Analytical and clinical validation of PATHWAY HER2 (4B5) assay for assessment of HER2-low/HER2-ultralow status and eligibility for trastuzumab deruxtecan in DESTINY-Breast06

Background: The DESTINY-Breast06 study demonstrated a statistically significant and clinically meaningful improvement in progression-free survival benefit with trastuzumab deruxtecan (T-DXd) versus chemotherapy treatment of physician's choice in patients with hormone receptor-positive metastatic breast cancer (mBC) whose tumors were scored as human epidermal growth factor receptor 2 (HER2)-low [immunohistochemistry (IHC) 1+, or IHC 2+/in situ hybridization (ISH)-negative] and who had received one or more lines of endocrine therapy and no previous chemotherapy in the metastatic setting. An exploratory analysis consistently showed a benefit for patients with HER2-low and HER2-ultralow (IHC 0 with membrane staining) expression. Materials and methods: Analytical validation of the PATHWAY HER2 (4B5) assay (Roche HER2 4B5 assay; Roche Diagnostic Solutions) at the HER2-ultralow cut-off was carried out, including intermediate precision, inter-reader precision, and inter-laboratory reproducibility. Patients with tumors historically classified as HER2-negative (IHC 0, 1+, and 2+/ISH-negative) based on pre-existing local test results were eligible for screening for DESTINY-Breast06; tumor samples taken in the metastatic setting were assessed centrally to confirm eligibility regarding HER2 status. Additionally, central and pre-existing HER2 test results were compared, and the prevalence of IHC scores based on central test results was assessed. Results: Intermediate precision met the pre-specified endpoints across all parameters tested; agreement was observed within and between readers and sites, demonstrating that the HER2-ultralow cut-off can be scored accurately and reproducibly using the Roche HER2 4B5 assay (intra- and inter-laboratory and inter-reader agreement ≥95%). A clinically meaningful progression-free survival (hazard ratios 0.43-0.78) and objective response rate (odds ratios 2.5-4.5) improvement for T-DXd over chemotherapy was consistently observed across IHC expression levels. For cases with a pre-existing HER2 IHC 0 result, central testing found 24% with HER2-low, 40% with IHC 0 with membrane staining, and 35% with IHC 0 absent membrane staining. Conclusions: For patients with HER2 IHC 0 mBC, rescoring and/or retesting with the Roche HER2 4B5 assay is encouraged to determine eligibility for T-DXd.