Molecular Testing Strategies for Pulmonary Adenocarcinoma An Optimal Approach With Cost Analysis Lester J. Layfield, MD; Richard D. Hammer, MD; Sandra K. White, MS; Larissa V. Furtado, MD; Robert L. Schmidt, MD, PhD  Context.—Molecular analysis of lung adenocarcinoma for therapeutically important genes is standard of practice, with multiple professional organizations recommending testing of all adenocarcinomas for mutations in EGFR, ALK, and ROS1. Some organizations recommend analyzing these genes in association with a panel. Few data exist as to optimal testing method or optimal sequence of testing from a cost perspective. Objective.—To determine which order of gene testing was least costly and whether sequential, small panel, or next-generation sequencing (NGS) was cheapest. Design.—Recent recommendations propose a set of essential molecular tests (EGFR, ALK, and ROS1) and an optional set of molecular tests that may be useful for selection of clinical trials. We compared the costs of different testing sequencing strategies for both the 3 essential genes and for 5 optimal genes. Testing costs were determined by a survey of prices from large laboratories. The strategy most frequently rated as the lowest cost strategy was designated the optimal testing strategy. Results.—Sequential testing of the essential genes in the order EGFR-ROS1-ALK was optimal from a cost perspec- tive. The expected cost of sequential testing was $2227 (95% CI, $1733–$2794). The cost of NGS was $2500. The expected cost per positive result was $11,362 using this strategy. Conclusions.—Molecular testing of lung adenocarcino- mas for the set of 3 essential genes and 5 optional genes can be performed by a variety of methods and in a variety of sequences. From a cost perspective, sequential testing in the order EGFR, ROS1, then ALK is optimal. NGS would be competitive if the price was less than $2200. NGS is optimal if testing for the 3 essential genes will be followed by testing for the 5 optional genes. NGS testing is optimal if the clinician plans to test both essential and optional genes. (Arch Pathol Lab Med. 2019;143:628–633; doi: 10.5858/ arpa.2018-0218-OA) R ecent advances in personalized medicine have resulted in the development of a number of targeted therapies for pulmonary adenocarcinomas. The initial therapeutic targets were pulmonary adenocarcinomas associated with mutations in epidermal growth factor receptor (EGFR) and anaplastic lymphoma kinase (ALK) genes. To reliably target these therapies, the American Society of Clinical Oncologists (ASCO), College of American Pathologists (CAP), and the International Association for the Study of Lung Cancer and Association for Molecular Pathology developed a set of recommendations for tumor testing. 1,2 These recommendations were based on the assumption that mutations in each gene would be tested individually. They recommended that EGFR should be tested first followed by testing for ALK translocations if EGFR was negative. 1,2 Precise recommendations for Kirsten rat sarcoma (KRAS) gene testing were less well developed. These testing recommendations used polymerase chain reaction methodology for EGFR testing and fluorescence in situ hybridization (FISH) techniques for ALK translo- cations. Since the development of those guidelines, a number of new targeted therapies have been developed, including ones based on molecular abnormalities in ROS1 and MET. These therapies also require molecular testing to select appropriate patients for treatment. The National Comprehensive Cancer Network (NCCN) published clinical practice guidelines for the treatment of pulmonary adenocarcinoma in 2017. 3 These guidelines recommended molecular testing for EGFR, ALK, and ROS1. They stated that such testing should be part of a broad molecular profile. Unfortunately, the ‘‘broad molecular profile’’ was not defined and the clinical utility of such a profile was not discussed. The NCCN 2017 guidelines panel strongly advised broader molecular profiling with the goal of identifying rare driver mutations for which effective drugs may already be available or to appropriately counsel patients regarding the availability of clinical trials. ASCO/ CAP have updated their molecular testing recommenda- Accepted for publication July 12, 2018. Published online December 21, 2018. From the Department of Pathology & Anatomical Sciences, University of Missouri, Columbia (Drs Layfield and Hammer); and the Department of Pathology and ARUP Laboratories, University of Utah, Salt Lake City (Ms White and Drs Furtado and Schmidt). The authors have no relevant financial interest in the products or companies described in this article. Corresponding author: Lester J. Layfield, MD, Department of Pathology and Anatomical Sciences, University of Missouri, One Hospital Drive, M263 Medical Sciences Bldg, Columbia, MO 65212 (email: layfieldl@health.missouri.edu). 628 Arch Pathol Lab Med—Vol 143, May 2019 Genetic Testing for Pulmonary Adenocarcinoma—Layfield et al