Background The preclinical screening of immune-modulatory therapies suffers from the absence of models that recapitulate in vivo heterogeneous tumor microenvironment (TME). 3D tumor organoid cultures provide a model that closely mimics in situ tumor architecture and is being aggressively used to evaluate therapeutic efficacy ex vivo. A vastly heterogenous TME impacts patient treatment response, and there is a dearth of human tumor models (2D or 3D), that mimic in vivo diversity of TME, including infiltrating immune populations. 3D organoid cultures typically contain neoplastic epithelium; however, they fall short in representing tumor to tumor-infiltrating lymphocytes (TILs) interactions, limiting their ability to generate a clinically relevant response to immunotherapeutics. Addition of immune cells from unrelated donors to organoids can simulate that microenvironment but is complicated by T cell alloreactivity. Here we describe 3D patient-derived xenograft organoid (PDXO) co-cultures with matching autologous human TILs to recapitulate the tumor-specific immune response, leveraging confocal high content analysis and luminex multiplex assays. This platform allows the evaluation and high throughput screening of novel immune targeting agents to determine impacts on patient-derived T cell function, T cell infiltration, and tumor cytotoxicity.
Methods Surgical resections from patients were used to generate patient-derived xenografts and tumor-infiltrating lymphocytes in parallel. PDX were resected and digested to establish PDXO. TILs and organoids from the same patient were fluorescent labeled and cultured together for four days to evaluate tumor infiltration and drug cytotoxicity in 3D cultures. CellInsight CX7 high content imaging platform was used to trace TILs and cancer cells and evaluate T cell infiltration and tumor cell killing in the presence and absence of immuno-modulatory therapies.
Results PDXO were established to mimic in vivo tumor biology. Tumor-specific TILs were successfully expanded and characterized by flow cytometry. Co-culture resulted in TIL infiltration in organoids from day one in culture and increased over four days. Cytotoxicity and TIL infiltration were quantified using fluorescent dyes via high throughput imaging platform. Significantly enhanced TIL infiltration was observed in autologous co-cultures compared to non-autologous co-cultures. The established unique autologous PDXO immune organoid co-cultures could be used as an improved simulation of the modulatory activity of therapeutic agents in patient-specific T cells against their own tumors.
Conclusions Patient autologous TILs – PDXO co-culture platform is an advanced model for evaluating IO therapeutics with the tumor-specific immune microenvironment. The platform provides an opportunity for precision medicine and high throughput drug screening of immuno-modulatory therapies.
Ethics Approval The study was approved by Champions Oncology’s Institutional Animal Care and Use Committee (IACUC).
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