Correlation of (D4D6) Immunohistochemistry with Fluorescence In Situ Hybridization Assay in a Contemporary Cohort of Pulmonary Adenocarcinomas

Sambit K. Mohanty  Repressor of Silencing ( ) gene rearrangement in the lung adenocarcinomas is one of the targetable mutually exclusive genomic alteration. Fluorescence in situ hybridization (FISH), immunohistochemistry (IHC), next-generation sequencing, and reverse transcriptase polymerase chain reaction assays are generally used to detect gene alterations. We evaluated the correlation between IHC and FISH analysis considering FISH as the gold standard method to determine the utility of IHC as a screening method for lung adenocarcinoma.  A total of 374 advanced pulmonary adenocarcinoma patients were analyzed for IHC on Ventana Benchmark XT platform using D4D6 rabbit monoclonal antibody. FISH assay was performed in parallel in all these cases using the Vysis Break Apart FISH probe.  The sensitivity, specificity, positive and negative likelihood ratios, positive and negative predictive values, and accuracy were evaluated.  A total of 17 tumors were positive either by IHC or FISH analysis or both (true positive). Four tumors were positive by IHC (H-score range: 120-270), while negative on FISH analysis (false positive by IHC). One tumor was IHC negative, but positive by FISH analysis (false negative). The sensitivity, specificity, positive likelihood ratio, negative likelihood ratio, positive predictive value, negative predictive value, and accuracy were 94.4% (confidence interval [CI]: 72.71-99.86%), 63.6% (CI: 30.79-89.07%), 2.6 (CI: 1.18-5.72), 0.09 (CI: 0.01-0.62), 80.95% (CI: 65.86-90.35%), 87.5% (CI: 49.74-98.02%), and 82.76%, respectively.   IHC has high sensitivity at a cost of lower specificity for the detection of gene rearrangement. All IHC positive cases should undergo a confirmatory FISH test as this testing algorithm stands as a reliable and economic tool to screen rearrangement in lung adenocarcinomas.