Clinical Validation and Implementation of a Targeted Sequencing Panel for Predisposition to Inherited Cancer

Next generation sequencing is a powerful technology that provides a wealth of information, however, for clinical laboratories development, validation and implementation of NGS assays is challenging due to costs, data analysis pipelines, and turnaround time. Smaller academic clinical laboratories may not have resources to develop individual gene panels and, therefore, require the use of targeted sequencing panels designed to cover multiple indications increasing incidental findings. In this ongoing validation, we are using the TruSight Cancer panel (Illumina) for use in patients at risk for Lynch syndrome. We tested 12 samples from patients with known germline mutations in a Lynch syndrome indicated gene (MLH1, MSH2, MSH6, PMS2, EPCAM) to evaluate assay performance characteristics. The data was pre-filtered for these 5 genes, and 100% of exonic bases were covered at 20× with 95% covered at >100×. On average there were 7 variants per sample requiring further evaluation and characterization. Pathogenic variants in 8 patients were identified, including SNVs, and small exonic and splice variant indels. Larger exonic deletions in the remaining 4 patients would require additional complementary del/dup testing in the absence of pathogenic variants detected by NGS. The TruSight Cancer panel consists of 94 genes that have been indicated in hereditary cancer. By pre-filtering the data for genes of interest, it is possible to use this assay to evaluate a limited panel of genes and reduce the concern for incidental findings. This allows for the gradual validation and implementation of new clinical targeted sequencing panels with the same performance characteristics and methodologies.