Nattokinase enhances the sensitivity of cancer cells to oxaliplatin through mitochondrial pathway and induction of apoptosis

Combination therapy consisting of anticancer drugs has been used to decrease the adverse effects and increase the effectiveness of the drugs. The present study aimed to examine the synergistic effect and potential mechanisms of oxaliplatin and nattokinase (NK) combined treatment for BGC-823, HepG-2, HeLa, and NCI-H460 cells. NK was isolated and purified from black soybean natto by dialysis and gel filtration chromatography, which had a nattokinase activity of 600,391 U/g. MTT experiment, clone formation assay, Hoechst 33,342 staining, DCFH-DA staining, and JC-1 staining showed that NK had no cytotoxicity and oxaliplatin could inhibit cell proliferation. Oxaliplatin combined with NK could enhance its anticancer effect and lead to mitochondrial dysfunction by causing the increase of intracellular ROS, which eventually induced apoptosis. Compared to oxaliplatin alone, nattokinase in combination with oxaliplatin can effectively inhibit DNA synthesis and increase cell uptake, ultimately inhibiting cell proliferation. Transwell and angiogenesis experiments proved that the combination of NK and oxaliplatin could more effectively inhibit cell migration, invasion, and in vitro angiogenesis, thereby preventing the spread and metastasis of tumors. RT-qPCR showed that the mechanism of inhibition of cell proliferation and thus induction of apoptosis by the combination of NK and oxaliplatin may be achieved through the upregulation of BAD, BAX, and Caspase-3 genes. Molecular docking results show that NK could act as the carrier of oxaliplatin. In conclusion, the above results suggested that NK could play a synergistic anticancer role with oxaliplatin by inducing apoptosis through the mitochondrial pathway, enhance the sensitivity of cancer cells to oxaliplatin, and provide a new strategy for cancer treatment.