Abstract PS2-11-08: Stemness-targeted therapies to enhance the efficacy of AURKA inhibitors in endocrine resistant er+ breast cancer

Abstract Significance: Emergence of resistance to endocrine therapies represents a major challenge in theclinical management of estrogen receptor positive (ER+) breast tumors. CDK4/6 inhibitors(palbociclib, ribociclib, abemaciclib) have improved survival when combined with endocrine therapy. Nonetheless, resistance to CDK4/6 inhibitors eventually occurs. One of the emerging mechanisms ofresistance toward endocrine therapy and CDK4/6 inhibitors is characterized by the enrichment ofbreast cancer stem cells (CSCs) that show stem-like features such as self-renewal capacity, highCD44 expression and ALDH activity. We have demonstrated that Aurora-A kinase (AURKA) has apivotal role in promoting cancer cellular plasticity and clonal expansion of endocrine resistantCD44high/ALDHhigh CSCs. Our preclinical studies on AURKA-driven tumor stemness provided thetherapeutic foundation for a Phase-II clinical trial (TBCRC041) of the AURKA inhibitor, alisertib, whichdemonstrated clinically meaningful activity in patients with endocrine and CDK4/6 inhibitors resistantER+ metastatic breast cancer (MBC). Despite such activity, alisertib resistance finally occurred. Therefore, we hypothesize that up-regulation of IL6R/STAT3 and PR stemness oncogenic pathwaysmay sustain the clonal expansion of CD44high/ALDHhigh CSCs after tumor progression on AURKAinhibitors. Experimental Design: We used variant MCF-7 breast cancer models that show resistance toendocrine therapy (letrozole and fulvestrant) and alisertib (MCF-7 LR/FR, MCF-7 LR/FR-AlisR) todefine in vitro and in vivo the efficacy of stemness-targeted therapies to enhance alisertib activity byimpairing the enrichment of hormone refractory CD44High/ALDHHigh CSCs. Parental MCF-7 cells wereused as control. We employed exclusive PDX-derived 3D-Mammospheres (MPS) that wereestablished from the Phase-II clinical trial TBCRC041 to test the efficacy of stemness-targetedtherapies in enhancing the therapeutic activity of alisertib. In Vitro Studies: Immunofluorescence andimmunoblot assays were used to assess the expression of breast cancer stemness biomarkers. Enrichment of ALDHHigh CSCs was measured using Red-Aldefluor kit. Cell proliferation and Red-Annexin-VHigh apoptotic cells were quantified in real-time using IncuCyte S3. In Vivo Studies: 1x106breast cancer cells were injected into the mammary fat pad of NSG-female mice and treated withfulvestrant, alisertib and tocilizumab as monotherapy or in combination. Control groups were treatedwith saline solution placebo. Drug Treatments: Tocilizumab and Onapristone were used to inhibit theIL6R/STAT3 and PR stemness pathways, respectively. Results: Our study demonstrates for the first time that nuclear STAT3/PR complex mediatesAURKA-induced expression of KLF4 stemness gene and enrichment of endocrine resistantCD44high/ALDHhigh CSCs. Alisertib resistant MCF-7 LR/FR cells showed increased expression ofIL6R, T288-AURKA, S727-STAT3 and S294-PR compared to alisertib naïve cells. MCF-7 LR/FR-AlisR-derived 3D-MPS had increased KLF4 and CD44 expression. Alisertib resistant PDX modelsalso showed high levels of CD44, IL6R/STAT3 and KLF4 expression. Remarkable, pharmacologicalblockade of PR and STAT3 transcriptional regulatory networks enhanced the efficacy of alisertib byreducing the enrichment of CD44High/ALDHHigh CSCs. Conclusions: These findings provide the strong preclinical rationale for the development of novelcombinatorial therapeutic strategies targeting IL6R/STAT3 and PR stemness pathways to enhancethe efficacy of AURKA inhibitors that will result in the selective eradication of CD44High/ALDHHighCSCs with consequent benefits on the overall survival of patients with advanced ER+ breast tumorsrefractory to endocrine therapies and CDK4/6 inhibitors.輀 Citation Format: A. B. D'ASSORO, M. Drago, T. Haddad, M. GOETZ, K. Giridhar, K. Kalari, L. Manzella, P. Vigneri, F. Drago, C. Vancheri, C. Lange, J. Ingle. Stemness-targeted therapies to enhance the efficacy of AURKA inhibitors in endocrine resistant er+ 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 PS2-11-08.