Assessment of Immune Responses to Neoantigen Vaccination in Lynch Syndrome

About This Grant

PROJECT SUMMARY Lynch Syndrome (LS) is the most common cause of hereditary colorectal (CRC) and endometrial cancer (EC), affecting >1 million Americans. LS is caused by germline mutations in one of the four DNA mismatch repair (MMR) genes. Normal epithelial cells in LS carriers become MMR deficient after a ‘second’ somatic hit, thus triggering the accumulation of hundreds of insertion-deletion mutations (indels) in microsatellite sequences (i.e., microsatellite instability, MSI). These mutations generate frameshift peptides (FSPs) that act as neoantigens (neoAgs), which are recognized by T cell receptors (TCRs) for activation of the immune system. We posit that LS carriers constitute a well-defined and prevalent population that can maximally benefit from immuneinterception strategies with neoAg vaccines to prevent LS-associated carcinogenesis. We have acquired genomic and immunogenicity data from our institutional LS cohort to catalog the most frequently recurrent and shared mutated neoAg across the LS precancer to early-stage tumors as well as associated TCR clonotypes. Expansion of neoAg-specific TCR clonotypes and structurally similar TCR clones eliciting immune responses have not yet been studied in healthy, at-risk populations for cancer interception. Thus, the central hypothesis of this proposal is that specific TCR clonotypes against shared mutated neoAg reside in the colorectal mucosa as memory T cells and can be expanded after neoAg vaccination to elicit adaptive anti-tumor immunity. We have already completed an investigator-initiated, NCI-sponsored, multi-center Phase I clinical trial of an adenoviral vaccine (Nous-209) for prevention of LS-associated cancers. We recently reported the findings on safety and immunogenicity of Nous-209 vaccine which includes a total of 209 shared mutated neoAg covering a from MMRdeficient CRC, EC, and gastric tumors. Here, we propose the following three specific aims (SA): 1. To profile the T cell repertoire in the colonic mucosa of LS trial participants post-Nous-209 vaccination, and to match LS neoplasm-associated neoAg to specific T cell clones using machine-learning models; 2. To assess the spatial organization of neoAg-specific T cell clones in the colonic mucosal microenvironment pre- and post-vaccination using digital spatial transcriptomics and RNAscope; and 3. To assess neoAg-T cell reactivity ex vivo in LS organoid co-cultures and in vivo using murine tumor xenograft models. We are proposing to orthotopically implant a MMR-deficient CRC cell line model in animals to recreate the human tumor microenvironment, thus allowing us to examine the infiltration of immune cells into tumor, to assess the T cell response to tumor burden, and to characterize the mechanism of T cell-mediated activity in the tumor setting, which cannot be directly done in humans outside of a clinical trial. This proposal is highly innovative by combining single-cell genomics, spatial transcriptomics, and systems biology tools to assess the immune response to neoAg vaccination. This proposal will impact the field of hereditary gastrointestinal syndromes by developing a novel cancer prevention strategy for a population at high risk for different types of cancers, which may have implications for other cancer syndromes.