Abstract P46: Development of a Robust Cryopreservation Protocol and Culture Platform for Patient-Derived Tumor Explants

Background: The discovery of effective anti-cancer drugs is limited by the use of reductionist tumor models, such as organoid or spheroid cultures, which fail to accurately recapitulate patient-specific heterogeneity in tumor architecture and composition. Patient-derived tumor explants (PDTE) preserve many features of the original tumor, but challenges such as short-term ex vivo viability and the often lengthy logistics of fresh sample transfer from clinical centers to laboratories remain. Establishing a robust workflow for PDTE cryopreservation and the development of a supportive ex vivo culture platform tailed for specific tumor types may enhance clinical utility of PDTEs. Methods: In this study, freshly resected patient tumors (melanoma, ovarian cancer, sarcoma) were collected from the National Cancer Center Singapore (NCCS). Tumors were immediately processed into fragments of varying sizes (0.3, 0.6, 1 cm in diameter) and precision-cut slices (300 μm thick), then cryopreserved using three different freezing media. Samples were thawed at designated time points (1, 6, 12 months) for evaluation of tumor viability through cell viability assays and histological analyses. Methacrylated hyaluronic acid (MeHA) hydrogels with tunable mechanical properties were used to identify optimized ex vivo conditions for culture of cryopreserved PDTEs. Results and Conclusions We found that freezing media formulations with reduced fetal bovine serum (20% and 40%) effectively maintained tumor viability, similar to conventional formulations (90% FBS + 10% DMSO). Notably, significant tissue damage was observed at the tissue periphery (1 mm depth) post-freeze-thaw, suggesting that larger tissue specimens (1 cm) better retain overall viability and structural integrity. Furthermore, MeHA hydrogels with varying stiffnesses (100-2,400 Pa) and pore sizes (50, 110, 250 μm) were successfully fabricated. Studies are ongoing to evaluate how hydrogels influence tissue macrostructure, cell viability, migration, and tumor architecture. Furthermore, we will determine whether different tumor types necessitate specific hydrogel conditions for optimal ex vivo PDTE culture. Citation Format: Kenny Zhuoran Wu, Ruochii You, Hengjia Zhou, Madhumathi Kalaichelvan, Haiyao Ding, Kevin Philip Ying Zhi Fong, Dong Hua Seah, Johnny Chin-Ann Ong, Eliza Li Shan Fong. Development of a Robust Cryopreservation Protocol and Culture Platform for Patient-Derived Tumor Explants [abstract]. In: Proceedings of Frontiers in Cancer Science 2024; 2024 Nov 13-15; Singapore. Philadelphia (PA): AACR; Cancer Res 2025;85(15_Suppl):Abstract nr P46.