Tracking the evolution of breast cancer through single cell analyses of premalignant breast tissues from women at high risk for cancer development

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Prophylactic mastectomy is currently the only breast cancer prevention strategy for women with inherited BRCA1 or BRCA2 mutations. While often effective, this strategy significantly impacts quality of life and does not prevent the development of cancers arising in other tissues (e.g., pancreatic and prostate cancers). Thus, better interception strategies are urgently needed. The overall goal of the parent grant is to characterize the earliest alterations in breast tissues in individuals at high risk of developing breast cancer and to lay the groundwork for the discovery of new prevention measures. Our collaborative studies with Dr. Sam Aparicio using single-cell copy number analyses have identified expanded population of ostensibly normal breast epithelial cells from BRCA1/2 mutation carriers that carry copy number alterations (CNAs). Interestingly, these CNAs are among those that are most commonly associated with early (DCIS) and established (IBC) forms of breast cancer and are present prior to BRCA1/2 loss of heterozygosity (LOH), suggesting that these changes may predispose mammary epithelial cells to tumor-initiating genetic alterations. This supplement proposal aims to test the overarching hypothesis that the genetic alterations that occur prior to LOH of BRCA1 or BRCA2 allow cells to survive eventual loss of both copies of BRCA1 or BRCA2, thereby increasing their cellular fitness and predisposition to malignant cancer following accumulation of additional genetic alterations caused by defects in homologous recombination. We plan to address this hypothesis by 1) complementing our CNA analysis by determining the single nucleotide variant and mutational landscape of precancerous breast tissues using the Tapestri Platform (MissionBio), a state-of-the-art, targeted single cell DNA and protein sequencing technology and 2) utilizing 3D breast organoid models to determine the functional role of CNAs/SNVs in survival after loss of BRCA1/2 and whether additional alterations downstream result in sensitization to tumor development. A better understanding of how early CNAs/SNVs influence tumorigenesis can lead to the identification of vulnerabilities of these cells in order to eliminate them before cancer progresses, as well as to develop strategies to detect expansion of aberrant cells as a means of monitoring pre-malignant cell expansion. Importantly, accomplishing the goals of this proposal will enable Dr. Oliphant to further strengthen the computational skills needed to analyze single-cell multiomic DNA- seq and protein expression datasets independently, as well as leverage 3D organoid models to identify and develop actionable therapeutic strategies more effectively. It will also provide Dr. Oliphant with opportunities for engagement in various career development experiences including participating at conferences and expanding his professional network. Overall, the funding of the proposed project will ensure that Dr. Oliphant expands his scientific and professional training, while establishing the foundation for a viable independent academic research program.