Spectrum of germline variants in moderate to high penetrance non-BRCA breast cancer susceptibility genes in a Pakistani early-onset cohort.

BACKGROUND: Hereditary factors account for a significant lifetime risk of breast cancer. Approximately 20% is attributable to pathogenic variants in the highly penetrant BRCA1/2 of the homologous recombination repair (HRR) pathway. Other HRR pathway genes (ATM, CHEK2, BARD1, TP53) are also linked to breast cancer susceptibility; however, the contribution of these genes remains underexplored in Pakistani populations. METHODS: A cohort of 66 early-onset breast cancer patients underwent next-generation sequencing (NGS) of BRCA1/2 followed by four non-BRCA breast cancer susceptibility genes (ATM, CHEK2, BARD1, and TP53) using a multigene panel. The present analysis focused primarily on variants identified in the four non-BRCA genes, which were annotated using ANNOVAR and in silico prediction algorithms. Variant classification was assessed using ClinVar and ACMG/AMP guidelines. RESULTS: A total of 478 variants were identified. Two pathogenic stop-gain variants in CHEK2 were detected in three patients (4.5%), while two missense and one 5'UTR variant of uncertain significance (VUS) in ATM were identified in three patients (4.5%). Further, based on MAF < 1%, n= 123 (47.85%) variants were recurrent, and 15 (5.8%) novel alleles were detected; of these, two novel missense variants were identified in exonic region in BARD1 and CHEK2. CONCLUSION: This study highlights the contribution of pathogenic variants in CHEK2, supports the clinical relevance in breast cancer, and identifies recurrent variants alongside potentially novel, population-specific alleles in BARD1 and CHEK2. The identification of VUS in ATM underscores the need for functional and population-based studies to elucidate their clinical significance and to advance precision oncology in Pakistan.