Background: Cancer progression and response to therapy are governed by complex cellular interactions in the tumor microenvironment (TME). A better understanding of TME and the immune microenvironment requires robust experimental systems modeling patient specific immune interactions. Conventional 2D cultures lack in recapitulating 3D morphological structures and complex immune biology of native human tumors. A vastly heterogenous TME impacts patient treatment response. There is a dearth of 2D or 3D in vitro models that mimic in vivo diversity of TME, including infiltrating immune populations. 3D organoid cultures typically contain neoplastic epithelium and fail to retain syngeneic tumor-infiltrating lymphocytes (TILs) interactions vital for exhibiting a clinically significant drug response. Here we describe patient-derived xenograft (PDX) derived 3D organoid (PDXO) cultures with autologous TILs to avoid alloreactivity and simulate tumor specific immune response. The described model allows the evaluation of TIL infiltration and high throughput screening of immune-modulatory therapies.
Methods: Patient-derived xenografts from mice were enzymatically digested to establish PDXO. PDXO were characterized for expression of cancer specific cell-surface markers. TILs were expanded in culture from PBMCs from matching patients and characterized using flow cytometry. Organoids and TILs from the same patient were fluorescently labeled and cocultured for four days, and tumor infiltration and drug cytotoxicity with and without immune-therapeutics was evaluated. CellInsight CX7 high content imaging platform was used to track TILs, and cancer cells, and TIL infiltration was confirmed via immunofluorescence. Luminex technology was used to quantify changes in the cytokine profile upon drug treatment.
Results: PDXO were established to mimic in vivo tumor biology. Tumor-specific TILs were expanded and characterized for expression of memory, inhibitory, proliferation, and regulatory T cell markers. TIL infiltration in organoids was observed from day one in culture and maintained over four days. Immunofluorescence for T cell marker CD3 (cluster of differentiation 3) within PDXO confirmed TIL infiltration. Infiltrating TILs and tumor cell killing evaluation using fluorescent dyes via high throughput imaging platform showed disease and patient-specific response. PDXO treatment with immune-modulatory therapeutics led to changes in the secreted levels of cytokines IL2 (interleukin 2), IFNγ (interferon gamma), and TNFα (tumor necrotic factor alpha).
Conclusion: Our patient autologous TILs - PDXO culture platform can be used for high throughput drug screening to model immune-therapeutic response with the tumor specific immune microenvironment.
Citation Format: Garima Kaushik, Padma Kamineny, Amy Wesa. Unique 3D organoids autologous TIL coculture platform enables high throughput immune-oncology drug response studies [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 2978.