In this study, compared with control cells, the mRNA and protein expression levels of RAD51 were decreased in HONE-1 and TW01 cells after O6BG (O6-methylguanine-DNA methyltransferase inhibitor) treatment in dose and time-dependent manners. Using confocal immunomicroscopy, the percentage of cells with gamma-H2AX and RAD51 co-localization, indicating the repair activity of DNA double-strand break, was decreased by 20% and 15% in HONE-1 and TW01 cells with the combination treatment, compared with the group with cisplatin alone. The activity of O6BG to suppress RAD51 translocation into the nucleus was also revealed by confocal immunomicroscopy and western blot with subcellular fractionation. Using a plasmid-based method, the homologous recombination capacity was decreased by 20% and 27% in HONE-1 and TW01 cells with O6BG treatment. In xenografts, the volume of HONE-1 cells was reduced by 20% in mice treated with cisplatin alone, and by 50% in those given combination treatment. Immunohistochemical analysis also showed that O6BG inhibited RAD51 expression in vivo. These findings suggest O6BG combined with cisplatin might provide a novel therapeutic strategy for human cancers.
