Immunohistochemical Analysis of ALK Fusion-Positive Lung Cancers: Correlation with FISH, Molecular Findings and Morphology.
Shahrazad T Saab, Christine Lovly, Clayton J Hollis, Yenamandra Ashwini, Hongbin Ji, William Pao, Adriana Gonzalez. Vanderbilt University, Nashville, TN; Chinese Academy of Sciences, Shanghai, China
Background: Activating fusions involving the gene encoding anaplastic lymphoma kinase (ALK) have been detected in a subset of lung cancers and define a clinically relevant molecular subset sensitive to ALK tyrosine kinase inhibitors. The majority of ALK fusions involve the upstream partner, EML4. This study correlates immunohistochemical (IHC) staining for ALK with tumor morphology, IHC for other common lung cancer antigens, and FISH and/or RT-PCR in a large cohort of non-small cell lung cancers.
Design: ALK IHC (Cell Signaling, rabbit monoclonal D5F3) was performed on 14 tissue microarrays (TMA) containing 434 adenocarcinomas and 226 squamous cell carcinomas. FISH was performed using an ALK break apart probe (Abbott Molecular, Abbott Park, IL). RT-PCR sequencing used 3 different 5' EML4 primers (that detect all known EML4 variants) and 1 ALK primer that hybridizes to all known ALK fusions. All ALK D5F3 stains were compared with routine IHC for CK7, CK20, TTF-1, CK5/6, p63, CDX-2 mucicarmine, and ALK1 (Dako, monoclonal mouse CD246). The presence of a fusion was verified by either FISH or RT-PCR in all cases.
Results: 8 ALK-positive cases were identified: 4 by TMA, 2 by FISH, 2 by RT-PCR. All cases were adenocarcinoma. 7 tumors were positive by IHC (D5F3 antibody). 5 of these were positive by the Dako antibody with generally weaker positivity and more nonspecific staining in FISH-negative tumors. ALK (D5F3) showed moderate-strong staining in >90% of cells in 5 cases, and weaker staining in >30% of cells in 2 cases. One case with a known EML4-ALK E20;A20 fusion by RT-PCR showed very weak staining in <30% of tumor cells (IHC neg). This sample and the 2 weaker ALK IHC positive cases all showed prominent mucinous features. Other histologic patterns included micropapillary to cribriform or solid architecture, sometimes mixed with mucinous/signet ring cells. 7/8 ALK positive tumors had material for IHC panel correlation and were positive for: CK7(7/7). TTF-1(7/7), p63(4/7), and mucicarmine(4/7), while CK5/6, CK20 and CDX2 were essentially negative.
Conclusions: ALK IHC (D5F3) antibody showed good correlation with FISH and RT-PCR and was easier to evaluate than ALK1 (CD246). Mucinous tumors may show lesser staining with ALK IHC. ALK positive tumor morphology can be suggested by a micropapillary-cribriform-solid architecture especially if mixed with mucinous features. Unlike some mucinous adenocarcinomas of the lung, CDX2 and CK20 were negative in these tumors.
Tuesday, March 1, 2011 1:00 PM
Poster Session IV # 259, Tuesday Afternoon