Interassay Comparison With Digital Image Analysis of Four Routine HER2 Immunohistochemistry Assays in Primary Breast Cancer and Its Metastasis

Trastuzumab-deruxtecan (T-DXd), an antibody-drug conjugate targeting human epidermal growth factor receptor 2 (HER2), improves overall survival in patients with breast cancer showing low or ultra-low HER2 expression. Differences in HER2 test results have been reported between various HER2 assays and between primary tumors and metastases, but an objective comparison with incorporation of the new HER2-ultralow cut-off values is needed. This study aimed to assess the performance of four routine clinical-grade HER2 assays across and between primary tumors and their metastases using digital image analysis (DIA). Primary tumors and metastases from 193 patients with breast cancer who participated in the IMPACT-MBC trial were incorporated into six tissue microarrays. Samples were stained by four laboratories using their routine HER2 immunohistochemistry protocols: 4B5 ultraView, 4B5 OptiView, SP3, and HercepTest (DG44). HER2 scores were determined using DIA. The four HER2 assays showed significant differences in HER2 status in both primary tumors and metastases. Eighty-five matched primary tumors and metastases were analyzed to investigate concordance in HER2 status. While no significant differences were found in HER2 scores between primary tumors and metastases for SP3 and both 4B5 assays, DG44 showed significantly higher HER2 scores in the metastasis (p = 0.004). Concordance between primary tumors and metastases was highest for 4B5 ultraView (69.4%), followed by SP3 (61.2%) and 4B5 OptiView (51.8%). DG44 showed the most variability, with only 36.5% of matched samples receiving the same HER2 category. DG44 identified a significantly higher proportion of HER2-(ultra)low cases and showed the most variability in HER2 status between matched primary tumors and metastases compared to 4B5 and SP3. The choice of HER2 assay can lead to discrepancies in HER2 status assessment, which could directly influence patient eligibility for T-DXd treatment.