Abstract P1-03-02: ASCO/CAP human epidermal growth factor receptor-2 (HER2) in situ hybridization (ISH) categories evaluated by quantitative HER2 protein diagnostic methodologies: A comparative analysis
Authors: Buscaglia B , Turner B , Goda H , Huang W , Leitzel K , Natori T , Nakano Y , Okada H , Sperinde J , Ali S , Vasekar M , D'Aguiar M , McMahon L , Henry J , Lipton A , Hicks D 
Published: May 2019

Abstract
Background: In 2013, the ASCO/CAP consensus panel published updated guidelines for HER2 testing in breast cancer that modified the definition of HER2 amplification by in situ hybridization (ISH), creating five new prognostic categories (group 1: classic amplified, group 2: monosomy, group 3: co-amplified (polysomy), group 4: equivocal, and group 5: classic non-amplified). Patients determined to be ISH amplified, were considered eligible for HER2-directed therapy. Concern over whether patients from non-classic groups 2-4 would benefit from treatment has led to the recent publication of the 2018 HER2 focused update. This update has modified the criteria for interpreting these ISH categories, recommending that the final diagnosis take into consideration a combination of HER2 immunohistochemistry (IHC) and ISH results. With increased emphasis on the HER2 protein assessment, it has prompted us to quantitatively examine HER2 protein expression in the ISH categories, using two different novel technologies. Materials & Methods: A cohort of 170 cases (URMC) and 102 cases (PSHMC) of invasive breast cancers, which had previously undergone HER2 IHC and ISH testing, were selected for this study. Cases were sorted and categorized into the HER2 ISH categories defined by ASCO/CAP. HER2 protein expression was quantitatively measured in the URMC and PSHMC cohorts using a novel immunodetection methodology (streptavidin-coated Phosphor-Integrated Dot (PID) fluorescent nanoparticles), and a novel dual-antibody, proximity-binding immunoassay (HERmark® Breast Cancer Assay, Monogram Biosciences, South San Francisco, California), respectively. HER2 protein expression was compared to the HER2 FISH and IHC results by ASCO/CAP category. Results: Cases in group 1 had a significantly (p &lt; 0.01) higher average PID/cell and HERmark compared to cases in groups 2-5 (Table 1). Cases in groups 2-4 showed lower quantitative levels of HER2 protein expression, similar to the classic non-amplified cases (group 5). Group 1 was further divided into three subgroups (Table 2): Group A - ISH high-level amplified (ratio &gt; 2, HER2 &gt; 6, CEP17 &lt; 2.7), Group B - amplified with elevated CEP17 (ratio &gt; 2, CEP17 &gt; 2.7), and Group C - low-level amplified (ratio &gt; 2, HER2 &gt; 4 and &lt; 6). Group A and B had a significantly (p &lt; 0.01) higher average PID/cell and HERmark compared to Group C. Group C was more comparable to cases in groups 2-5 (Table 1). Conclusion: Our results suggest that quantitative assessment of HER2 protein expression may help to further classify cases for HER2 status for targeted therapy, supporting the 2018 ASCO/CAP recommendation that non-classic ISH results might be resolved by evaluating protein expression. Follow up studies with a larger patient cohort and dual quantitative assessment are warranted.
Average PID/cell and HERmark in ASCO category groupsASCO category groupN (URMC)PID/cell (URMC)*N (PSHMC)HERmark (PSHMC)*18888.07761.521011.20N/A32016.0213.84238.5315.95296.3208.3*averageTable 2:Average PID/cell and HERmark in subgroups of Group 1SubgroupN (URMC)PID/cell (URMC)*N (PSHMC)HERmark (PSHMC)*A24157.66465.7B34101.61044.1C3016.9329.8*average
Citation Format: Buscaglia B, Turner B, Goda H, Huang W, Leitzel K, Natori T, Nakano Y, Okada H, Sperinde J, Ali S, Vasekar M, D'Aguiar M, McMahon L, Henry J, Lipton A, Hicks D. ASCO/CAP human epidermal growth factor receptor-2 (HER2) in situ hybridization (ISH) categories evaluated by quantitative HER2 protein diagnostic methodologies: A comparative analysis [abstract]. In: Proceedings of the 2018 San Antonio Breast Cancer Symposium; 2018 Dec 4-8; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2019;79(4 Suppl):Abstract nr P1-03-02.