Abstract 2843: Neoantigen evaluation in viably cryopreserved dissociated tissue via large scale proteomic profiling

Abstract Genomic instability is a hallmark of cancer where the acquisition of numerous genetic alterations leads to transformation and carcinogenesis. These alterations result in the generation of potentially immunogenic neoantigens, and the clinical success of checkpoint inhibitor therapies has highlighted the importance of neoantigens in cancer immunotherapy. Evaluation of the next generation of neoantigen therapies is complicated by the difficulty in sourcing primary patient material. During tumor resection and pathology review, the tissue is fixed, which prevents the ability to test in vitro model systems for therapy efficacy. While fresh tissue is amendable to in vitro culture, it is not possible to perform large scale genomic and proteomic screens in a timeframe for the culture to occur. By leveraging viably cryopreserved dissociated tissue, the tumor can be analyzed for genetic alterations and then, when novel neoantigens are determined, cultured at a later date. To test this workflow, we profiled dissociated tissues from cancer indications with a high degree of genomic instability for microsatellite instability (MSI). Consistent with previous studies, 17-55% of endometrial, colorectal, and gastric cancer samples were MSI-high. MSI-high and microsatellite stable (MSS) samples were further analyzed for peptide identification using LC-MS/MS (Orbitrap Astral) and over 10,000 protein groups and 80,000 peptides were identified. Samples were also amendable to T cell receptor (TCR) repertoire sequencing to uncover tumor-reactive T cell clones. These deep molecular analyses provided the foundation for the development of short-term spheroid and long-term tumoroid culture systems to understand immune responses using autologous tumor-infiltrating and peripheral lymphocytes. Collectively, this workflow provides a scalable system to identify neoantigens and evaluate the immune response to these antigens in a robust and reproducible pipeline. Citation Format: Shawn P. Fahl, Casey Smith, Kerri Colwell, Jessica Moore, Jaison Arivalagan, Danielle Gutierrez. Neoantigen evaluation in viably cryopreserved dissociated tissue via large scale proteomic profiling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 2843.