Mapping lineage and functional diversity of the mammary epithelium in women at high risk of breast cancer

Breast cancer risk is shaped by the vast heterogeneity of mammary epithelial cells, comprising basal, luminal progenitor, and mature luminal populations. While transcriptional variation among these lineages has been extensively studied, protein-level features—particularly in high-risk women—remain underexplored, limiting insight into early cellular and molecular determinants of susceptibility. Moreover, little is known about how clinical covariates influence clonogenic capacity, proteomic states, and epithelial proportions. We combine low-input proteomics with functional clonogenic assays to profile mammary epithelial cell subpopulations from a cohort of 22 breast tissues encompassing different germline mutation backgrounds, parity status and age. We quantify 5,555 proteins and observed marked inter-donor variation in epithelial composition, proteomic programs, and colony-forming capacity. Multivariable modeling reveals that clinical covariates—including age, parity, and germline mutation status—modulate both global proteomic architecture and lineage-specific pathway activity. Parity is associated with reduced basal cell abundance, altered luminal progenitor and mature luminal proteomes, and changes in clonogenicity. Pathway analyses identify both conserved and lineage-restricted responses to shared risk factors. Projection of clonogenic signatures onto METABRIC and TCGA tumors further links functional programs to tumor subtypes and molecular phenotypes. This study provides the most comprehensive proteomic atlas of cell-type resolved diversity in the high-risk breast to date. By defining how clinical covariates shape epithelial composition and molecular state, it clarifies key sources of biological variability that challenge controlled study design and offers a resource for improving mechanistic insight, risk assessment, and prevention strategies.