Testing for ROS1 in non-small cell lung cancer: a review with recommendations

Abstract

Rearrangements of the ROS1 gene occur in 1–2 % of non-small cell lung cancers (NSCLCs). Crizotinib, a highly effective inhibitor of ROS1 kinase activity, is now FDA-approved for the treatment of patients with advanced ROS1-positive NSCLC. Consequently, focus on ROS1 testing is growing. Most laboratories currently rely on fluorescence in situ hybridisation (FISH) assays using a dual-colour break-apart probe to detect ROS1 rearrangements. Given the rarity of these rearrangements in NSCLC, detection of elevated ROS1 protein levels by immunohistochemistry may provide cost-effective screening prior to confirmatory FISH testing. Non-in situ testing approaches also hold potential as stand-alone methods or complementary tests, including multiplex real-time PCR assays and next-generation sequencing (NGS) platforms which include commercial test kits covering a range of fusion genes. In order to ensure high-quality biomarker testing, appropriate tissue handling, adequate control materials and participation in external quality assessment programmes are essential, irrespective of the testing technique employed. ROS1 testing is often only considered after negative tests for EGFR mutation and ALK gene rearrangement, based on the assumption that these oncogenic driver events tend to be exclusive. However, as the use of ROS1 inhibitors becomes routine, accurate and timely detection of ROS1 gene rearrangements will be critical for the optimal treatment of patients with NSCLC. As NGS techniques are introduced into routine diagnostic practice, ROS1 fusion gene testing will be provided as part of the initial testing package.

Non–Small Cell Lung Cancer, Version 5.2017, NCCN Clinical Practice Guidelines in Oncology

“…80,115,118 Similar to testing for ALK rearrangements, testing for ROS1 is also performed using FISH. 65,116,[119][120][121] NGS can also be used to assess whether ROS1 rearrangements are present, if the platform has been appropriately designed and validated to detect ROS1 rearrangements. 115 Because a companion diagnostic test has not been approved for ROS1, clinicians should use an appropriately validated test to detect ROS1.…”

“…Testing for ROS1 has typically been performed using FISH; however, a validated NGS platform that can detect this gene fusion may also be used. 120 The NCCN panel recommends that EGFR mutation testing be performed as part of broad molecular profiling (eg, multiplex mutation screening assays or NGS). Testing for ALK gene rearrangements can be performed with FISH or with NGS if the platform is validated and can identify gene fusions.…”

Recent Advances in Targeting ROS1 in Lung Cancer

“…The ROS1 D4D6 rabbit monoclonal antibody (Cell Signaling Technology, MA) is commercially available and exhibits sensitivity nearing 100% and specificity ranging between 92–100% (28, 47, 48). However, the ROS1 IHC readout can be more difficult to interpret and operator-dependent compared to ALK IHC for several reasons.…”

Molecular Pathology: A Requirement for Precision Medicine in Cancer

“…Diagnosis is performed using FISH [23]. IHC and PCR are possible; however, so far these have not been standardized.…”