3D printed microfluidic chip for multiple anticancer drug combinations

•First 3D printed multi-drug combinations microfluidic chips.•Microchannels with a helical structure promote rapid and efficient mixing of solutions.•The four drugs are spatially mixed in the channels and formed 36 combinations of different concentrations and ratios.•The combined effect of multiple drugs are demonstrated by the viability test of A549 cells. Multi-drug combinations therapy is a promising method in the fight against cancer cells. Multi-drug combinations cannot be practically elucidated using conventional devices owing to structural limitations or operational complexity. 3D printing has recently attracted attention as a way to fabricate complicated and interconnected microfluidic channels, which is of design flexibility. Here, an easy-to-use, high-throughput, multi-drug combinations 3D printed microfluidic chips were brought forward. 36 discrete concentration combinations have been generated by repeated splitting-and-mixing of four inlet drugs in interconnected network channels, which are used to determine the optimal concentration of inhibition of cancer cells survival. We evaluate the potential to advance multi-drug combinations efficiency by optimizing the geometric parameters of tree-shaped branch unit, as well as the chip performance by dye visualization. The microfluidic chip was further applied to study multi-drug combinations responses, and the 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium, inner salt (MTS) assay was employed to test the cytotoxic effect of four drugs combinations on A549. The experiment results demonstrated clinically relevant antagonistic, synergistic interactions between the four different antitumor drugs. Moreover, this chip has outstanding features, e.g., more compact structure, more accurate combination of concentrations, greater space for scalability. As such, it will be an effective multi-drug combinations screening platform for applications in biomedical research and clinical medicine.