Abstract P113: Image-based quantification of immunotherapeutic effect on the tumor-immune interactions in 3D co-cultures

Abstract

Abstract INTRODUCTION Despite the increasing number of immunotherapies available in preclinical cancer research and drug discovery pipelines, the number of patients benefiting from these immunotherapies remains extremely low. Moreover, the failure of immunotherapies in clinical trials remains very high to date. These high failure rates could be attributed to the lack of deep understanding the mechanism of action of drug candidates and to the absence of clinically relevant preclinical models that are suitable for drug screening purposes. To address these issues, we have developed an in vitro drug screening platform suitable for high-throughput testing in physiologically relevant 3D environment. Our preclinical drug testing platform is based on the co-culture of tumor cells with immune cells in 3D. Image analysis is applied to read out effects of drug candidates on immune cell migration, immune cell infiltration and tumoroid killing. MATERIALS AND METHODS 3D tumor cultures from human cancer cell lines (including breast, lung and colon cancer) or patient-derived colorectal organoids (from HUB Organoid Technology) were generated by embedding tumor cells in 3D ECM-like matrix constituted from protein hydrogel. Subsequently, different immune subsets, including PBMCs from healthy donors, T cells, NK cells or macrophages, were stained with cell tracker and added to the cultures. The cultures were treated with different immune-modulators (e.g. superantigens, activating antibodies, T cell engagers, CSFR1 inhibitor or STING agonist). The effect of immune-modulators on immune cell infiltration and their killing was assessed by high-content imaging and quantified after morphometric analysis with the proprietary Ominer® software. Immune cell-mediated killing was confirmed by measurement of IFNγ secretion in the culture supernatants. RESULTS Image-based analysis allowed for the dissection of complex tumor-immune cell interactions in the 3D cultures. Moreover, morphometric analysis revealed different levels of immune cell infiltration and tumoroid killing upon treatment with different immune-modulators. These effects could be reduced by inhibitory signals from the tumor microenvironment (TME) and were confirmed by the levels of IFNγ secreted by the immune cells. CONCLUSION Our in vitro platform allows quantitative image-based analysis of 3D tumor-immune cell co-cultures in a high-throughput manner, based on spatially resolved information in a more physiologically-relevant setting compared to traditional 2D cultures. This image-based analysis could be employed to dissect the effect of immunotherapy on different cell populations with the TME representing a promising tool to improve our understanding of the mechanism of action of novel treatments, treatment relapse and combination strategies, to eventually lead to a better clinical performance. Citation Format: Lidia Daszkiewicz, Gera Goverse, Nataliia Beztsinna, Marjan van de Merbel, Benjamin Visser, Tomas Veenendaal, Emma Spanjaard, Kuan Yan, Leo Price. Image-based quantification of immunotherapeutic effect on the tumor-immune interactions in 3D co-cultures [abstract]. In: Proceedings of the AACR-NCI-EORTC Virtual International Conference on Molecular Targets and Cancer Therapeutics; 2021 Oct 7-10. Philadelphia (PA): AACR; Mol Cancer Ther 2021;20(12 Suppl):Abstract nr P113.