A 3D-Printed Biochemical-Mechano-Thermally Responsive Multicellular Tumor Organoid Platform Decodes Biochemical-Physical Regulation of Cancer-Immune Crosstalk and Chemoresistance.

Oral tumors are not shaped by biochemical signals alone; they also evolve within mechanically active and thermally fluctuating microenvironments that critically regulate malignant progression, immune remodeling, and therapeutic resistance. Here, we develop a dynamic bioorthogonal organoid platform that couples a 3D-printed photothermal elastic scaffold with an interpenetrating hydrogel network derived from decellularized oral mucosa extracellular matrix, enabling the reconstruction of in vivo-like structural support, matrix-derived biochemical cues, spatial cellular compartmentalization, and dynamic physical stimulation. Through click chemistry-mediated cell anchorage and hydrogel encapsulation, oral cancer cells and macrophages are spatially organized into a compartmentalized cancer-immune niche. Under cyclic mechanical stretch and near-infrared-triggered periodic photothermal stimulation, dynamic culture markedly enhances cancer cell proliferation and glycolytic reprogramming while concomitantly polarizing macrophages toward a pro-tumoral M2 phenotype. In both monoculture and coculture settings, biochemical-mechano-thermal stimulation substantially increases cancer cell chemoresistance. Mechanistically, these biochemical-physical cues activate stress-adaptive and survival programs while enriching cancer stem-like populations. In a xenograft mouse model exposed to in vivo biochemical, mechanical and thermal stimulation, dynamic conditioning markedly accelerates tumor progression, enhances metastatic potential, and confers pronounced chemoresistance. Together, this study establishes a biomimetic multicellular tumor platform for uncovering how dynamic biochemical-physical cues orchestrate cancer-immune crosstalk and therapeutic failure.