FISH Testing Misses Drug-Targetable EML4-ALK Rearrangements in Lung Adenocarcinoma

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A study published in The Oncologist on February 26, 2015, challenges the role of fluorescence in situ hybridization (FISH) testing as the current gold standard for detecting EML4-ALK rearrangements and offers an alternative algorithm for detecting this drug-targetable mutation in patients with advanced lung adenocarcinoma.

Dr. Pekar-Zlotin and colleagues present an innovative retrospective analysis using newer NGS platforms to confirm that some patients who were negative for ALK rearrangement by the current standard FISH test were positive by IHC and NGS.

A study published in The Oncologist on February 26, 2015, challenges the role of fluorescence in situ hybridization (FISH) testing as the current gold standard for detecting EML4-ALK rearrangements and offers an alternative algorithm for detecting this drug-targetable mutation in patients with advanced lung adenocarcinoma.

After endothelial growth factor receptor (EGFR) mutations, mutations in the fused gene product of echinoderm microtubule-associated protein-like 4 (EML4) and anaplastic lymphoma kinase (ALK) are the second most common drug-targetable genetic abnormality in lung carcinoma. Approximately 3%-7% of lung carcinomas harbor EML4-ALK rearrangements, which are susceptible to treatment with ALK-targeted tyrosine kinase inhibitors (TKIs). Current guidelines recommend testing with an ALK-specific FISH assay to select patients for ALK-targeted TKI therapy based on the presence of EML4-ALK rearrangements. However, immunohistochemistry (IHC) testing detects EML4-ALK rearrangements in a subset of patients with false-negative FISH results who may also benefit from ALK-targeted therapy.

A team of researchers led by Prof Nir Peled, MD, PhD, at the Davidoff Cancer Center in Petach Tiqwa, Israel, compared the accuracy of FISH and IHC as tools for detecting EML4-ALK rearrangements in lung adenocarcinoma. The team also used genetic testing with next-generation sequencing (NGS) to examine tumor samples with discordant FISH and IHC results.

“Our data suggest that the FISH-based approach to detecting EML4-ALK rearrangements in lung cancer may miss a substantial number of patients who could benefit from ALK-targeted therapy,” Dr. Peled said. “Clinicians should strongly consider using IHC to screen for EML4-ALK rearrangements, with follow-up next-generation sequence testing for borderline results.”

In the retrospective study, the research group tested for EML4-ALK rearrangements in lung adenocarcinoma samples from 51 patients treated between 2011 and 2013. All samples underwent testing with FISH and IHC (D5F3 antibody), and those with discordant results underwent further testing with NGS analysis. In total, 4 samples (7.8%) were positive for EML4-ALK by FISH [FISH(+)] while 8 samples (15.7%) were positive by IHC [IHC(+)]. Three samples were FISH(+) IHC(+), while 5 samples were FISH(-) IHC(+). Among the 5 discordant samples, DNA sequencing with the NGS assay confirmed that 4 were positive for ALK rearrangements.

Using the NGS results for reference, the sensitivity and specificity of IHC were 100% and 97.7%, respectively. By comparison, the sensitivity and specificity of FISH were 42.9% and 97.7%, respectively.

In this historical cohort, only 2 of the 5 patients with discordant FISH(-) IHC(+) test results were treated with crizotinib, an ALK-targeted TKI. One patient experienced a complete response to therapy, with a progression-free survival (PFS) duration of 18 months and overall survival in excess of 3.5 years. The second patient had stable disease, with a PFS of 6 months. These findings suggest that patients with ALK FISH(-) IHC(+) lung cancer may benefit from ALK-targeted therapy, the most efficient form of therapy in lung cancer.

Based on these findings, the study authors propose an algorithm for molecular testing to improve the selection of individualized therapy for patients with advanced lung adenocarcinoma. The algorithm includes standard EGFR mutation analysis and IHC D5F3 testing for ALK. If IHC testing is inconclusive for ALK, clinicians should consider ALK genetic analysis with the NGS assay. Using IHC testing has the potential to identify more patients who may benefit from treatment with ALK-targeted TKIs. These cases may be missed based on false-negative results with FISH testing alone. The authors further suggest that the new gold standard test for ALK rearrangement should be NGS rather than FISH.

Lecia V. Sequist, MD, Deputy Editor of The Oncologist, noted, “Dr. Pekar-Zlotin and colleagues present an innovative retrospective analysis using newer NGS platforms to confirm that some patients who were negative for ALK rearrangement by the current standard FISH test were positive by IHC and NGS. Because IHC antibodies can vary and require validation, prospective studies are now needed to examine validated IHC antibodies, FISH and NGS, so that our diagnostic toolbox can expand.”

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About The Oncologist: Established by oncologists to help physicians better manage their practices in an ever-changing environment, The Oncologist® is the official journal of the Society for Translational Oncology (STO). Now celebrating its 20th edition, this internationally peer-reviewed journal focuses on clear and concise interpretation addressing the multimodality diagnosis, treatment, and quality of life of the cancer patient. Each issue is meant to impact the practice of oncology and to facilitate significant communication in the introduction of new medical treatments and technologies. For more information, visit http://www.TheOncologist.com.

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