Abstract 2841: Characterization of ALK fusions in circulating tumor cells (CTCs) of NSCLC

Abstract

Abstract Background: Many NSCLC patients have insufficient tumor sample or have high co-morbidities preventing access to tissue biopsies for FISH or IHC to determine ALK fusion and subsequent eligibility to Crizotinib therapy. We developed a gene/protein assay for the ALK fusion and examined CTCs and CTC subpopulation incidence and molecular characterization in newly diagnosed NSCLC patients. Methods: 10 samples from newly diagnosed NSCLC patients prior to therapy were recruited and blood specimens were collected and shipped to Epic Sciences. 2/10 had known ALK rearrangement through tissue analysis. All nucleated cells were plated onto glass slides and subjected to IF staining and CTC identification by fluorescent scanners and algorithmic analysis. CTCs, defined as classic (CK+ CD45- w/intact DAPI nuclei and morphologically distinct), apoptotic (CK+, CD45-, non-intact nuclei) and CK- (CK-, CD45-, intact and distinct) were identified. Samples were characterized with ALK IF to assess expression. Patients with known ALK rearrangements in tissue also had their CTCs assessed with ALK FISH. Results: Assays for the ALK protein and ALK gene rearrangement were developed and specificity confirmed utilizing H2228 (ALK positive) & A549 (ALK negative) cells spiked into donor blood and run through the Epic Assay. Expression of ALK fusions was visualized in CTCs of 1/2 patients with known ALK rearrangements and 0/8 patients with no ALK rearrangements. ALK fusion protein expression was seen in classic, apoptotic, small and CK- CTCs. Conclusion: ALK fusion protein and ALK rearrangement assessment with FISH utilizing the Epic CTC platform demonstrates sensitivity and specificity to identification of patients eligible to Crizotinib. The identification of ALK in CTC subpopulations identifies unique tumor cell morphology and suggests evidence of epithelial plasticity. Further clinical studies are ensuing to determine the sensitivity and specificity of ALK in CTCs and the relevance of CTC subpopulations related to Crizotinib therapy and NSCLC progression. Citation Format: David Lu, Rachel Krupa, Natalee Bales, Jessica Louw, Dena Marrinucci, Ryan Dittamore. Characterization of ALK fusions in circulating tumor cells (CTCs) of NSCLC. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 2841. doi:10.1158/1538-7445.AM2014-2841