A ۵-Gene Panel for Testing High-risk Individualsfor Breast Cancer Using Next Generation Sequencing

Introduction: Breast cancer is one of the most common anddeadly diseases in women which occurs due to hereditary mutationsin about ۱۰% of patients. Finding the pathogenic andlikely pathogenic mutations is crucial for identification of highriskpeople, which can be followed by the preventive actionssuch as mastectomy in mutation carriers and also determiningthe appropriate treatment process in patients with breast cancer.Next generation sequencing is a clinical method for studyingmultiple genes in breast cancer panel simultaneously.Materials and Methods: A library preparation kit and analysispipeline for HBOC amplicon sequencing was designed, developedand validated (۵ genes, including BRCA۱, BRCA۲, TP۵۳,PALB۲ and CHEK۲). The study was conducted on ۲۷۵ individualsidentified as high-risk according to the NCCN criteria,such as the history of breast cancer diagnosis at the age before۴۵ years or a history of breast cancer in male family members.The next generation sequencing performed by illumina MiSeqsystem for detecting single nucleotide variations and smallindels. The results were analyzed using multiple packages includingTrimmomatic ۰.۳۹, FastQC ۰.۱۱.۹, Burrows-WheelerAligner, Picard Tools, GATK ۴.۲.۶.۱, and Jannovar ۰.۳۸ andthen confirmed by Clinvar and HGMD databases.Results: We observed pathogenic and likely pathogenic variantsin BRCA۱/۲, CHEK۲, and PALB۲. The frequencies in ourpopulation were ۸.۹%, ۳.۰%, and ۱.۵% respectively. All of the mutations were checked and confirmed by Sanger sequencingmethod.Conclusion: Next-generation sequencing is an accurate, costeffectiveand appropriate method for molecular analysis in peoplewith family or personal history of breast and ovarian cancer,which can be used to enhance the prevention and treatment.