These results confirm the efficacy of the automated MN assay for fast population triage in a multicenter setting, in the case of large radiation accidents.

No abstract

As enhanced chromosomal radiosensitivity (CRS) results from non- or misrepaired double strand breaks (DSBs) and is a hallmark for breast cancer and single nucleotide polymorphisms (SNPs) in DSB repair genes, such as non homologous end-joining (NHEJ) genes, could be involved in CRS and genetic predisposition to breast cancer. In this study, we investigated the association of five SNPs in three different NHEJ genes with breast cancer in a population-based case-control setting. The total patient population composed of a selected group of patients with a family history of the disease and an unselected group, consisting mainly of sporadic cases. SNP analysis showed that the c.2099-2408G>A SNP (XRCC5Ku80) [corrected] has a significant, positive odds ratio (OR) of 2.81 (95% confidence interval (CI): 1.30-6.05) for the heterozygous (He) and homozygous variant (HV) genotypes in the selected patient group. For the c.-1310 C>G SNP (XRCC6Ku70)[corrected] a significant OR of 1.85 (95%CI: 1.01-3.41) was found for the He genotype in the unselected patient group. On the contrary, the HV genotype of c.1781G>T (XRCC6Ku70) [corrected] displays a significant, negative OR of 0.43 (95%CI: 0.18-0.99) in the total patient population. The He+HV genotypes of the c.2099-2408G>A SNP (XRCC5Ku80) [corrected] also showed high and significant ORs in the group of "radiosensitive," familial breast cancer patients. In conclusion, our results provide preliminary evidence that the variant allele of c.-1310C>G (XRCC6Ku70) [corrected]and c.2099-2408G>A (XRCC5Ku80) [corrected] are risk alleles for breast cancer as well as CRS. The HV genotype of c.1781G>T (XRCC6Ku70) [corrected] on the contrary, seems to protect against breast cancer and ionizing radiation induced micronuclei.

These results show that the variant allele of c.-1310 C>G, located in the Ku70 promoter, is a risk allele for breast cancer. Furthermore, the association of the c.-1310 C>G SNP with breast cancer risk was stronger in women with a long oestrogen exposure.

Abstract. Enhanced in vitro chromosomal radiosensitivity (CRS) has been proposed as a marker for low-penetrance gene mutations predisposing to breast cancer (BC). Since the double strand break (DSB) is the most detrimental form of DNA damage induced by ionizing radiation, it is possible that mutations in genes encoding proteins involved in DSB repair affect breast cancer risk. The purpose of the present study was to examine whether five single nucleotide polymorphisms (SNPs) in Rad51 and Xrcc3 (rs1801320, rs1801321, rs1799796, rs861539 and rs1799794) exhibited an association with breast cancer susceptibility in a Belgian population of BC patients with a known or putative genetic predisposition. We also ascertained whether a relationship exists between the occurrence of the 'variant' alleles of these variations and in vitro CRS. Blood samples were obtained from BC patients and from healthy female individuals. Variations in the 5' UTR of Rad51 and Xrcc3 were genotyped, and statistical analysis was performed. The results showed that low-penetrant variations in Rad51 and Xrcc3, two proteins belonging to the homologous recombination DSB repair pathway, may modify BC risk in patients already carrying a pathological mutation in the highly penetrant BC genes BRCA1 and BRCA2. Combined risk genotype analysis revealed that Rad51 SNPs enhance BC risk in BRCA2 patients, whereas Xrcc3 SNPs significantly enhance BC risk in carriers of BRCA1 mutations and in patients with hereditary BC. When four putative risk genotypes of Rad51 and Xrcc3 were combined, positive significant odds ratios were obtained in the entire patient population and in patients with a hereditary history of disease. Although obtained from a limited number of patients, our data are supportive of a polygenic model whereby combinations of weak variations are responsible for an enhanced BC risk by acting jointly with high-penetrant mutations in BRCA1 or BRCA2.

