Focal HER2/neu amplified clones partially account for discordance between immunohistochemistry and fluorescence in-situ hybridization results: data from NCCTG N9831 Intergroup Adjuvant Trial

Abstract

568 Background: Accurately and reliably identifying HER2/neu status is a critical priority in the treatment decision process. Current HER2 testing strategies include immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH), but discrepancies between them (3+IHC and FISH-, 0–2+ IHC and FISH+) interfere with interpretation and patient management. We postulate that some of these discrepancies may reflect tumor heterogeneity, derived from focal HER2 amplified clones (FHAC) within a tumor. Methods: As part of the ongoing N9831 “Phase III trial of doxorubicin and cyclophosphamide followed by weekly paclitaxel with or without trastuzumab treatment for women with HER2 overexpressing or amplified node positive or high–risk node negative breast cancer” adjuvant Intergroup trial, breast tumors were evaluated for HER2 status by both IHC and FISH using HercepTest™ and PathVysion™ assays. Samples with FHAC were defined as containing 2% to 40% of cells demonstrating unequivocal amplification (cells with >10 HER2 signals and HER2:CEP17 ratio >5.0), regardless of the overall HER2:CEP17 ratio. HER2/FISH discordant cases were identified and the number with FHAC determined. Results: 2006 samples from 1738 patients were studied. 1169 cases demonstrated FISH amplification (HER2:CEP17 ratio ≥2) and were IHC positive (3+). 114 cases contained FISH amplification, but were IHC negative (0–1+) or equivocal (2+). 2 % of the IHC 3+/FISH amplified, 21% of the IHC 0–1+/FISH amplified, and 30% of the IHC 2+/FISH amplified cases contained FHAC (Table 1). Conclusions: The presence of FHAC seems to be a factor in a substantial number of cases exhibiting discordance between IHC and FISH evaluations of HER2 positivity. The prognosis and response to trastuzumab among patients with FHAC have yet to be determined. Author Disclosure Employment or Leadership Consultant or Advisory Stock Ownership Honoraria Research Funding Expert Testimony Other Remuneration Eli Lilly; Merck; Vysis, Inc.; Wyeth Merck Aventis Genentech