Quantitative measurements of HER2 expression and HER2 homodimer using a novel proximity based assay: comparison with HER2 status by immunohistochemistry and chromogenic in situ hybridization in the FinHer study.
Authors: Joensuu H , Weidler J , Lie Y , Sherwood T , Whitcomb J , Leinonen M , Bono P , Isola J , Kellokumpu-Lehtinen P , Bates M , Huang W 
Published: December 2010

Abstract #2071

Background: The accuracy and reliability of current methods, immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH), to assess HER2 status has recently been a subject of debate. The best method to assess HER2 status remains controversial. We developed a novel assay (HERmark, Monogram Biosciences) that provides precise quantification of HER2 expression (H2T) and HER2 homodimer (H2D) in formalin-fixed paraffin-embedded (FFPE) tissues. We compared H2T and H2D to local IHC, central chromogenic in situ hybridization (CISH) and central IHC retesting of breast cancers from the FinHer study.&#x2028; Methods: H2T and H2D were detected through light-dependent release of fluorescent tags (VeraTag reporters) conjugated to a HER2 antibody, requiring proximity to a second HER2 “scissors” antibody. The VeraTag signal was quantified by capillary electrophoresis and normalized to tumor area. Assay comparisons correlated H2T and H2D with HER2 testing by local IHC and central CISH from FinHer (Joensuu et al, N Engl J Med 2006;354), as well as central HER2 status reassessment by combination of externally performed central IHC retesting (PhenoPath labs, Seattle, WA) and central CISH (FinHer) according to ASCO/CAP guideline for HER2 testing in breast cancer (Wolff et al, Arch Pathol Lab Med 2007;131).&#x2028; Results: H2T and H2D in 899 evaluable FinHer samples described a continuum over a wide dynamic range (∼ 3 logs), in contrast with conventional IHC categories (0-3+). The correlation between H2T and IHC categories was significant (P &lt; .0001). Overlap of H2T among the IHC categories was observed. H2D showed a similar correlation with IHC and a general correlation with H2T (P &lt; .0001). A H2T cutoff value, based on its ability to distinguish high and low responders in a cohort of metastatic breast cancer patients treated with trastuzumab-based regimens (log10 H2T= 1.14, Leitzel et al, 2008 ASCO, abstract 1002), was used to define HERmark negative (-) and positive (+), which were then compared with IHC and CISH results. The concordances between HERmark (-) and local IHC (-), central CISH (-), and central HER2 reassessment (-) were 89%, 84%, and 91%, respectively. The concordances between HERmark (+) and local IHC (+), central CISH (+), and central HER2 reassessment (+) were 71%, 89%, and 92%, respectively. The HERmark test showed greater overall concordance with central HER2 reassessment (91%) than with local IHC (84%) and central CISH (87%)&#x2028; Conclusions: HERmark reliably measures H2T and H2D in FFPE tissues. H2T showed excellent concordance with central HER2 status according to ASCO/CAP guideline for HER2 testing. The precise quantification of H2T and H2D may provide novel, quantifiable, predictive tests for targeted HER2 therapy.

Citation Information: Cancer Res 2009;69(2 Suppl):Abstract nr 2071.