Breast cancer patients are characterised by an enhanced chromosomal radiosensitivity pointing to a defect in the repair of DNA double strand breaks (dsb). In mammalian cells, radiation induced dsb are mainly repaired by the non homologous end-joining (NHEJ) repair pathway, a pathway in which the Ku70/Ku80 heterodimer plays a key role as it binds to the broken DNA ends. In this study we wanted to investigate the radiosensitizing effect of Ku70/80 knockdown, by lentivirus-mediated RNAinterference, in a spontaneous immortalised human mammary epithelial cell line (MCF10A). Several endpoints for measuring radiosensitivity were taken into acount: micronucleus formation (chromosomal radiosensitivity), cell survival, apoptosis and senescence. For all endpoints, MCF10A cells were infected with lentiviral vectors for RNAi of Ku70 (pLVTHM/shKu70/GFP). Western blot analysis showed that the Ku70 lentiviral vector was effective in silencing the expression of both Ku70 and Ku80. When a satisfactory knockdown was obtained (70-90% vs. mock-infected (pLVTHM/GFP) cells), the cells were used to perform radiation experiments. For the in vitro MN assay, cells were irradiated with doses of 2 and 4 Gy 60Co gamma-rays. A significantly higher radiationinduced MN yield was obtained in the Ku70/80 knock down cell line compared to the mock-infected cell line, pointing to an increased chromosomal radiosensitivity. This increased chromosomal radiosensitivity demonstrates that the repair genes, Ku70 and Ku80, are involved in the repair of DNA double strand breaks, which are the main DNA lesions resulting in chromosomal aberrations such as micronuclei. Besides chromosomal radiosensitivity we also investigated radiosensitivity at the cellular level by cell survival experiments. Cells were irradiated with doses ranging between 0 and 8 Gy and cultured for 5 days before being analysed. The results of the cell survival assay show that Ku70/80 knockdown cells have a lower survival yield after irradiation compared to mock-infected cells, pointing to an enhanced cellular radiosensitivity. Analysis of the cell death pattern showed that MCF10 cells (Ku70/80 knockdown and mock-infected) do not undergo apoptosis but go into cellular senescence. In conclusion, we can state that knockdown of Ku70 and Ku80 by RNAi of Ku70 resulted in an increased chromosomal and cellular radiosensitivity in an immortalised MCF10 human mammary cell line after irradiation with low LET 60Co gamma-rays. These results may further support the role of DNA dsb repair genes in breast cancer. 230

No abstract

Abstract. More than 60 years after their isolation and characterization, aminoglycoside (AG) antibiotics remain powerful agents in the treatment of severe gram-negative, enterococcal or mycobacterial infections. However, the clinical use of AGs is hampered by nephrotoxicity and ototoxicity, which often develop as a consequence of prolonged courses of therapy, or of administration of increased doses of these drugs. The discovery of non-ototoxic antibacterial agents, showing a wider spectrum of activity, has gradually decreased the use of AGs as first line antibiotics for many systemic infections. However, AGs are now undergoing an unexpected revival, being increasingly indicated for the treatment of severe emerging infections caused by organisms showing resistance to most first-line agents (e.g., multidrug-resistant tuberculosis, complicated nosocomially-acquired acute urinary tract infections). Increasing adoption of aminoglycosides poses again to scientists and physicians the problem of toxicity directed to the kidneys and to the inner ear. In particular, aminoglycoside-induced deafness can be profound and irreversible, especially in genetically predisposed patients. For this reason, an impressive amount of molecular strategies have been developed in the last decade to counteract the ototoxic effect of aminoglycosides. The present article overviews: i) the molecular mechanisms by which aminoglycosides exert their bactericidal activity, ii) the mechanisms whereby AGs exert their ototoxic activity in genetically-predisposed patients, iii) the drugs and compounds that have so far proven to prevent or modulate AG ototoxicity at the preclinical and/or clinical level, and iv) the dosage regimens that have so far been suggested to decrease the incidence of episodes of AG-induced ototoxicity. Aminoglycoside antibiotics: a revival?Aminoglycoside (AG) antibiotics have been extensively used for the prophylaxis and the treatment of a wide variety of systemic infections, for the outstanding features displayed by these antibacterial agents, i.e., the concentration-dependent bactericidal activity, the post-antibiotic effect, the favorable pharmacokinetic profile, and the strong synergism with other antibiotics such as vancomycin and ß-lactams (1). More than 60 years after their isolation and characterization, these drugs remain powerful tools for treatment of severe gram-negative, enterococcal or mycobacterial infections. AGs are usually administered in combination with ß-lactam antibiotics for a variety of systemic infections (e.g., bacterial endocarditis or various pseudomonal infections), but are also used as monotherapy for infections of the urinary tract. Streptomycin, the first AG isolated and adopted for human therapy (Fig. 1), MOLECULAR MEDICINE REPORTS 1: 3-13, 2008 3 Prevention and modulation of aminoglycoside ototoxicity (Review) GIANPAOLO