Deregulated estrogen metabolism in BRCA1 deficient cells induces chromosomal instability.

Objectives: Germline mutations in BRCA1 predispose carriers to a high<br/>incidence of breast and ovarian cancers. The BRCA1 protein functions to maintain<br/>genomic stability via important roles in DNA repair, transcriptional regulation, and<br/>post-replicative repair. Despite functions in processes essential in all cells, BRCA1<br/>loss or mutation leads to tumours predominantly in estrogen-regulated tissues.<br/>Here, we aim to determine if endogenous estrogen metabolites may be an initiator<br/>of genomic instability in BRCA1 deficient cells.<br/><br/>Methods: We analysed DNA DSBs by ?H2AX, 53BP1, and pATM1981<br/>foci and neutral comet assay, estrogen metabolite concentrations by LC-MS/MS,<br/>and BRCA1 transcriptional regulation of metabolism genes by ChIP-chip, ChIP,<br/>and qRT-PCR.<br/><br/>Results: We show that estrogen metabolism is perturbed in BRCA1 deficient<br/>cells resulting in elevated production of 2-hydroxyestradiol (2-OHE2) and 4-hydroxyestradiol (4-OHE2), and decreased production of the protective metabolite<br/>4-methoxyestradiol. We demonstrate that 2-OHE2 and 4-OHE2 treatment leads<br/>to DNA double strand breaks (DSBs) in breast cells, and these DSBs were exacerbated<br/>in both BRCA1 depleted cells and BRCA1 heterozygous cells (harbouring<br/>185delAG mutation). Furthermore, the DSBs were not repaired efficiently in either<br/>BRCA1 depleted or heterozygous cells, and we found that 2-OHE2 and 4-OHE2<br/>treatment generates chromosomal aberrations in BRCA1 depleted cells. We suggest<br/>that the increase in DNA DSBs in BRCA1 deficient cells is due to loss of<br/>both BRCA1 transcriptional repression of estrogen metabolising genes (such as<br/>CYP1A1 and CYP3A4) and loss of transcriptional activation of detoxification<br/>genes (such as COMT).<br/><br/>Conclusions: We suggest that BRCA1 loss results in estrogen driven tumourigenesis<br/>through a combination of increased expression of estrogen metabolising<br/>enzymes and reduced expression of protective enzymes, coupled with a defect in<br/>the repair of DNA DSBs induced by endogenous estrogen metabolites. The overall<br/>effect being an exacerbation of genomic instability in estrogen regulated tissues in<br/>BRCA1 mutation carriers.