166P Clinical use of tumor sequencing for identification of germline pathogenic variants (gPV) in breast cancer (BC): The SEQ project

The aim of this study is to assess the performance of the accredited in-house NGS panel VHIO300 in the detection of gPV in tumor sequencing from patients with BC. We selected 90 patients (pts) with BC with prior germline genetic diagnosis and available tumor tissue (primary tumor or metastatic samples); 45 had a gPV in a hereditary breast and ovarian cancer (HBOC) gene and 45 had no gPV detected. Blinded tumor sequencing was carried out using VHIO300 panel. Frequency of true positives (TP), true negatives (TN), false positives (FP) and false negatives (FN) of tumor sequencing was calculated, as well as sensitivity, specificity, positive and negative predictive value (PPV, NPV). A Cohen's kappa coefficient of 0.80 was required. 2 samples (1 from a pt with PALB2 gPV and 1 germline negative) did not fulfill technical requirements for tumor sequencing. So, 88 pts had germline and tumoral sequencing results. The VHIO300 identified the gPV in 40/44 (91%) tumor samples (TP). Four gPV were not identified in tumor sequencing (FN): 2 large rearrangements (LR) (100% of LR) and 2 intronic variants (29% of intronic variants). The 4 variants were identified after being revisited in a targeted manner (Table). Among the 44 pts with no gPV, the VHIO300 identified 3 HBOC-associated PV potentially of germline origin due to their variant allele frequency (1 BRCA1, 2 BRCA2). Ad hoc germline analysis of these 3 PV was negative, so they were considered as FP (7%) (Table). The sensitivity, specificity, PPV and NPV of the VHIO300 to detect gPV was 91%, 93%, 93%, and 91%, respectively. Cohen's kappa coefficient was 0.84 (95% CI 0.73 – 0.95).Table: 166PGeneVariantType of variantVAFBC tumorgPV not detected by tumor sequencing (FN)ATMc.902-19_1065+869del1052LR----Lum B HER2 neggPV not detected by tumor sequencing (FN)CHEK2Deletion of exons 3 and 4LR----Lum B HER2 neggPV not detected by tumor sequencing (FN)CHEK2c.793-1G>AIntronic variant----Lum AgPV not detected by tumor sequencing (FN)BRCA2c.8332-13T>GIntronic variant----Lum B HER2 negPV in HBOC gene in tumoral sequencing not gPV (FP)BRCA1c.5193+1G>TIntronic variant45.32%Triple negPV in HBOC gene in tumoral sequencing not gPV (FP)BRCA2c.8695C>TPoint mt nonsense57.66%Lum B HER2 negPV in HBOC gene in tumoral sequencing not gPV (FP)BRCA2c.8023A>GPoint mt missense35.19%Lum A Open table in a new tab The VHIO300 panel demonstrated acceptable sensitivity and specificity in this BC population, validating its clinical use as a surrogate initial method to identify patients with HBOC gPV. However, if there is a high clinical suspicion of HBOC, germline testing should be performed if no putative gPV is identified in tissue.