Optimizing the ER cutoff: Using ER≥50% to define a distinct HER2+ subgroup characterized by low pCR rates, ADC resistance, and CDK4 dependency.

e12646 Background: While estrogen receptor (ER)-positive/HER2-positive breast cancer generally demonstrates better long-term survival than ER-negative/HER2-positive disease, it shows significantly lower rates of pathological complete response (pCR) to standard neoadjuvant HER2-targeted therapies. Currently, the optimal ER positivity cutoff for stratifying patients remains undefined. Methods: We analyzed a retrospective cohort of 741 HER2-positive patients treated with neoadjuvant chemotherapy plus dual HER2 blockade (trastuzumab and pertuzumab). Receiver operating characteristic (ROC) analysis was used to define the optimal prognostic ER cutoff. Integrated multi-omics analysis (TCGA, SCAN-B) and CRISPR dependency screening (DepMap) were utilized to elucidate biological mechanisms and identify therapeutic vulnerabilities. Results: ROC analysis identified 50% ER positivity as the optimal threshold for stratifying response. In multivariate analysis, ER≥50% subgroup was characterized by activated estrogen signaling, downregulated cell cycle pathways, and predicted resistance to taxanes and antibody-drug conjugates (e.g., T-DXd). Conversely, CRISPR screening prioritized CDK4 as a top essential survival dependency specifically in ER-positive/HER2-positive models, suggesting functional addiction to the Cyclin D1-CDK4 axis. Conclusions: An ER cutoff of 50% optimally defines a distinct, chemo-resistant luminal subgroup within HER2-positive breast cancer. These findings support a precision medicine strategy integrating CDK4/6 inhibitors with endocrine and anti-HER2 therapies for patients with ER≥50% tumors.