Abstract
BACKGROUND: Lung cancer remains the leading cause of cancer-related mortality, with many patients responding poorly to immunotherapy due to limited tumor recognition. Neoantigen-based strategies offer a promising solution, but current discovery methods often miss key targets, particularly those with low or heterogeneous expression. To address this, we developed ImmuniT, a three-phase platform for enhanced neoantigen discovery and validation. METHODS: Under an IRB-approved protocol, patients with lung cancer consented to tumor collection for ex vivo processing to modulate antigen expression. Autologous T cells from matched blood were co-cultured with treated cancer cells to expand tumor-reactive populations. The nextneopi pipeline integrated mutational, transcriptomic, and HLA data to predict candidate neoantigens, which were validated using MHCepitope tetramer staining. RESULTS: In five patient samples, ImmuniT identified a broader spectrum of neoantigens and induced stronger T cell activation in vitro compared to conventional approaches. Notably, in one case, two neoantigens missed by standard methods were confirmed to elicit tumor-specific T cell responses in both the tumor-infiltrating and peripheral compartments. CONCLUSIONS: These findings highlight ImmuniT's potential to expand the repertoire of actionable tumor antigens and improve personalized immunotherapy strategies, particularly for patients with limited response to existing treatments.