Analysis of how mutations accumulate in pretumor tissue, although widely presumed to occur, has been extremely difficult to study. This is principally because, with most such mutations being unique to individual cells within a tissue, their detection is technically challenging. In this study we propose to apply Single Cell Multiple Displacement Amplification (SCMDA) that we recently developed for high accuracy detection of a spectrum of mutations from single nucleotide substitutions to indels and aneuploidy in individual cells within pre-tumor tissues of women who inherited mutations in the BRCA1 or BRCA2 genes. We hypothesize that mutations from single nucleotide substitutions to indels, large genomic rearrangements, and aneuploidy accumulating as consequence of defects in homology dependent DNA repair in mammary epithelial cells are the underlying cause of increased cancer risk in these women. We further hypothesize that estrogen, which is known to generate metabolites that directly damage DNA, mechanistically acts as a modifier ofBRCA1/2 cancer penetrance by working in concert with the BRCA1/2 repair defects to increase the somatic mutation rate in the cells of BRCA1/2 carriers. In Aim 1, we will utilize SCMDA to test if mutation frequencies are elevated in individual BRCA1/2 heterozygous mammary epithelial cells. In Aim 2, we will directly test the hypothesis that estrogen increases mutation frequencies in BRCA1/2 mutant cells.