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J Thorac Oncol. 2018 Oct;13(10):1474-1482. doi: 10.1016/j.jtho.2018.05.041. Epub 2018 Jun 20.

Comparison of Molecular Testing Modalities for Detection of ROS1 Rearrangements in a Cohort of Positive Patient Samples.

Author information

1
Department of Pathology, University of Colorado - Anschutz Medical Campus, Aurora, Colorado.
2
Department of Medicine - Division of Medical Oncology, University of Colorado - Anschutz Medical Campus, Aurora, Colorado.
3
Department of Pediatrics - Section of Hematology, Oncology, and Bone Marrow Transplant, University of Colorado - Anschutz Medical Campus, Aurora, Colorado.
4
Department of Pathology, University of Colorado - Anschutz Medical Campus, Aurora, Colorado; Department of Medicine - Division of Medical Oncology, University of Colorado - Anschutz Medical Campus, Aurora, Colorado.
5
Department of Medicine - Division of Medical Oncology, University of Colorado - Anschutz Medical Campus, Aurora, Colorado. Electronic address: robert.doebele@ucdenver.edu.

Abstract

INTRODUCTION:

ROS1 gene fusions are a well-characterized class of oncogenic driver found in approximately 1% to 2% of NSCLC patients. ROS1-directed therapy in these patients is more efficacious and is associated with fewer side effects compared to chemotherapy and is thus now considered standard-of-care for patients with advanced disease. Consequently, accurate detection of ROS1 rearrangements/fusions in clinical tumor samples is vital. In this study, we compared the performance of three common molecular testing approaches on a cohort of ROS1 rearrangement/fusion-positive patient samples.

METHODS:

Twenty-three ROS1 rearrangement/fusion-positive clinical samples were assessed by at least two of the following molecular testing methodologies: break-apart fluorescence in situ hybridization, DNA-based hybrid capture library preparation followed by next-generation sequencing (NGS), and RNA-based anchored multiplex polymerase chain reaction library preparation followed by NGS.

RESULTS:

None of the testing methodologies demonstrated 100% sensitivity in detection of ROS1 rearrangements/fusions. Fluorescence in situ hybridization results were negative in 2 of 20 tested samples, the DNA-based NGS assay was negative in 4 of 18 tested samples, and the RNA-based NGS assay was negative in 3 of 19 tested samples. For all three testing approaches, we identified assay characteristics that likely contributed to false-negative results. Additionally, we report that genomic breakpoints are an unreliable predictor of breakpoints at the transcript level, likely due to alternative splicing.

CONCLUSIONS:

ROS1 rearrangement/fusion detection in the clinical setting is complex and all methodologies have inherent limitations of which users must be aware to correctly interpret results.

KEYWORDS:

FISH; ROS1; lung cancer; molecular testing; next-generation sequencing

PMID:
29935306
PMCID:
PMC6286810
[Available on 2019-10-01]
DOI:
10.1016/j.jtho.2018.05.041

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