Abstract PS2-10-02: Liquid Biopsy Tracking of PI3K-mTOR Residual Disease Signatures in Metastatic Breast Cancer

Abstract Background Breast cancer is the most commonly diagnosed cancer globally and remains the leading cause of cancer-related death in women. HER2-positive breast cancer, comprising 15-20% of cases, is defined by overexpression of the HER2 tyrosine kinase receptor. While HER2-targeted therapies have transformed clinical outcomes, resistance, recurrence, and metastasis continue to pose significant challenges. Immunotherapy has revolutionised the treatment of several solid tumors by enhancing anti-tumor immunity; however its efficacy in HER2-positive breast cancer has been limited. There is a pressing need for biomarkers that can both track metastatic burden and identify patients most likely to benefit from immunotherapeutic strategies. In this study, we present novel preclinical data from liquid biopsy analyses and propose a biomarker-driven approach targeting the PI3K-mTOR pathway to enhance immune reinvigoration in HER2-positive metastatic breast cancer. Methods Twenty five stage IV HER2-positive metastatic breast cancer (mBC) patients underwent liquid biopsy profiling using an epigenetic biomarker targeting the PI3K-mTOR signaling pathway. We applied digital pathology, Duolink® proximity ligation assays, global immune profiling, and flow cytometry to characterize biomarker expression and immune landscape features. Ex vivo drug screening with PI3K-mTOR inhibitors, including the brain-penetrant paxalisib, was performed to evaluate their effects on mesenchymal phenotypes, disruption of circulating tumor cell (CTC) clusters, and immune reinvigoration. Results We identified a novel epigenetic switch that drives the transition to a more aggressive, drug-resistant mesenchymal phenotype in HER2-positive breast cancer, associated with activation of the nuclear chromatinised PI3K signaling pathway. This phenotype, detectable via liquid biopsy, correlates with metastatic burden and immune exhaustion. In our pilot cohort, we evaluated the interplay between PI3K mutational burden and activation of this mesenchymal epigenetic program. Functional drug response profiling of patient-derived samples demonstrated that dual PI3K-mTOR inhibition not only facilitated mesenchymal-to-epithelial transition but also suppressed key metastatic signatures, including those linked to the highly aggressive persister cell phenotype (p65, FOXQ1, NRF2, and NNMT) and cancer drug resistance (ABCB5, SNAIL, and ALDH1). Importantly, PI3K-mTOR inhibition disrupted metastatic progenitors, including circulating tumor cell (CTC) clusters, and enhanced anti-tumor immune responses, as evidenced by modulation of both progenitor-like and terminally exhausted T cell populations. Conclusions Our preclinical findings support a biomarker-driven therapeutic strategy targeting the PI3K-mTOR pathway to induce epigenetic reprogramming and immune modulation. This dual mechanism may enhance anti-tumor efficacy by converting immunologically 'cold' tumors into 'hot', more immunogenic tumors. By reshaping the tumor microenvironment, this approach has the potential to improve the clinical utility of immunotherapy and expand treatment options for patients with HER2-positive metastatic breast cancer. Citation Format: M. Melino, W. Tu, S. Goh, A. Bain, J. Friend, T. Prasanna, D. Yip, A. Ives, M. Leahy, D. Lewis, M. Nottage, S. Rao. Liquid Biopsy Tracking of PI3K-mTOR Residual Disease Signatures in Metastatic 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 PS2-10-02.