Abstract 928: Advancing CAR-Ts and anti-immunosuppressive therapy development: Immunosuppressive TME 3D co-culture platform grounded on a fully characterized PDX tumor bank

Abstract Solid tumors represent a significant challenge in oncology due to their complex and highly immunosuppressive tumor microenvironment (TME). This hostile environment, shaped by immunosuppressive cytokines, and immune evasion mechanisms, limits the efficacy of immune-based therapies, including checkpoint inhibitors, chimeric antigen receptor (CAR) T cells, and bispecific antibodies. Although therapeutic strategies have been developed to overcome immune suppression, accurate preclinical models with predictive value remain critically needed. To address this, Champions Oncology has developed a robust ex-vivo 3D co-culture platform based on its extensive patient-derived xenograft (PDX) tumor bank, comprising over 1,500 fully characterized models. These models are annotated with comprehensive multi-omic data, including RNA sequencing, proteomic, and phospho-proteomic profiles, enabling the selection of tumor models aligned with specific indications, clinical history, and target expression, mimicking patient cohorts. The platform uses rigorously characterized organoids derived from this tumor bank as its core ensuring fidelity to the original tumor. Developing the ex-vivo immunosuppressive co-culture system requires careful integration of relevant cellular players, including human cancer-associated fibroblasts (CAFs), infiltrating immune cells such as M2-polarized macrophages, and tumor cells from 3D PDX organoids. Co-culture of these cell types creates a biologically accurate representation of the human TME. Here we describe a novel complex co-culture platform, with fully optimized conditions enabling the differentiation of cellular components and functional co-cultures. This system was used to test the potency of diverse CAR-T therapies engineered to overcome immune suppression. A custom high-throughput flow cytometry panel was developed to interrogate the system, measuring both cancer cell killing and immune activation. The co-culture system proved ideal for evaluating CAR-T cancer-killing capacity and understanding mechanisms of immunosuppression-mediated therapeutic resistance. Additionally, the platform supports testing combination therapies, such as checkpoint inhibitors paired with TME reprogramming agents, to combat resistance mechanisms. In conclusion, this ex-vivo co-culture platform represents a transformative tool in preclinical immuno-oncology. Combining a deeply characterized tumor bank with a robust organoid-based system enables testing anti-immunosuppressive therapies in a clinically relevant setting. This innovation promises to accelerate the development of next-generation therapies, offering unparalleled insights into tumor biology and immune dynamics to improve outcomes for patients with immunosuppressive tumors. Citation Format: Mara Gilardi, Veena Jagannathan, Brandon Walling, BanuPriya Sridharan, Stefano Cairo, Marianna Zipeto, Michael Ritchie. Advancing CAR-Ts and anti-immunosuppressive therapy development: Immunosuppressive TME 3D co-culture platform grounded on a fully characterized PDX tumor bank [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 928.