Abstract B040: High-Content Screening to Enhance the Preclinical Development of Immunotherapies, Validated for 100 Patient-Derived Organoids

Abstract Introduction Significant drug developments in immuno-oncology (IO) have been made in the last decade, which have led to clinical success in haematological cancers. To further unravel the potential of immunotherapies in solid cancers, relevant translational in vitro models are needed. Unlike traditional 2D assays with cancer cell lines, with the advancements in organoid technology immune responses can be monitored in a complex 3D co-culture of immune cells with tumor material, reflecting the original patient tumor morphology, genetics and transcriptomics. Using our biobank of over 650 patient-derived organoids (PDOs) from 21 different tissue types, a high content imaging (HCI)-based screening platform has been developed. This platform is highly effective for evaluating immunotherapy responses in vitro replicating the complex cellular interactions of the tumor microenvironment. Here, we assessed the immune sensitivity of 100 organoid models from 10 different cancer indications and tested the efficacy of T cell engagers and CAR-T cells. Methods and Materials PDOs and PDX-derived organoids (PDXO) from different indications (breast, cervical, colorectal, gallbladder, head and neck, lung, melanoma, mesothelioma, ovarian, and pancreatic) were cultured in a protein hydrogel. Isolated PBMCs from allogenic healthy donors were added to the 3D cultures to determine the immune-sensitivity of the models upon stimulation with superantigen or CD3/28 beads. Selected models were treated with EGFR targeting CAR-T cells or T cell engagers. HCI analysis was performed to extract morphometric information on immune cell infiltration and immune cell-mediated killing of the organoids. Additional cytokine analysis on the supernatants was performed by multiplex MSD. Results The clinical variability of sensitivity or resistance of cancer patients to immune cell-mediated killing was reflected in the 100 organoid models analyzed. Although most tested models demonstrated an increase in IFNγ levels upon treatment with EGFR targeting T cell engagers, these findings did not always correlate with increased killing of the organoids measured via HCI. These results correspond to the data obtained with EGFR targeting CAR-T cells. These CAR-T cells demonstrated an increased infiltration into the organoids, which resulted in reduced organoid volumes. Conclusion The preclinical in vitro model of organoid and immune cell co-cultures presented here allows for the functional testing of immunotherapies in a physiologically relevant assay via HCI. This platform enables the combination of classical biomarker analysis with read-outs on immune cell infiltration into organoids and subsequent organoid killing. This approach provides a deeper understanding of the mechanisms of action of the tested immunotherapies. With a large array of validated organoid models from various cancer indications, this innovative platform enhances preclinical testing of cancer immunotherapies, ultimately bringing the promise of these therapies to more patients. Citation Format: Saskia de Man, Sergei Chavez-Abiega, Tomas Veenendaal, Michelle Kop, Jarmil Hanrath, Daniel Okkes, Emma Spanjaard, Leo Price, Gera Goverse. High-Content Screening to Enhance the Preclinical Development of Immunotherapies, Validated for 100 Patient-Derived Organoids [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor Immunology and Immunotherapy; 2024 Oct 18-21; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2024;12(10 Suppl):Abstract nr B040.