Reverse Engineering Tumors to Enhance Neoantigen Expression for Immunotherapy

Many cancer types lack an immunogenic target for immune cells to detect, thereby limiting the effectiveness of immunotherapy. These immunologically “cold” tumors are marked by low neoantigen expression and minimal immune infiltration. We hypothesize that strategies that increase tumor immunogenicity such as inducing neoantigen expression or enhancing antigen presentation can convert cold tumors into “hot”, thus improving responsiveness to immune-based treatments. Using a novel pH low insertion peptide (pHLIP) that penetrates cell membranes in an acidic microenvironment, we have successfully shown tumor-targeted delivery of cargo peptides (e.g. SIINFEKL) into transplanted B16 (melanoma) and MC38 (colon adenocarcinoma) cell lines. Furthermore, SIINFEKL was proteolytically processed through the MHC-I antigen presentation pathway, induced OT-I antigen-specific T cells, and led to delayed tumor growth (Yurkevicz et al, Mol. Cell Therapy). Using a virus-induced Kras mutant p53 null (KP) mouse model of sarcoma, we tested whether pHLIP could deliver SIINFEKL into autochthonous de novo tumors and induce OT-I-specific T cells. We isolated T cells from these KP tumors and observed that only mice treated with pHLIP-SIINFEKL exhibited OT-I-specific T cell infiltration (SIINFEKL-MHC-I-Tetramer+) and effector activation (CD44+, IFNg+), indicating that pHLIP can function in a physiological context to deliver neoantigens and trigger an immunogenic response. These findings demonstrate a novel strategy to enhance sarcoma immunogenicity and may inform future immunotherapy approaches. Citation Format: Julie F. Cheung, Annali Yurkevicz. Reverse engineering tumors to expression neoantigens [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2026; Part 1 (Regular Abstracts); 2026 Apr 17-22; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2026;86(7 Suppl):Abstract nr 4255.