A 3D Ex Vivo Tumor-Immune Coculture System Mimicking In Vivo Tumor Environmental Stress on CD8+ T Cells Exhaustion.

Dissection of exhaustion trajectories of immune cells under tumor selection pressure in the tumor microenvironment (TME) elucidates the underlying machinery in anti-tumor immunity, which still lacks easy-to-use models to decipher. Herein, gelatin methacryloyl (GelMA)-poly (ethylene oxide) (PEO) based 3D hydrogel microspheroids are constructed with non-immunogenicity and controllable macroporous structure to establish a tumor-immune cell coculture (3D-HyGTIC) system. In 3D-HyGTIC system, when immune cells embarked, stepwise up-regulation of main immune checkpoints (ICs) molecules is observed with compromised cytokine production in CD8 T cells, the trajectory of which is in lineage correlation with in vivo grafted tumors. Reinvigoration of CD8 T cells is more obvious with the addition of an anti-PD-1 regimen at the early time point, which is recapitulated during the coculture of patient-derived tumor fragments (PDTF) and autologous T cells. Moreover, the upregulation of LAG-3 on CD8 T cells after anti-PD-1 treatment is uncovered. Sequential addition of anti-LAG-3 successfully rescues the otherwise failed reactivation of CD8 T cells. Therefore, the 3D-HyGTIC system is not only inclined to mimic the early differentiation trajectories of tumor-infiltrating CD8 T cells but also may facilitate an evaluation of the efficacy of IC blockades and guide the designing of combination immunotherapy.