Abstract 809: nCounter for detection of clinically relevant alterations in exosomes of non-small cell lung cancer cells and patients

Background: ALK, ROS1 and RET fusions and MET exon 14 skipping variant (METex14) are present in 10-15% of advanced non-small-cell lung cancer (NSCLC) patients and their accurate identification is critical to guide targeted therapies. In a significant number of cancer patients, the tumor tissue available is insufficient for genetic analysis and repeated tissue biopsies for monitoring the course of the disease and the emergence of resistance are not feasible. Liquid biopsies constitute the only alternative available in these cases, but NGS techniques have shown insufficient sensitivity for fusion detection in blood samples. The nCounter technology has been adapted to detect fusions and skipping variants in FFPE tumor biopsies and we aimed to validate it for exosomes. Methods: Exosomes were purified using a miRCURY kit (Qiagen) and RNA was extracted using the TRI reagent (MRC Inc). A customized nCounter panel (Nanostring) for detection ALK, ROS1 and RET fusion transcripts and MET ex14 mRNA was used with a 10-cycles preamp step. First, proof-of-concept experiments were run by testing exosomes isolated from the culture medium of cell lines. Next, we tested exosomes isolated from the blood of NSCLC patients with know genotypes. Results: nCounter fusion probes successfully detected ALK, RET and ROS1 fusion transcripts in exosomes isolated from the culture medium of the cell lines H3122 (EML4-ALKv1), H2228 (EML4-ALKv3), HCC78 (SLC34A2-ROS1) and LC/2-Ad (CCDC6-RET). Exosomes from a cell line established from a patient progressing to alectinib were also positive for EML4-ALKv1and showed high MET expression levels, while exosomes from the fusion-negative cell lines A549 and H23 tested negative. Finally, fusion transcripts were detected in exosomes purified from the blood of fusion positive NSCLC patients but not in fusion negative cases. Conclusions: nCounter can detect ALK, RET and ROS1 fusion transcripts in exosomes purified from the blood of advanced NSCLC patients Citation Format: Ana Giménez Capitán, Jill Bracht, Chung-Ying Huang, Cristina Teixidó, Noemí Reguart, Rich Boykin, Sarah Warren, Joseph M Beechem, Santiago Viteri Ramirez, Juan José García, Andrés Aguilar, Rafael Rosell Costa, Jay Gerlach, Miguel Angel Molina-Vila. nCounter for detection of clinically relevant alterations in exosomes of non-small cell lung cancer cells and patients [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and