MS02.03 Achieving Effective Lung Cancer Genotyping While Balancing Constrained Resources

Abstract

Modern oncologic practice for patients with non-small cell lung carcinoma (NSCLC) demands real time data for an increasing numbers of tumor biomarkers. As a result, laboratories are embracing panel-based approaches to tumor molecular profiling, a trend that is facilitated by the adoption of next generation sequencing (NGS) assays. NGS assays may be designed for highly sensitive and focused detection of hotspot mutations (typically by amplicon sequencing) or broader, more comprehensive profiling for detection of a wide variety of alterations in oncogenes and tumor suppressor genes (typically by hybrid capture technology). There are benefits and drawbacks to both approaches. In general, amplicon sequencing offers speed and sensitivity but limited scope and a propensity for PCR-related artifacts that may adversely affect detection of certain mutation types. In contrast, hybrid capture techniques offer tremendous breadth, enabling capture of 100s of genes to whole genomes, but are limited by sensitivity and longer turnaround time.1 Preanalytic steps including sample acquisition, pathologist review to confirm sample adequacy, and nucleic acid extraction often comprise the majority of the turnaround time required to complete focused tumor tissue molecular profiling. Some assays have been optimized for turnaround times of just a few hours- these assays bypass the separate nucleic acid extraction step,2 but are limited by the few number of mutational targets detected, ultimately requiring the use of multiple simultaneous or sequential focused panels.3 This practice, while perhaps the optimal current model from the standpoint of patient care, may be financially prohibitive for many laboratories. This approach additionally requires careful stewardship of patient tissues, as the use of multiple competing assays and may lead to exhaustion of tumor tissue and incomplete tumor molecular profiling. For patients with limited tissue, a considered plan incorporating input from the treating physician, surgical pathologist, and molecular laboratorian should developed early in order to guide assay priority and ensure adequate tissue is available to confirm any unexpected or contradictory results. When the turnaround time for comprehensive large panel testing is clinically acceptable, this approach may be theoretically more cost effective and in most cases will deliver information for essential and emerging biomarkers.4 It is important to keep in mind, however, that even comprehensive DNA-based assays may lack optimal sensitivity for certain structural variants (fusions, large insertion-deletion events), and it may be necessary to couple these tests with focused RNA-based panels optimized for transcript fusion detection.5 1. Rizzo JM, Buck MJ. Key principles and clinical applications of "next-generation" DNA sequencing. Cancer Prev Res (Phila). 2012;5(7):887-900. 2. Ilie M, Butori C, Lassalle S, et al. Optimization of EGFR mutation detection by the fully-automated qPCR-based Idylla system on tumor tissue from patients with non-small cell lung cancer. Oncotarget. 2017;8(61):103055-103062. 3. Lindeman NI, Cagle PT, Aisner DL, et al. Updated Molecular Testing Guideline for the Selection of Lung Cancer Patients for Treatment With Targeted Tyrosine Kinase Inhibitors: Guideline From the College of American Pathologists, the International Association for the Study of Lung Cancer, and the Association for Molecular Pathology. J Mol Diagn. 2018;20(2):129-159. 4. Sireci AN. Single Genes, Panels, and Next-Generation Sequencing Platforms: A Financial Perspective. Arch Pathol Lab Med. 2018;142(7):790-791. 5. Benayed R, Offin M, Mullaney K, et al. High Yield of RNA Sequencing for Targetable Kinase Fusions in Lung Adenocarcinomas with No Mitogenic Driver Alteration Detected by DNA Sequencing and Low Tumor Mutation Burden. Clin Cancer Res. 2019. amplicon sequencing, hybrid capture, turnaround time