A new diagnostic method based on RT-PCR and CLART technology for non-small cell lung cancer (NSCLC) to detect major EML4-ALK and ROS.1 translocations: CLART CMA ALK and ROS.1.

Abstract

e20045 Background: EML4-ALK and ROS1 translocations represent 3-6% and 1-2% of NSCLC patients, respectively. These patients would benefit for a tyrosine kinase inhibitor therapies. Currently, the gold standard for ALK and ROS.1 detection are break-apart fluorescence in situ hybridization (FISH) or inmunodetection (IHC) of the chimeric fusion proteins. FISH has both high sensitivity and specificity, although often present technical interpretation problems due to signal instability and scoring. IHC is a rapid and low cost technique; however it presents several challenges regarding sensitivity and reproducibility. CLART CMA ALK & ROS.1 kit is based RT-PCR and detection of the amplified product was carried out by a low-density microarray platform: CLART (Clinical Arrays Technology) of major fusion transcripts of EML4-ALK and ROS.1 translocations ( V1: E13;A20 , V6: E13; ins69;A20 , V2: E20;A20 , V3a: E6:A20 , V3b: E6;ins33 A20 ,V5a: E2;A20 and V5b: E2;ins117A20 for EML4-ALK and SDC4-ROS1 S2;R32/ S2;R34, CD74-ROS1 C6;R34 and SLC34A2-ROS1 S4;R32/S4;R34). Methods: The translocations analysis was performed with CLART CMA ALK & ROS.1 kit. Clinical testing was performed using the RNA extracted from 115 samples of NSCLC patients being 34 patients with translocations detected in ALK or ROS.1 genes and 81 patients without any translocations. The results were cross-checked by comparison to Next GenerationS methodology performed on the PGM platform with the Ion Ampliseq Panel Oncomide Solid Tumor Fusion and the concordance was 95.6%. Results: Analytical sensitivity was assessed using fusion transcripts cloned in recombinant plasmids. Results ranged from 10 to 100 copies/5µl of recombinant plasmids. The analysis of 115 clinical samples allowed establishing the diagnostic sensitivity and specificity in 90.9-100% and 99-100% respectively. Conclusions: CLART CMA ALK & ROS.1 emerges as a suitable alternative of FISH/IHQ for diagnosis of the most common translocations in EML4-ALK and ROS.1 genes in NSCLC samples and therefore selecting patients that would best benefit from a target therapy.