Abstract 1878: 3D Tumor models in drug discovery for high throughput analysis of immune cell infiltration
Authors: Halder Tithi Ghosh , Rheinschmidt Shelby , Thode Trason , Sampson Samuel , Rodriguez del Villar Ryan , Ng Serina , Weston Alexis , Kaadige Mohan , Trent Jeffery , Altin John , Moser Justin , Borazanci Erkut , Gordon Michael , Larson Anna , Soldi Raffaella , Sharma Sunil 
Published: June 2022

Abstract
The interaction between tumor cells and immune system has been studied mostly in animal models and co-cultures with cell lines. However, it is difficult to study the unique features of human immune cells in mice, and cell lines are often transformed or immortalized by genetic modification. Organoid platform has emerged as a powerful tool which maintains the architectures and distinctive functions of a specific organ. Although organoids still fail to model immune system accurately, co-cultures of organoids and lymphocytes may have promising applications. In this study, we developed a unique methodology to use organoids for understanding the role of epithelial cell-immune cell interactions in cancer. Using this immune-organoid tool we tested the infiltration and efficacy of TILs generated from patient tumor cells. We also tested if checkpoint blockade could amplify the immune cell infiltration in tumor using this organoid platform bringing hope for further application of this technology in immunotherapy. Single cell suspensions were prepared from patient tumor biopsy and seeded in ultra-low attachment (ULA) microplates in appropriate media to generate organoids. TILs were isolated from the same tumor and rapidly expanded in appropriate media. Tumor organoids and TILs were incubated with PD-L1 and CTLA-4 monoclonal antibody respectively for check point blockade. RFP-stained TILs were then seeded into inserts of a transwell receiver plate using specific tumoroid:TIL ratio. After 48 h organoids were analyzed by 3D Z-stack imaging and morphometric analysis with Cytation 5 software to quantify the lymphocyte infiltration. Using this immune organoid model, we were able to quantify the number of TILs that infiltrates inside the tumor organoids generated from a melanoma and a pancreatic cancer patient. We also found that PDL-1 checkpoint Blockade in tumor cells enhanced the TIL infiltration inside organoids. Moreover, we were able to isolate the infiltrated lymphocytes from the tumor organoids and characterized them by analyzing different markers and cytokines using FACS, RT-PCR and Immunohistochemistry. Therefore, this study has potential to be an excellent preclinical model for human tumors, translating basic cancer research to clinical practice as well as providing a new meaning to precision medicine by identifying and validating novel immune therapies for individual cancer patients.
Citation Format: Tithi Ghosh Halder, Shelby Rheinschmidt, Trason Thode, Samuel Sampson, Ryan Rodriguez del Villar, Serina Ng, Alexis Weston, Mohan Kaadige, Jeffery Trent, John Altin, Justin Moser, Erkut Borazanci, Michael Gordon, Anna Larson, Raffaella Soldi, Sunil Sharma. 3D Tumor models in drug discovery for high throughput analysis of immune cell infiltration [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 1878.