Abstract PR009: Generation of T cell responses against broad KRAS hotspot neoantigens using a single large-scale process as the basis for an adoptive cell therapy or for TCR discovery
Authors: Conn Brandon P , Dietze Jared L , Yee Christian J , Hallisey Margaret M , Ortiz-Caraveo Irais , van Buuren Marit M , Gaynor Richard B , Foley Kendra C , Choi Jaewon , Juneja Vikram R 
Published: February 2025

Adoptive cell transfer (ACT) with T cells targeting KRAS neoantigens can drive anti-tumor immunity. However, due to the broad range of immunogenicity, the majority of ACT therapies have been focused on a small number of the over twenty different known KRAS neoantigens. Here, we develop a single process starting from peripheral blood that can prime and expand T cell responses to any KRAS neoantigen based on each individual’s HLA profile. We started with an ex vivo induction process, NEO-STIM, previously developed to induce T cell responses to a broad range of neoantigens. To account for the low fraction of antigen-specific T cells anticipated against individual KRAS neoantigens with low immunogenicity, we used a simple peptide pulse to upregulate 4-1BB (CD137) on cells specific for mutant KRAS and enriched using magnetic beads. We optimized both the initial stimulation and expansion steps such that nine of ten runs reached our target dose. Between the initial and optimized process runs, we conducted the process in 21 different healthy donors and generated T cell responses to 48 of 49 evaluated neoantigens with a range of antigen-specific frequency commensurate with the immunogenicity of the neoantigen. G12V neoantigens appeared more immunogenic on average than G12D neoantigens, with several HLA alleles (A*11:01, C*03:03, and C*03:04) presenting these epitopes in a particularly immunogenic fashion. Using tumor lines expressing the target KRAS mutation and HLA restriction elements, we demonstrated that the T cells generated through this process were able to slow tumor growth in vitro using a kinetic cytotoxicity assay and in vivo using a mouse xenograft model. We additionally cloned more than 150 KRAS T cell receptors (TCRs) in total, with multiple to each neoantigen. The strongest TCRs specific for KRAS G12D presented on HLA-A*11:01 and HLA-C*08:02 were similar or better than clinically active benchmark TCRs against these same targets, demonstrating that this approach can also be used for the discovery of clinically relevant TCRs from healthy donors. Together, this process can now be used as the basis for the development of a therapeutic or to discover a library of TCRs against each KRAS neoantigen regardless of immunogenicity.

Citation Format: Brandon P Conn, Jared L Dietze, Christian J Yee, Margaret M Hallisey, Irais Ortiz-Caraveo, Marit M van Buuren, Richard B Gaynor, Kendra C Foley, Jaewon Choi, Vikram R Juneja. Generation of T cell responses against broad KRAS hotspot neoantigens using a single large-scale process as the basis for an adoptive cell therapy or for TCR discovery [abstract]. In: Proceedings of the AACR IO Conference: Discovery and Innovation in Cancer Immunology: Revolutionizing Treatment through Immunotherapy; 2025 Feb 23-26; Los Angeles, CA. Philadelphia (PA): AACR; Cancer Immunol Res 2025;13(2 Suppl):Abstract nr PR009.