.

Effect of pre- and post-irradiation hyperthermia on the induction of gene conversion (non-reciprocal recombination) in diploid yeast cells was investigated. Post irradiation heat treatment does not significantly modify the frequency of gene conversion. Both low-level (51 degrees C-10 min) and lethal (51 degrees C, 40 min) pre irradiation heat treatments enhanced the gene conversion frequency by 17-49%. There was no quantitative correlation between the observed enhancement and the radiation dose. These results suggest that hyperthermia given prior to radiation not only sensitizes the cells to killing but also increases their convertogenic response. It has been suggested that recombination may be involved in carcinogenesis and tumour promotion. Based on these observations along with other reports, wherein hyperthermia was shown to modify the carcinogenic effects in animals as well as the genetic effects of radiation in vitro, it is possible to suggest that hyperthermia may have a potential to increase the second primary cancers, especially in long term survivors. Certain differences in the mechanism of interaction between radiation and heat in the pre- irradiation and post irradiation modalities appear likely.

Modification of radiation-induced damage by hyperthermia--role of repair processes.

The role of repair inhibition in hyperthermic sensitization has been investigated by carrying out interaction studies (non-lethal heat plus radiation) using three radiosensitive mutant strains rad 9-4, rad 51, rad 52 of diploid yeast. The radiosensitization observed in these mutants is compared with that obtained in a wild type strain X2180. Stationary phase cells were heat treated at 51 degrees C for 30 min and log-phase cells at 49 degrees C for 5 min such that the heat treatment per se was non-lethal. Thermal enhancement ratios (TER) were calculated as the ratio of D0 values as well as D10 values, for radiation alone and for combination of heat and radiation. In stationary phase cells TER values ranged from 1.25 to 2 in different strains. There was no significant difference in the degree of sensitization in wild type and mutant strains. However in log-phase cells TER calculated as the ratio of D10 values was 1.84 for wild type strain; whereas the same was 1.2, 1.18 and 1.50 for rad 9-4, rad 51 and rad 52 strains. TER value obtained from ratio of D0 values was also significantly higher for wild type cells in log-phase cultures. These results suggest that the radiosensitization by heat occurs prominently in radioresistant log-phase cells and reflects as a reduction in the shoulder width of survival response curves. Prompt sensitization as seen by the increase in slope of survival curve also contributes to the TER. Inhibition of recovery from sublethal damage by heat appears to be an important mechanism of radiosensitization in log-phase cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Modification of recovery from radiation damage by hyperthermia in yeast.

Recovery from radiation damage was studied in the diploid yeast Saccharomyces cerevisiae BZ 34 subjected to pre-irradiation, post-irradiation or simultaneous hyperthermic treatment. Pre-irradiation heat treatment at 51 degrees C for 20 m min sensitized the cells to lethal damage; however, these cells could efficiently recover from potentially lethal damage (PLD) on liquid holding. In contrast, post-irradiation hyperthermia (51 degrees C for 20 min or 52 degrees C for 15 min) partially inhibited recovery from PLD. For treatments at 40 and 45 degrees C the inhibition of recovery was restricted to the duration of heat treatment. As soon as the temperature of the cells was reduced to 30 degrees C for 48 h, they showed a significant recovery from PLD. The kinetics of the liquid holding recovery of cells exposed to 600 and 900 Gy of radiation followed by heat treatment at 51 degrees C, 20 min or 52 degrees C, 15 min was studied. Post-irradiation heat treatment inhibits PLD repair (PLDR) during the first 3 h. Subsequent liquid holding for longer durations results in recovery from PLD, reaching a maximum of nearly 50% of that obtained in cells exposed to radiation alone. When the cells were held for 0-48 h between radiation and heat treatment, they showed recovery from radiation damage such that sensitization by subsequent heat treatment was reduced significantly over the 48 h interval. The inhibition of PLDR by hyperthermia also decreases progressively with an increase in the time interval between the two treatments. When both heat and radiation treatments were carried out simultaneously by irradiating the cells at 48 degrees C, the inhibition of PLDR was partial. The recovery factor, calculated as the ratio of survival after recovery to that on immediate plating (without PLDR), was 2.6 +/- 0.9 for simultaneous heat and radiation treatment compared to 11.9 +/- 2.7 for radiation alone.