P3.02a-010 Evaluation of Aberrant ALK Expression in Lung Cancer by RT-PCR and Comparison with FISH and Immunohistochemistry: Topic: ALK Biomarkers

Abstract

In advanced lung cancer patients the gold standard for detecting ALK gene rearrangements is fluorescence in situ hybridization (FISH), and ALK protein expression can be also evaluated by immunohistochemistry (IHC). A single analysis performed alone may not detect all the ALK-positive cases and some patients with discordant FISH and IHC respond to tyrosine kinase inhibitors (TKIs). In this study we evaluated ALK aberrant expression in lung cancer patients by a reverse-transcription (RT)-PCR, to investigate its clinical utility and its concordance with FISH and IHC. ALK aberrant expression was retrospectively investigated on RNA from formalin-fixed paraffin-embedded tissue (FFPEt) of 24 advanced lung adenocarcinoma patients, previously evaluated by FISH and IHC. We used a one-step Scorpion RT-PCR that allowed in a single reaction either the mRNA reverse transcription and the cDNA amplification for ALK kinase-domain, normally not expressed, and a control gene, to assess RNA quality. Results are reported in Table 1. Despite the instability of mRNA from FFPEt, only 2 samples resulted inadequate for RT-PCR. RT-PCR was in disagreement with both FISH and IHC in one case, which is likely to be a RT-PCR false positive. RT-PCR did not detect ALK aberrant expression in a FISH positive case, which was negative also by IHC; unfortunately, this patient died after a cycle of pemetrexed therapy, before undergoing a second line TKI treatment. The presence of ALK rearrangements does not necessarily imply increased protein levels, because of the complex transcriptional and post-transcriptional regulations, so further analysis at RNA levels may clarify discrepancy between FISH and IHC allowing a better stratification of patients who could benefit from TKIs. Therefore, according to our results the RT-PCR evaluating ALK aberrant expression regardless of the fusion partners should be considered for introduction into routine ALK testing in lung cancer.