ABSTRACT: Benzo[a]pyrene diol epoxide (BPDE) is a highly carcinogenic metabolite of the environmental contaminant Benzo[a]pyrene, which is commonly found as a by-product of incomplete combustion of organic matter. BPDE reacts with the DNA to form BPDE-DNA bulky adducts which, if not removed, can lead to mutations due to DNA base-pair substitution and consequently to tumorigenesis. Due to its lipophilic nature, B[a]p can cross the placenta to reach the developing fetus and the placenta of mice, rats and humans can metabolize B[a]p into BPDE, which can lead to fetal genotoxic exposure. Besides direct embryonic exposure, BPDE-DNA adducts are detectable in the sperm and ovarian cells of cigarette smokers, and these DNA modifications can be paternally transmitted through the spermatozoa to the embryo. Human induced pluripotent stem cells (hiPSCs) can be used as a model of human embryonic stem cells (hESCs), an with that in mind, we treated hiPSCs with BPDE and performed a microarray analysis to evaluate for the first time the effects of BPDE exposure in pluripotent stem cells. Furthermore, hiPSCs derived from patients suffering from Nijmegen Breakage Syndrome, a chromosomal instability disorder characterized by microcephaly, defective DNA repair and increased risk of malignancies, were also investigated in the context of BPDE exposure and compared to their healthy counterparts. Transcriptomics analysis, coupled with protein content analysis through immunostaining and western blots, revealed that hiPSCs have a robust reaction to BPDE exposure, with an enhanced expression of several targets related to the p53 mediated DNA damage response, including DNA lesion bypass, cell cycle checkpoints and extrinsic apoptosis. Additionally, hiPSC harbouring an NBS mutation reacted differently to BPDE treatment than WT cells, showing less apoptotic response, no p53 or MDM2 protein increase, increased transcription of cancer-related targets and the repression of DNA-repair pathways transcription.
