Abstract PS3-11-20: Tumor-intrinsic signatures associated with response to chemotherapy-free HER2/PD-L1 blockade in HER2-enriched early breast cancer

Abstract Background: HER2-positive breast cancers have demonstrated sensitivity to de-escalated regimens combining HER2-targeted therapy with immune checkpoint inhibition. In our recent Phase II trial evaluating a chemotherapy-free regimen of durvalumab, trastuzumab, and pertuzumab (DTP), we observed a 49% pathologic complete response (pCR) rate among patients with HER2-Enriched tumors (BluePrint ®). Understanding tumor-intrinsic and microenvironment factors driving response variability remains a critical goal. Methods: To investigate mechanisms of therapeutic response and resistance, we performed single nucleus RNA sequencing (snRNA-seq) on formalin-fixed, paraffin-embedded tumor specimens from all 37 evaluable DTP trial participants. Paired pre-treatment needle biopsies and post-treatment surgical resections were profiled to capture longitudinal transcription dynamics and changes in cellular composition. Patients were stratified by Residual Cancer Burden (RCB) following surgical pathology. While previous analyses focused on immune cell dynamics, this study presents preliminary findings from 201,788 total epithelial cells. Epithelial populations were initially classified as aneuploid (“tumor”) or diploid (“normal”) epithelial cells based on inferred copy number alterations, with tumor cell calling validated by concordance with RCB status. Results: Tumor cells from Responder patients (RCB0, n = 18), RCB1 patients (n = 7), and Non-Responder patients (RCB2/3, n = 6) exhibited Chromosome 17 amplification consistent with HER2 positive status. snRNA-seq of tumor samples collected at surgery revealed a stepwise increase in tumor content—absent in RCB0, low in RCB1, and high in RCB2/3—supporting the accuracy of epithelial cell classification. Notably, Non-Responders harbored multiple tumor subclones, whereas Responders retained a single dominant clone from pre- to post-DTP treatment. Gene Set Enrichment Analysis of pre-treatment tumor cells from Responders versus Non-Responders revealed enrichment of endoplasmic reticulum (ER) stress and unfolded protein response pathways, increased ER-Golgi trafficking, and altered translational programs in tumor cells in Responders, suggesting that DTP-sensitive tumors are undergoing apoptosis or immunogenic cell death. In contrast, tumor cells from Non-Responders were enriched for Wnt/β-catenin and PI3K/Akt/mTOR signaling, indicating activation of tumor-intrinsic resistance pathways. Conclusions: Single-cell profiling revealed epithelial gene expression programs with potential to predict response or resistance to dual HER2 blockade and immune checkpoint inhibition. Ongoing spatial transcriptomic studies aim to map immune-epithelial interactions and tissue architecture in HER2-enriched breast cancer. Clinical trial information: NCT03820141. Research Sponsor: Houston Methodist Hospital and Astrazeneca. Citation Format: M. Li, J. Deng, X. Hoi, J. Zheng, R. Hashmani, W. Qian, J. Zhou, J. Guan, K. Sun, H. Mai, T. Sheu, S. Haley, M. Schwartz, S. Wong, F. Nikolo, K. Chan, P. Niravath, J. Chang. Tumor-intrinsic signatures associated with response to chemotherapy-free HER2/PD-L1 blockade in HER2-enriched early breast cancer abstract. In: Proceedings of the San Antonio Breast Cancer Symposium 2025; 2025 Dec 9-12; San Antonio, TX. Philadelphia (PA): AACR; Clin Cancer Res 2026;32(4 Suppl):Abstract nr PS3-11-20.