title: Combined genetic and splicing analysis of BRCA1 c.[594-2A>C; 641A>G] highlights the relevance of naturally occurring in-frame transcripts for developing disease gene variant classification algorithms.
abstract: A recent analysis using family history weighting and co-observation classification modeling indicated that BRCA1 c.594-2A > C (IVS9-2A > C), previously described to cause exon 10 skipping (a truncating alteration), displays characteristics inconsistent with those of a high risk pathogenic BRCA1 variant. We used large-scale genetic and clinical resources from the ENIGMA, CIMBA and BCAC consortia to assess pathogenicity of c.594-2A > C. The combined odds for causality considering case-control, segregation and breast tumor pathology information was 3.23 × 10-8Our data indicate that c.594-2A > C is always in cis with c.641A > G. The spliceogenic effect of c.[594-2A > C;641A > G] was characterized using RNA analysis of human samples and splicing minigenes. As expected, c.[594-2A > C; 641A > G] caused exon 10 skipping, albeit not due to c.594-2A > C impairing the acceptor site but rather by c.641A > G modifying exon 10 splicing regulatory element(s). Multiple blood-based RNA assays indicated that the variant allele did not produce detectable levels of full-length transcripts, with a per allele BRCA1 expression profile composed of ≈70-80% truncating transcripts, and ≈20-30% of in-frame Δ9,10 transcripts predicted to encode a BRCA1 protein with tumor suppression function.We confirm that BRCA1c.[594-2A > C;641A > G] should not be considered a high-risk pathogenic variant. Importantly, results from our detailed mRNA analysis suggest that BRCA-associated cancer risk is likely not markedly increased for individuals who carry a truncating variant in BRCA1 exons 9 or 10, or any other BRCA1 allele that permits 20-30% of tumor suppressor function. More generally, our findings highlight the importance of assessing naturally occurring alternative splicing for clinical evaluation of variants in disease-causing genes.
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mesh_terms: D000328:Adult; D000368:Aged; D017398:Alternative Splicing / Q000235:genetics; D019313:BRCA1 Protein / Q000235:genetics*; D001943:Breast Neoplasms / Q000235:genetics* / Q000473:pathology; D004252:DNA Mutational Analysis; D005091:Exons / Q000235:genetics; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D006801:Humans; D008875:Middle Aged; D009154:Mutation / Q000235:genetics*; D010051:Ovarian Neoplasms / Q000235:genetics* / Q000473:pathology; D022821:RNA Splice Sites / Q000235:genetics; D012326:RNA Splicing / Q000235:genetics; D025521:Tumor Suppressor Proteins / Q000235:genetics*
chemical_list: D019313:BRCA1 Protein; D022821:RNA Splice Sites; D025521:Tumor Suppressor Proteins