Abstract
In 2011, the Vysis Break Apart ALK fluorescence in situ hybridization (FISH) assay was approved by the United States Food and Drug Administration as a companion diagnostic for detecting ALK rearrangement in lung cancer patients who may benefit from treatment of tyrosine kinase inhibitor therapy. This assay is the current “gold standard”. According to updated ALK testing guidelines from the College of American Pathologists, the International Association for the Study of Lung Cancer and the Association for Molecular Pathology published in 2018, ALK immunohistochemistry is formally an alternative to ALK FISH, and simultaneous detection of multiple hot spots, including, at least, ALK, ROS1, RET, MET, ERBB2, BRAF and KRAS genes is also recommended while performing next generation sequencing (NGS)-based testing. Therefore, ALK status in a specimen can be tested by different methods and platforms, even in the same institution or laboratory. In this review, we discuss several clinically relevant technical aspects of ALK FISH, including pros and cons of the unique two-step (50- to 100-cell) analysis approach employed in the Vysis Break Apart ALK FISH assay, including: the preset cutoff value of ≥15% for a positive result; technical aspects and biology of discordant results obtained by different methods; and incidental findings, such as ALK copy number gain or amplification and co-existent driver mutations. These issues have practical implications for ALK testing in the clinical laboratory following the updated guidelines.
Highlights
Lung cancer is the most common cause of cancer-related deaths in the United States [1] and in many other countries [2,3]
Camidge et al [16] has shown that analysis of ≥60 cells from ≥4 selected tumor areas of a non-small cell lung cancers (NSCLC) specimen can reach 100% specificity and 100% of sensitivity for anaplastic large-cell lymphoma kinase (ALK) rearrangement testing, the Vysis ALK Break Apart fluorescence in situ hybridization (FISH) probe set used in their study is slightly different for coverages of both 5 ALK and 3 ALK probe from the one approved by the Food and Drug Administration (FDA)
According to the updated molecular testing guidelines for the selection of lung cancer patients for treatment with targeted tyrosine kinase inhibitors recommended by the College of American Pathologists, the International Association for the Study of Lung Cancer and the Association for Molecular Pathology in 2017 [68], as well as the FDA having approved the VENTANA ALK (D5F3) CDx Assay (Roche Diagnostics) for a qualitative detection of ALK protein in NSCLC tissue specimens, both ALK FISH and the ALK IHC are considered as the first line methods for ALK rearrangement testing
Summary
Lung cancer is the most common cause of cancer-related deaths in the United States [1] and in many other countries [2,3]. Camidge et al [16] has shown that analysis of ≥60 cells from ≥4 selected tumor areas of a NSCLC specimen can reach 100% specificity and 100% of sensitivity for ALK rearrangement testing, the Vysis ALK Break Apart FISH probe set used in their study is slightly different for coverages of both 5 ALK and 3 ALK probe from the one approved by the FDA. The ≥15% result has been widely applied for clinical decisions, including patient selection for targeted therapy and the prediction of treatment response, this cutoff value for a positive ALK rearrangement in a NSCLC specimen was initially not defined by a clinical end point Instead, it was established based on an analytical assessment of the background signals (“background noise”) [16,21,22]. Terms such as “borderline negative result (i.e., 10–14%)” and “borderline positive result (i.e., 15–24%)” have been suggested for further analysis of these cases by different methods/technologies (e.g., IHC, NGS or other methods) by others [19,33,34,35]
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