Success and failure rates of tumor genotyping techniques in routine pathological samples with non-small-cell lung cancer
Introduction
In the United States in 2013, the expected number of new cases of and deaths from lung cancer will exceed 220,000 and 159,000, respectively [1]. The overall, of all stages combined, five-year survival for the most prevalent form of lung cancer – non-small-cell lung cancer (NSCLC) – does not exceed 15% despite use of surgical resection, radiotherapy and systemic chemotherapy [1]. The last decade of research in lung cancer has yielded important advances in the development of targeted therapies that target driver oncogenes [2]. The most prevalent mutated or rearranged oncogenes identified in non-small cell lung cancers (NSCLCs) are v-ki-ras2 Kirsten rat sarcoma viral oncogene homolog (KRAS), epidermal growth factor receptor (EGFR), anaplastic lymphoma kinase (ALK), ROS1, BRAF, ERBB2 and RET [3]. Specifically, mutations in EGFR and rearrangements (either inversions or translocations) involving ALK are part of the pathogenesis of some lung adenocarcinomas, predominantly in never and/or light smokers, and predict for improved outcomes with tyrosine kinase inhibitors (TKIs), such as erlotinib and crizotinib, respectively, that target these aberrant kinases [4], [5], [6]. As such, molecular testing of lung cancer specimens has become part of routine clinical practice in the care for patients with advanced NSCLC.
Currently, the proposed testing guidelines from the College of American Pathologists, the International Association for the Study of Lung Cancer, the Association for Molecular Pathology, and the National Comprehensive Cancer Network recommend testing all advanced NSCLCs with an adenocarcinoma component at the time of diagnosis for EGFR mutation and for ALK fluorescence in situ hybridization (FISH) analysis [7], [8]. These specimens are more frequently encountered as small biopsy or cytology specimens derived from either the primary tumor or from lymph node or distant metastatic sites, and processed as formalin-fixed, paraffin embedded tissue samples. Limited tumor cellularity in small biopsy or cytology specimens can lead to molecular testing failure. Additionally, it is recognized that different tissue processing techniques, including acid decalcification or heavy metal fixatives, can cause DNA degradation and impede molecular analysis [9]. These issues can influence the selection and success rates of specimens submitted for mutational analysis when considering sampling of bony lesions. Practically, this can pose a dilemma for the clinician and pathologist alike, in determining whether an initial biopsy used to establish the diagnosis of NSCLC is sufficient for additional molecular testing, or whether another specimen must be procured for such a purpose.
As molecular testing of NSCLCs has become a standard of clinical practice, various institutions have reported their testing protocol and experience [10], [11], [12], [13]. From these studies, one can occasionally glean the failure rates of their specimens, though to our knowledge a systematic study of genetic testing failure rates for NSCLC using commercially available testing in a typical clinical practice setting has not been published to date. Here, we present the molecular testing efficiency for EGFR, KRAS, and ALK FISH analysis of clinical specimens from NSCLC patients from our institution over a five year time period, reflecting real-life clinical practice experience.
Section snippets
Patient selection
Patients with a diagnosis of lung cancer who were seen by our providers and whose tumors were genotyped for at least EGFR mutations were identified through an ongoing Institutional Review Board (IRB) approved protocol at Beth Israel Deaconess Medical Center (BIDMC2009-P-000182). Patients and tumor pairs were excluded if genotyping was not performed. There were 381 patient-tumor specimens that were submitted to a commercial vendor for tumor genotype techniques between 2007 and 2012. The data cut
Patient and tumor characteristics
Table 1 summarizes the clinical and pathological characteristics of the 381 patient-tumor pairs that were included in our cohort.
Success and failure rates of tumor genotype techniques
Out of the 381 samples, all were sent for EGFR mutation analysis, 226 for KRAS mutation analysis, and 275 for ALK FISH (Table 1). In 207 tumors (54.3%), all three tests were ordered simultaneously. Table 2 details the success and failure rates for EGFR, KRAS, and ALK analyses techniques. The success rate for EGFR mutation analysis was 94.2% and the failure rate 5.8%;
Discussion
We have shown that our institutional success rate for EGFR, KRAS, and ALK testing of clinical lung cancer specimens exceeds 90% using routine tissue samples and a commercial vendor. As molecular testing of non-squamous, non-small cell lung carcinomas has become the standard of care for patients with advanced stage lung cancer, numerous studies by high-volume academic centers have been published outlining mutation frequency in different patient populations [17]. Less has been published regarding
Funding
This work was funded in part through a fellowship from the American Society of Clinical Oncology Conquer Cancer Foundation (DBC), an American Cancer Society grant (RSG 11-186 to DBC), a Lung Cancer Foundation of America-International Association for the Study of Lung Cancer grant (to DBC), and National Institutes of Health (NIH) grant CA090578 (to DBC).
Conflict of interest statement
Daniel B. Costa has received consulting fees from Pfizer, Roche and AstraZeneca. Paul A. VanderLaan, Erik Folch, David H. Boucher, Michael S. Kent, Sidharta P. Gangadharan, Adnan Majid, Olivier N. Kocher, Michael A. Goldstein, Mark S. Huberman have no conflicts to disclose. No other conflict of interest is stated.
Acknowledgements
We would like to thank all current and former members of the Thoracic Oncology Clinic at Beth Israel Deaconess Medical Center, and our patients.
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These authors contributed equally.