Herein, we first describe a perfusion chip integrated with an AC electrothermal (ACET) micromixer to supply a uniform drug concentration to tumor cells. The in-plane fluid microvortices for mixing were generated by six pairs of reconstructed novel ACET asymmetric electrodes. To enhance the mixing efficiency, the novel ACET electrodes with rotating angles of 0°, 30°, and 60° were investigated. The asymmetric electrodes with a rotating angle of 60° exhibited the highest mixing efficiency by both simulated and experimental results. The length of the mixing area is 7 mm, and the mixing efficiency is 89.12% (approximate complete mixing) at a voltage of 3 V and a frequency of 500 kHz. The applicability of our micromixer with electrodes rotating at 60° was demonstrated by the drug (tamoxifen) test of human breast cancer cells (MCF-7) for five days, which implies that our ACET in-plane microvortices micromixer has great potential for the application of drug induced rapid death of tumor cells and mixing of biomaterials in organs-on-a-chip systems.