Mutational Landscape of DNA Damage Response Genes in Locally Advanced and Advanced Ovarian Cancer
This study aimed to investigate mutations in 14 DNA Damage Response (DDR) genes in patients with Locally Advanced and Advanced Ovarian Cancer (OC) treated at the Instituto Nacional de Cancerologia (INCAN) in Mexico City, Mexico. A prospective, sequential, observational study was conducted and approved by the Ethics and Review Board of INCAN (015/037/ICI) (CEI/1000/15), with all patients providing informed consent. Tumor biopsies from 36 patients were obtained post-surgery, where tissue samples were divided for pathological confirmation and DNA extraction.Tumor DNA was extracted using the QIAamp DNA Blood Mini Kit (Qiagen, Hilden, Germany), and the integrity and concentration of DNA were assessed by agarose gel electrophoresis and RNase P Detection Reagent, respectively. We focused on sequencing 14 DDR genes (ARID1A, BRCA1, BRCA2, ATM, CHK2, FANCA, FANCB, FANCC, FANCD2, PALB2, PARP1, RAD50, RAD51, and TP53) using targeted amplification. BRCA1 and BRCA2 were amplified using the Ion Ampliseq BRCA 1/2 panel (Thermo Fisher Scientific), while the other genes were amplified with a custom-designed panel (IAD94476_197_Designed). A total of 25 ng of DNA was used for library preparation, with samples uniquely barcoded using Ion Xpress adapters (Thermo Fisher Scientific).Sequencing data were aligned to the hg19 human reference genome (GRCh37), and variant analysis was conducted using Ion Reporter version 5.16. Pathogenic and likely pathogenic variants were classified according to the American College of Medical Genetics and Genomics guidelines. This comprehensive analysis aims to identify critical genetic alterations in DDR genes that could be potential biomarkers for therapeutic strategies and prognostic evaluation in OC.