caa a22 4500 606190635 CHVBK 20210128100856.0 cr unu---uuuuu 210128e20151201xx s 000 0 eng 10.1007/s10577-015-9478-4 doi (NATIONALLICENCE)springer-10.1007/s10577-015-9478-4 Progression of chromosomal damage induced by etoposide in G2 phase in a DNA-PKcs-deficient context [Elektronische Daten] [Micaela Palmitelli, Marcelo de Campos-Nebel, Marcela González-Cid] Etoposide (ETO), a drug used for the treatment of human tumors, is associated with the development of secondary malignancies. Recently, therapeutic strategies have incorporated chemosensitizing agents to improve the tumoral response to this drug. ETO creates DNA double-strand breaks (DSB) via inhibition of DNA topoisomerase II (Top2). To repair DSB, homologous recombination (HR) and non-homologous end-joining (NHEJ), involving D-NHEJ (dependent of the catalytic subunit of DNA-dependent protein kinase, DNA-PKcs) and B-NHEJ (backup repair pathway) are activated. We evaluated the progression of the DNA damageinduced by the Top2 poison ETO in G2 phase of human HeLa cells after chemical inhibition ofDNA-PKcs with NU7026. Compared to ETO treatment alone, this combined treatment resulted in a twofold higher rate of chromatid breaks and exchanges when analysis was performed in the following metaphase. Moreover, when analysis was performed in the second metaphase following treatment, increases in the percentage of micronuclei with H2AX (biomarker for DSB) foci in binucleated cells and dicentric chromosomes were seen. In post-mitotic G1 phase, a close association between unresolved DSB and meiotic recombination 11 homolog A (MRE11) signals was observed, demonstrating the contribution of MRE11 in the DSB repair by B-NHEJ. Hence, chemical inhibition of DNA-PKcs impaired both D-NHEJ and HR repair pathways, altering the maintenance of chromosomal integrity and cell proliferation. Our results suggest that the chemosensitizing effectiveness of the DNA-PKcs inhibitor and the survival rate of aberrant cells may contribute to the development of therapy-related tumors. Springer Science+Business Media Dordrecht, 2015 Topoisomerase II poison nationallicence DNA damage nationallicence DSB repair pathway nationallicence G2 phase nationallicence DNA-PKcs chemical inhibition nationallicence A-NHEJ : Alternative non-homologous end-joining nationallicence ATM : Ataxia-telangiectasia mutated nationallicence B-NHEJ : Backup non-homologous end-joining nationallicence BN : Binucleated cells nationallicence BrdU : 5-Bromo-2′-deoxyuridine nationallicence C-NHEJ : Classic non-homologous end-joining nationallicence CENP-F : Centromere protein F nationallicence D-NHEJ : DNA-PKcs-dependent non-homologous end-joining nationallicence DAPI : 4,6-Diamidino-2-phenylindole nationallicence DMSO : Dimethyl sulfoxide nationallicence DNA-PKcs : DNA-dependent protein kinase catalytic subunit nationallicence DSB : Double-strand breaks nationallicence ETO : Etoposide nationallicence FISH : Fluorescent in situ hybridization nationallicence FITC : Fluorescein isothiocyanate nationallicence γH2AX : Histone H2AX-phosphorylated on serine 139 nationallicence HR : Homologous recombination nationallicence MI : Mitotic index nationallicence MN : Micronuclei nationallicence MRE11 : Meiotic recombination 11 homolog A nationallicence NU7026 : 2-(Morpholin-4-yl)-benzo[h]chromen-4-one nationallicence PBS : Phosphate-buffered saline nationallicence PFA : Paraformaldehyde nationallicence PI : Propidium iodide nationallicence Rad51 : DNA repair protein RAD51 homolog 1 nationallicence SCE : Sister chromatid exchanges nationallicence Top2 : Topoisomerase II nationallicence Palmitelli Micaela Laboratorio de Mutagénesis, Instituto de Medicina Experimental, IMEX-CONICET, Academia Nacional de Medicina, J. A. Pacheco de Melo 3081, 1425, Buenos Aires, Argentina aut de Campos-Nebel Marcelo Laboratorio de Mutagénesis, Instituto de Medicina Experimental, IMEX-CONICET, Academia Nacional de Medicina, J. A. Pacheco de Melo 3081, 1425, Buenos Aires, Argentina aut González-Cid Marcela Laboratorio de Mutagénesis, Instituto de Medicina Experimental, IMEX-CONICET, Academia Nacional de Medicina, J. A. Pacheco de Melo 3081, 1425, Buenos Aires, Argentina aut Chromosome Research Springer Netherlands 23/4(2015-12-01), 719-732 0967-3849 23:4<719 2015 23 10577 https://doi.org/10.1007/s10577-015-9478-4 text/html Onlinezugriff via DOI BK010053 XK010053 XK010000 Metadata rights reserved Springer special CC-BY-NC licence nationallicence 1 research-article jats NATIONALLICENCE NATIONALLICENCE NL-springer NATIONALLICENCE 856 40 https://doi.org/10.1007/s10577-015-9478-4 text/html Onlinezugriff via DOI NATIONALLICENCE 700 1- Palmitelli Micaela Laboratorio de Mutagénesis, Instituto de Medicina Experimental, IMEX-CONICET, Academia Nacional de Medicina, J. A. Pacheco de Melo 3081, 1425, Buenos Aires, Argentina aut NATIONALLICENCE 700 1- de Campos-Nebel Marcelo Laboratorio de Mutagénesis, Instituto de Medicina Experimental, IMEX-CONICET, Academia Nacional de Medicina, J. A. Pacheco de Melo 3081, 1425, Buenos Aires, Argentina aut NATIONALLICENCE 700 1- González-Cid Marcela Laboratorio de Mutagénesis, Instituto de Medicina Experimental, IMEX-CONICET, Academia Nacional de Medicina, J. A. Pacheco de Melo 3081, 1425, Buenos Aires, Argentina aut NATIONALLICENCE 773 0- Chromosome Research Springer Netherlands 23/4(2015-12-01), 719-732 0967-3849 23:4<719 2015 23 10577