Diagnostic evaluation of HER-2 as a molecular target: An assessment of accuracy and reproducibility of laboratory testing in large, prospective, randomized clinical trials

Purpose: To critically assess the accuracy and reproducibility of human epidermal growth factor receptor type 2 (HER-2) testing in outside/local community-based hospitals versus two centralized reference laboratories and its effect, on selection of women for trastuzumab (Herceptin)based clinical trials. Experimental Design: Breast cancer specimens from 2,600 women were prospectively evaluated by fluorescence in situ hybridization (FISH) for entry- into Breast Cancer International Research Group (BCIRG) clinical trials for HER-2-directed therapies. Results: HER-2 gene amplification by FISH was observed in 657 of the 2,502 (26%) breast cancers successfully analyzed. Among 2,243 breast cancers with central laboratory immunohistochemistry (10H8-IHC) analysis, 504 (22.54%) showed overexpression (2+ or 3+). Outside/ local laboratories assessed HER-2 status by immunohistochemistry in 1,536 of these cases and by FISH in 131 cases. Overall, the HER-2 alteration status determined by outside/local immunohistochemistry showed a 79% agreement rate [kappa statistic, 0.56; 95% confidence interval (95%,Cl), 0.52-0.60], with FISH done by the central laboratories. The agreement rate comparing BCIRG central laboratory 10H8-IHC and outside/local laboratory immunohistochemistry was 77.5% (K statistic, 0.51; 95% Cl, 0.46-0.55). Finally, HER-2 status, determined by unspecified FISH assay methods at outside/local laboratories, showed a 92% agreement rate (kappa statistic, 0.83; 95% Cl, 0.73-0.93), with FISH done at the BCIRG central laboratories. Conclusions: Compared with the HER-2 status determined at centralized BCIRG reference laboratories, these results,indicate superiority of FISH to accurately and reproducibly assess tumors-for the HER-2 alteration at outside/local laboratories for entry to clinical trials.