Vysis ALK non-small cell lung cancer assay performance on archival tissue and cytology specimens.

Abstract

e21060 Background: Rearrangements of ALK gene on chromosome 2p23 create a constitutively activated tyrosine kinase implicated in the development of NSCLC and targeted by novel therapeutic agents. The Vysis ALK Break Apart (BAP) FISH Probe assay was developed to detect chromosome 2p23 rearrangements in NSCLC FFPE tissue by fluorescence in situ hybridization. Performance of the ALK assay was assessed on FFPE tissue specimens and FFPE cell lines. The ALK probe was also tested on lung cytology cell suspensions. Methods: The ALK FISH assay success rate on primary NSCLC was tested with 30 FFPE tumor resection specimens, four 5-μm section slides each, judged by qualitative rating of fluorescent signal specificity, intensity, and background. Assay performance was further assessed on 15 FFPE biopsy specimens of 4 metastatic sites including mediastinal and axillary lymp nodes, pleural cavity, brain, and adrenal gland. Each specimen was tested in duplicate and evaluated by 2 reviewers. Assay reproducibility on FFPE cell pellets was tested with 3 lots of ALK Negative and ALK Positive Control Slides (cell lines) on 5 non-consecutive days. Slides were enumerated by 3 reviewers to determine % ALK positive cells, to a total of 45 results per control slide type. Additionally, the ALK BAP probe was tested with a modified assay protocol on ThinPrep lung cytology samples from 8 specimens. Results: The assay success rate on FFPE resection specimens was 99.2 % (90% CI 96.1% - 100%). The core biopsy specimens yielded 95% overall success rate. All Control Slides (FFPE cell lines) tested in the reproducibility study were enumerable within assay ranges, and were correctly classified per specifications of 0-8% and ≥20% ALK positive cells in ALK Negative and ALK Positive Control Slides, respectively. All of the lung cytology specimens prepared by ThinPrep hybridized successfully, however 2 specimens had low cellularity for ALK enumeration. Conclusions: The Vysis ALK BAP FISH assay demonstrated reproducible performance on FFPE cell lines and was shown to yield interpretable results on FFPE lung tissue specimens retrieved by resection or biopsy, from primary tumor or metastatic sites. Feasibility using lung cytology cell suspensions was demonstrated.