Interrogating the relationship between translational dynamics and non-canonical antigen presentation in lung cancer - ABSTRACT Antigen presentation is a fundamental component of cancer immunity. The primary cytotoxic effectors of cancer immunity are CD8+ T cells, which recognize peptide antigens presented in the context of Major Histocompatibility Complex Class I (MHC-I). These foundational interactions between cancer cells and immune cells underlie the success of immunotherapy. Antigens derived from somatic mutations, called “neoantigens”, have garnered significant interest for the design of next generation immunotherapies. However, neoantigens suffer from significant clonal heterogeneity within tumors and are specific to individual patients, limiting their widespread use in engineered immunotherapies like cancer vaccines. To address this limitation, advances in mass spectrometry have enabled direct measurement of the full repertoire of peptides known to bind MHC-I which is collectively known as the “immunopeptidome”. Ever increasing depth and resolution in measuring the immunopeptidome has revealed numerous sources of cancer specific antigens beyond neoantigens, including peptides derived from cryptic translation events from novel unannotated open reading frames (nuORFs). Notably, immunotherapy trials in cancer have revealed that antigen specific CD8+ T cell responses extend far beyond just mutated neoantigens, especially in low tumor mutation burden patients, highlighting a critical need to understand additional classes of tumor specific antigens like nuORFs. A deeper understanding of the mechanisms by which nuORFs are presented may reveal novel pathways that could be pharmacologically targeted to increase cancer immunogenicity. We hypothesize that nuORF antigens are synthesized, processed, and presented through a non-canonical mechanism that utilizes cap-independent translation and co-translational quality control to facilitate their presentation on MHC-I for immunosurveillance. Our preliminary analysis demonstrates that specific translation inhibitors that promote cap-independent translation alter the antigenic landscape of lung cancer. In this proposal, we will build upon these observations and utilize a combination of sophisticated genetically engineered mouse models (GEMMs) and human cancer cell lines to define the mechanisms that influence nuORF biosynthesis, processing, antigen presentation, and CD8+ T cell recognition. In Aim 1, we will evaluate how pharmacological and physiological manipulation of translation alters presentation and recognition of nuORF derived antigens. In Aim 2, we will assess the contribution of co-translational quality control mechanisms in shaping non-canonical antigen presentation. Ultimately, the goal of this proposal is to understand the mechanisms by which nuORFs are presented to more effectively engage this class of targets for immunotherapy.
