Intrinsic radiosensitivity of human pancreatic tumour cells and the radiosensitising potency of the nitric oxide donor sodium nitroprusside.

A panel of eight human pancreatic tumour cell lines displayed high intrinsic radioresistance, with mean inactivation doses between 2.4 and 6.5 Gy, similar to those reported for melanoma and glioblastoma. The radiosensitising potency of sodium nitroprusside, a bioreductive nitric oxide donor, was assessed in a model of metabolism-induced hypoxia in a cell micropellet. Sodium nitroprusside at 0.1 mM revealed a radiosensitising effect with an overall enhancement ratio of 1.9 compared with 2.5 for oxygen. Radiosensitising activity correlated with the enhancement of single-strand DNA breakage caused by radiation. In suspensions with cell densities of between 3% and 30% (v/v), the half-life of sodium nitroprusside decreased from 31 to 3.2 min, suggesting a value of around 1 min for micropellets. Despite this variation, the radiosensitising activity was similar in micropellets and in diluted cell suspensions. S-nitroso-L-glutathione was found to possess radiosensitising activity, consistent with a possible role of natural thiols in the storing of radiobiologically active nitric oxide adducts derived from sodium nitroprusside. As measured by a nitric oxide-specific microsensor, activation of sodium nitroprusside occurred by bioreduction, whereas S-nitroso-L-glutathione showed substantial spontaneous decomposition. Both agents appear to exert radiosensitising action through nitric oxide as its scavenging by carboxy phenyltetramethylimidazolineoxyl N-oxide (carboxy-PTI0) and oxyhaemoglobin resulted in attenuated radiosensitisation. Sodium nitroprusside was at least 10-fold more potent than etanidazole, a 2-nitroimidazole used as a reference. Our data suggest that sodium nitroprusside, a drug currently used for the treatment of hypertension, is a potential tumour radioresponse modifier

Radiosensitization of hypoxic tumour cells by S-nitroso-N-acetylpenicillamine implicates a bioreductive mechanism of nitric oxide generation

The radiosensitizing activity of S-nitroso-N-acetylpenicillamine (SNAP), a nitric oxide (NO) donor, was assessed in a model of non-metabolic hypoxia achieved in an atmosphere of 95% nitrogen–5% carbon dioxide. A 10 min preincubation of hypoxic EMT-6 cells (10 × 106 ml−1) with 0.1 and 1 mM SNAP before radiation resulted in an enhancement ratio of 1.6 and 1.7 respectively. The level of spontaneous NO release, measured by a NO specific microsensor, correlated directly with the concentration of SNAP and was enhanced 50 times in the presence of cells. Dilution of the cell suspension from 10 to 0.1 × 106 ml−1 resulted in a 16-fold decline in NO release, but only a twofold decrease in radiosensitization was observed. Preincubation of hypoxic cells with SNAP for 3 min up to 30 min caused an increasing radiosensitizing effect. Extended preincubation of 100 min led to the loss of radiosensitization although the half-life of SNAP is known to be 4–5 h. Taken together, these observations suggest that SNAP generates NO predominantly by a bioreductive mechanism and that its biological half-life is unlikely to exceed 30 min. The lack of correlation between free NO radical and radiosensitizing activity may reflect a role of intracellular NO adducts which could contribute to radiosensitization as well. © 1999 Cancer Research Campaig

NF-κB inhibition impairs the radioresponse of hypoxic EMT-6 tumour cells through downregulation of inducible nitric oxide synthase

Hypoxic EMT-6 tumour cells displayed a high level of inducible nitric oxide synthase (iNOS) and an increased radiosensitivity after a 16 h exposure to lipopolysaccharide, a known activator of nuclear factor-κB (NF-κB). Both iNOS activation and radioresponse were impaired by the NF-κB inhibitors phenylarsine oxide and lactacystin. Contrasting to other studies, our data show that inhibition of NF-κB may impair the radioresponse of tumour cells through downregulation of iNOS. © 2003 Cancer Research UK.SCOPUS: ar.jinfo:eu-repo/semantics/publishe

DNA breakage, cytotoxicity, drug accumulation and retention in two human ovarian tumor cell lines AZ224 and AZ364 treated with adriamycine, modulated by verapamil.

We investigated the cytotoxicity of adriamycin in two human ovarian tumor cell lines, AZ224 and AZ364, by the MTT-test and we analysed the formation of DNA single-strand (SSB) and double-strand breaks (DSB) by means of the alkaline and neutral elution technique. The AZ364 cell line was 15 times more resistant to ADR (ID 50 = 10.0 μg/ml) than the AZ224 cell line (ID 50 = 0.66 μg/ml) after 1 hr of drug exposure. Immediately after exposure, we observed a biphasic dose response for SSB in the AZ224 cells over a concentration range of 0.1 to 32.0 μg/ml, while practically no DSB were found. Upon drug removal and incubation in drug-free medium, full repair of SSB was observed for an ADR concentration of 1 μg/ml. On the contrary, the DSB became significantly increased for all tested concentrations and persisted as observed after 3 hr and 7 hr in drug-free medium. The resistant cell line AZ 364 showed consistently less DNA breakage than the AZ 224 cell line. This inherent difference in sensitivity to ADR could, however, not be explained on the basis of the cellular pharmacokinetics of the drug. Verapamil induced a 3 to 4 fold potentiation of the ADR cytotoxicity in both cell lines after continuous exposure and was associated with an increase in DNA-breakage. The results of our study confirm that there is a lack of correlation between cytotoxicity of ADR and DNA strand breakage immediately after 1 hr of drug exposure. Instead, we emphasize the importance of the formation, extent and persistence of protein-associated DSB upon drug removal to the cytotoxic action of ADR in vitro.SCOPUS: ar.jinfo:eu-repo/semantics/publishe