Measuring DNA lesions induced by chemotherapeutic agents – DNA repair and DNA Damage

Abstract

DNA-damaging chemotherapy is amongst the main kinds of cancer treatment. Conversely, our cells are exposed to a wide variety of factors that may cause DNA lesions which, by themselves, may be precursors of cancer development. Cancer genomes, as a whole, accumulate mutations (‘driver’ and ‘passenger’ mutations) which may amount to more than 1000 in cancer-associated genes. These mutations may ensue from a multiplicity of exogenous and endogenous DNA damaging agents, as well as by less proficient DNA repair processes. The first line of cell defense against DNA damage is attributed to BER pathway. The main aim of this study was evaluate the role of BER pathway through the measure of DNA lesions induced by chemotherapeutic agents. Three genes acting on BER pathway were individually silenced in HeLa cells: PARP1, XRCC1 and APE1. These silenced cell-lines were then exposed individually to three different chemotherapeutic agents used in cancer therapy: doxorubicin (DOX); paclitaxel (PAX); and 5-fluorouracil (5-FU), and the DNA damage induced by these agents was measured by comet assay. Our results showed that, globally, silenced cell lines were slightly resistant to drugs exposure than wild-type ones. Furthermore, the effect of DNA lesion measured by % DNA in tail induced by 5-FU and PAX was similar to the one reported for negative control in all cell lines. However, the effect measured for H2O2 (Positive Control) exposure was the inverse, the silenced cell lines showed much more lesion that the wild-type, which might be representative of the lesion type. In conclusion, the impact of each gene silencing on the genotoxicity of individual agents was not fully clear. Additional approaches able to measure other end-points (e.g. apoptosis) are fundamental to better understand the role of BER genes in DNA induced lesion.