P1-07-01: Comparison of Four HER2 Testing Methods in the Detection of HER2−Positive Breast Cancer: Results from the FinHer Study Cohort.
Authors: Huang W , Wirtz R , Weidler J , Lie Y , Sherwood T , Leinonen M , Bono P , Isola J , Kellokumpu-Lehtinen P-L , Joensuu H 
Published: June 2012

Abstract
Background: Accurate assessment of the HER2 status is essential for identifying patients who may benefit from HER2 targeted therapy. The current methods, immunohistochemistry (IHC) and in situ hybridization (ISH), determine HER2 status semi-quantitatively as positive (+), equivocal (+/−) and negative (−) with predefined cutoff values. Recent studies have suggested that current HER2 cutoffs may not be optimal for all clinical settings of HER2 targeted therapy. In a small subset of adjuvant NCCTG N9831 patients confirmed as HER2−normal by round-robin review of HER2 testing, trastuzumab benefit was observed (Perez et al, SABCS 2010). Quantification of HER2 as continuous variable may enable a more accurate optimization of HER2 cutoffs for various HER2 targeted therapies. In this study, we measured continuous HER2 protein expression by the HERmark™ assay and continuous mRNA expression by quantitative real time polymerase chain reaction (qPCR), and compared these results with central IHC and central chromogenic in situ hybridization (CISH) results of FinHer.
Methods: Total HER2 protein expression (H2T) was quantified using the HERmark assay as previously described (Huang et al. Am J Clin Pathol 2010;134:303). HER2 mRNA expression (H2N) was measured by qPCR as previously published (Noske et al. Br Cancer Res Treat 2011;126:109). The results of H2T and H2N as continuous variables and as predefined categories were compared with central CISH results from FinHer (Joensuu et al, N Engl J Med 2006;354), and central IHC retesting.
Results: H2T in 899 evaluable samples described a continuum of 0.4 to 721.2 (relative HERmark unit); while H2N in 915 evaluable samples showed a continuum of 31.4 to 42.8 (delta-Ct). Significant correlation between H2T and H2N as continuous variable was found (R2= 0.56, P&lt; .0001). Paired method comparison was performed for samples with valid results in any two of the four testing methods. Overall concordance of H2T and H2N with predefined categories (+, +/−, -) was 81%, and concordance of (+) and (−) subsets was 95% when (+/−) cases (H2T 11%; H2N 6%) were excluded. Overall concordance of central IHC and H2T categories (+, +/−, -) was 75%, and concordance of (+) and (−) subsets was 96% when (+/−) cases (IHC 16%; H2T 11%) were excluded. Overall concordance of IHC and H2N categories (+, +/−, -) was 84%, and concordance of (+) and (−) subsets was 99% when (+/−) cases (IHC 16%; H2N 6%) were excluded. Concordance of central CISH (+, -) with H2T and H2N categories (+, -) was 89% and 91%, respectively, when (+/−) cases were excluded from H2T (13%) and H2N (8%), respectively.
Conclusions: All four methods identified HER2−positive breast cancers. The discordance rate between the methods tested was approximately 10 to 20% despite careful delineation of cancerous tissue in the sample and analysis of adjacent tumor sections. No combination of assays could be identified with concordance rate &gt;95% when the equivocal subsets were included in comparisons. Exclusion of the equivocal subsets (about 10% of samples) yielded high concordance rates of approximately 95% or higher. H2T and H2N showed comparable continuous distribution patterns and significant concordance with standard HER2 status by central IHC and CISH.
Citation Information: Cancer Res 2011;71(24 Suppl):Abstract nr P1-07-01.