Abstract 1772: A novel approach for autologous pan-cancer cellular immunotherapy reveals dramatic expansion of ab and gd TCR T cell clonotypes indicative of an antigen-driven response

Abstract

Abstract Introduction: We developed a unique autologous cellular therapeutic strategy using a proprietary engineered leukocyte stimulator cell lines (ENLIST cells) that express an array of immunomodulatory proteins to activate and expand tumor cell killing effector cells from human peripheral blood mononuclear cells (PBMCs). Our clinical aim is to use these expanded cells, called SUPLEXA therapy in cancer patients. Results of prior immunophenotyping of SUPLEXA cells by mass cytometry (CyTOF) reveals that they are comprised of NK cells, CD8+ T cells and γδ T cells that express markers indicative of cytolytic function, e.g. granzymes and perforin. Using an unbiased approach to better define the specificity of SUPLEXA, we performed TCR repertoire analysis using the iRepertoire and 10X Genomics RNA sequencing platforms. Results: SUPLEXA was prepared from healthy donor PBMCs. RNA was subsequently prepared from SUPLEXA cells and donor PBMCs to compare TCR αβ and γδ chain clonality and diversity using the iRepertoire DAM-PCR sequencing platform. SUPLEXA cells showed dramatic and significantly increased clonality among the 4 TCR chains that were analyzed as compared to normal PBMCs. We also observed a diversification of TCRαβ and TCRγδ sequences in SUPLEXA as compared to PBMCs. In addition, to the increased diversity, clonal expansion was also observed. Single-cell RNA sequencing of flow-sorted CD3+ T cells using the 10X Genomics platform with paired TCRαβ and TCRγδ PCR sequencing feature overlays revealed high frequencies of T cells with paired TCR clonotypes. We discovered that the top 5 paired TCRαβ clonotype sequences accounted for 32.7% of T cells, while CD8+ T cells from a normal donor contains only 3.0% in the top 5 clonotypes. Similar high frequency clonotypes were observed for γδ T cells by 10X single-cell RNA sequencing. Conclusions: The creation of SUPLEXA from healthy adult PBMCs results in dramatic expansion of αβ and γδ TCR T cell clonotypes. Given that SUPLEXA cells demonstrate broad, dose-dependent tumor cytolytic activity, these findings suggest that the expanded αβ and γδ T cell clonotypes may react to tumor neoantigens that are present during the activation process by ENLIST cells, which are derived from the SK-MEL-2 melanoma cell line. Accordingly, TCR reagents made using the paired αβ and γδ T cell sequences from the expanded clonotypes will be useful for future tumor neoantigen discovery. Plans for a phase 1 clinical trial are currently underway. Citation Format: Frank Borriello, Joshua W. Keegan, Brandon L. Hancock, Fei Guo, James R. Lederer. A novel approach for autologous pan-cancer cellular immunotherapy reveals dramatic expansion of ab and gd TCR T cell clonotypes indicative of an antigen-driven response [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 1772.