Investigating the contribution of rare non-coding variants in BRCA1, BRCA2 and PALB2 to hereditary breast cancer

Pathogenic coding variants in BRCA1, BRCA2 and PALB2 confer hereditary breast/ovarian cancer risk, yet these regions comprise less than 10% of the genomic footprint of these genes, leaving most sequence unexplored. We investigated the contribution of non-coding variation to hereditary breast cancer by analyzing intronic variants and 5' upstream regions of BRCA1, BRCA2 and PALB2 in the BEACCON case-control study of over 11,000 participants. Full-gene sequencing showed that 46.3% of cases carried at least one rare non-coding variant. This was associated with a modest increase in breast cancer risk (OR = 1.2, p < 0.0001), most likely reflecting the presence of a small proportion of pathogenic variants within a larger background of predominantly neutral variation. Stronger enrichment was observed for triple-negative disease, particularly for BRCA1 (OR = 1.5, p = 0.0001). Tumor sequencing of 42 high-priority variants identified 11 (26.2%) with wild-type allele loss and high homologous recombination deficiency. Functional CRISPR/Cas9 knock-in assays in MCF10A cells confirmed that two deep intronic variants created aberrant splice sites, disrupted splicing and impacted transcript expression.