Abstract 2697: Targeting tumor microenvironments of triple negative breast cancer with a novel adeno-associated virus-mediated virotherapy

Abstract Triple-negative breast cancer (TNBC) is a group of aggressive breast cancers with a higher mortality rate due to their resistance to chemotherapy and radiotherapy. Unfortunately, there are no specific targeted therapies for TNBC, making it a critical medical need. In basal-like TNBC, Notch signaling regulates the recruitment of tumor-associated macrophages (TAMs) and the activation of cancer stem-like cells (CSCs) by TAMs through TGF-β pathways. TGFβ-responsive CSCs have enhanced resistance to T cell immunotherapy and are responsible for tumor relapse. Therefore, TGF-β pathway inhibition is a promising therapeutic target in TNBC. However, due to systemic toxicity, the FDA has not approved TGF-β investigational inhibitors. To avoid these toxicities, we use a recombinant adeno-associated virus (r-AAV) vector encoding a soluble TGF-β receptor decoy (r-AAV-TGFβRD) to inhibit TGF-β intratumorally in TNBC. r-AAVs are non-replicating viruses and, therefore, are a safe platform for in vivo gene therapy delivery. We found that rAAV-GFP transduced both human and mouse TNBC cells. Our rAAV constructs contain a C1QTNF3-derived collagen domain (CD) that facilitates the trimerization and stabilization of secreted fusion proteins. We purified these proteins from cellular supernatants of r-AAV vector-transduced cells using His-Tag affinity chromatography and found that they significantly decreased human TNBC cell's mammosphere growth. Intratumoral injection of the r-AAV-TGFβRD virus significantly reduced syngeneic mouse TNBC tumor growth and increased the tumor-infiltrating lymphocytes (TILs), including CD3 and CD8. NanoString analyses (nCounter Immunology) suggest changes in tumor microenvironments after the treatment. We found significant increases in the innate immune system, host-pathogen interactions, and chemokine signaling with increasing influx of macrophages, DC, and CD8 as TILs. Interestingly, we found decreased B cells and Tregs within TME as TILs. Notably, the r-AAV-TGFβRD virus significantly enhances the efficacy of chemotherapy (paclitaxel) in mediating the anti-tumor tumor effect in the syngeneic TNBC mouse model. These findings indicate that r-AAV-TGFβRD virus can be a promising approach for treating TNBC alone or in combination with chemotherapy for translational application in the future. Citation Format: Md. Manirujjaman, Yong Ran, Soroor Heidari, Jone Garai, Samarpan Majumder, Luis Del Valle, Jovanny Zabaleta, Barbara Osborne, Lucio Miele, Todd Golde, Fokhrul Hossain. Targeting tumor microenvironments of triple negative breast cancer with a novel adeno-associated virus-mediated virotherapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2697.