Construction of oral squamous cell carcinoma organoids in vitro 3D‐culture for drug screening

Abstract Objective Patient‐derived organoids are potent pre‐chemotherapy models. Due to limited research on diverse types of oral squamous cell carcinoma (OSCC) and construction efficiency, our goal was to optimize OSCC organoid models from various sites and assess drug responsiveness. Methods We screened and optimized culture media, employing three‐dimensional techniques to construct human‐derived oral squamous cell carcinoma (OSCC) organoid models in vitro. Morphological validation, immunofluorescence analysis, tissue origin verification, and Short Tandem Repeat (STR) sequencing confirmed the consistency between organoids and source tissues. These organoid models were then subjected to varying concentrations of anticancer drugs, with subsequent assessment of cell viability to calculate IC50 values. Results Twenty‐nine surgical specimens yielded an 86.2% success rate in culturing 25 organoids in vitro. Morphological consistency confirmed nuclear atypia and positive expression of K5, P40, and E‐cadherin, indicating squamous epithelial origin. Cultured complex organoids included α‐SMA+ tumour‐associated fibroblasts and tumour stem cells expressing CD44 and Ki67. STR sequencing affirmed genomic homogeneity between cultured organoids and source tissues. Drug sensitivity testing revealed diverse responses among organoids, highlighting their value for assessing drug sensitivity. Conclusions An efficient OSCC organoid culture system for personalized in vitro drug sensitivity screening was established, laying the foundation for precise treatment development.