Abstract P4-10-21: Utility of HER2 IHC intensity in selecting areas for FISH testing in equivocal (2+) cases

Intro: The 2018 ASCO/CAP HER2 guidelines eliminated the equivocal category for in situ hybridization (ISH). Additional workup is recommended for definitive classification of tumors that are sorted into ISH groups 2, 3 or 4. Evaluation of HER2 IHC using sections from the same tissue sample used for ISH is recommended to guide the selection of areas to score by ISH. Here we seek to evaluate the utility of selecting the area for ISH testing in IHC equivocal (2+) cases based on the staining intensity of the IHC staining. Methods: A total of 66 cases with equivocal HER2 IHC (2+) and reflexed to FISH were prospectively collected from 8/22/18 to 6/26/2019. The areas of most and least intense HER2 IHC staining were separately identified and scored by FISH. The mean HER2:CEP17 ratio and mean HER2 signals/tumor cell were calculated for both populations and statistical significance was evaluated using the Wilcoxon Ranked Sum test. Results: The distribution of the high intensity area was 58 ISH group 5, 4 ISH group 4, and 8 ISH group 1. The distribution of the low intensity area was 51 ISH group 5, 9 ISH group 4, 2 ISH group 2, and 5 ISH group 5. In the high and low intensity areas, 59 of 66 (95.5%) cases sorted into the same ISH group and 7 (10.6%) sorted into different ISH groups. Of these 7, 3 (4.5%) had different final FISH categorizations. All 3 cases were ISH Group 1 (amplified) in the high intensity area and ISH group 2, 4 or 5 (not amplified in the absence of 3+ IHC) in the low intensity area. The other 4 cases with different ISH groups did not have different final HER2 categorizations. The mean HER2/CEP17 ratios for the high and low intensity areas were 1.50 and 1.39 (p = 0.075). There was no statistically significant difference in mean HER2 copy number for the high and low intensity areas (p = 0.92). Discussion: When areas with high and low HER2 IHC intensity were compared, there was a small difference in the mean HER2:CEP17 ratio with a trend toward statistical significance. However, the absolute difference in the means was small (0.11) and only a minority of cases (4.5%) showed discordant final HER2 categorization between the two groups. All of the cases with discordant HER2 categorization were amplified (ISH Group 1) in the IHC high intensity area and not amplified (ISH Groups 2, 4 and 5) in the low intensity area. The data suggests that using HER2 IHC to guide selection of areas for ISH testing may identify amplified tumors (or clones) in a minority of cases (4.5%). Additional analysis of a larger number of cases is required to validate these findings. Citation Format: Johann D Hertel, Kathleen A Kaiser-Rogers, Siobhan O9Connor, Benjamin Calhoun. Utility of HER2 IHC intensity in selecting areas for FISH testing in equivocal (2+) cases [abstract]. In: Proceedings of the 2019 San Antonio Breast Cancer Symposium; 2019 Dec 10-14; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2020;80(4 Suppl):Abstract nr P4-10-21.