Ultra-rare functional variants reveal early-onset breast cancer risk genes and pathways in the UK Biobank and All of Us Research Program.

Breast cancer (BRCA) affects one in eight women, and while past studies have identified susceptibility genes and variants, those driving early-onset disease remain opaque. We introduce EA-Pathways, a control-free association method that aggregates ultra-rare germline coding variants weighted by Evolutionary Action to detect functional impact distribution biases in genes and pathways. In UK Biobank women with BRCA, EA-Pathways prioritized candidate risk genes across pathways enriched in known familial genes. Ultra-rare, high-impact variants in these genes were BRCA specific, and variants from eight pathways were associated with 2-year-earlier disease, including homology-directed DNA repair, TP53, and pexophagy pathways. The early-onset effect replicated in All of Us with diagnoses up to 6, 9, and 3 years earlier in European, admixed American, and African genetic ancestries, highlighting risk alleles with differing frequencies between ancestry groups. These findings demonstrate EA-Pathways as a sensitive pathway association method capable of identifying cancer-predisposing genes and ancestry-informed BRCA screening opportunities.