Dual-channel six-step linear concentration gradient microfluidic chip for orthogonal combination drug screening in head and neck tumor cells.

Head and neck cancer, as the seventh most common cancer worldwide, is in urgent need of an efficient drug screening platform due to tumor heterogeneity and the lack of objective markers for conventional chemotherapy resulting in limited efficacy. In this study, we developed a dual-channel microfluidic chip integrating organoid technology and hydrodynamic modulation, and realized six-chamber linear drug concentration gradient generation (<3% fluctuation at a flow rate of 10-100 μL min-1, stable for 24 hours) through a three-layer PDMS structure and serpentine microchannel design. After 24 h of treatment with four drugs, CAL-27 cell viability in the chip decreased with increasing drug concentration, showing no significant difference from conventional assays. Using a Matrigel 3D scaffold, the culture time of spheroids was reduced by 24 h compared with monolayer cells, and the organoid morphology remained unchanged before and after drug addition (P > 0.05). Fluorescence staining revealed distinct responses to the four drugs near their IC50 concentrations (P < 0.01). The platform integrates fluorescence imaging and CFD simulation, which has the advantages of automation, high mimicry and low consumables compared with the off-chip method, providing a reliable tool for the optimization of the precision treatment plan for head and neck cancer.