The Effects Of LDH-A Inhibition By Oxamate On Thegrowth And Radiosensitivity In Nasopharyngealcarcinoma Cells

Purpose: An elevated rate of glucose consumption and the dependency on aerobicglycolysis for ATP generation have long been observed in cancer cells, a phenomenonknown as the Warburg effect, the altered energy metabolism in cancer cells provides anattractive opportunity for developing novel cancer therapeutic strategies. Lactatedehydrogenase(LDH), which catalyzes the transformation of pyruvate to lactate, plays avital role in the process of glycolysis. It has been reported that the level of LDH-Aexpression was increased both in head and neck cancer cells and blood serum of NPCpatients, and was associated with poor prognosis. However, the effect of LDH-A inhibitionis little known in NPC cells. In this study,the effect of LDH-A inhibitor Oxamate on thegrowth and radiosensitivity was explored and the involving mechanism was also studied.Methods: Human NPC cell lines CNE-1and CNE-2were choosenas the study obeject.1) We used MTT (methye thiazolye telrazlium) assay to test the influence of Oxa on cellviability at different concentrations and times.2) Scratch healing assay and the transwell experiment were used to study the influence ofoxamate sodium on the cell migration and invasion.3) Flow cytometry was applied to analyze cell cycle progression and Annexin V-FITC cellapoptosis kit was used detect apoptosis and necrosis.4) ROS content, LDH activity and ATP content were also detected by related kits.5) JC-1staining was used to test the change of mitochondrial membrane potential.6) Western blot was used to detect the changes of protein expression. 7) Colony formation method was employed to study the effects of oxamate sodium onradiosensitivity of NPC cells.8) H2AX foci analysis was used to detect DNA damage after irradiation.Results:1) Oxamate inhibited the growth and proliferation of CNE-1and CNE-2cells in dose-andtime-dependent way, the IC50s were74.6mmol/L,32.4mmol/L and17.8mmol/L forCNE-1cells at24h,48h,72h and62.3mmol/L,44.5mmol/L,31.6mmol/L forCNE-2cells at24h,48h,72h repectively.2) LDH-A inhibition by oxamate induced G2/M cell cycle arrest via down-regulation ofCDK1/cyclinB1pathway.3) LDH-A inhibition by oxamate promoted apoptosis through enhancing mitochondrialROS generation,4) We also identified that oxamate increased sensitivity to irradiation. The SER for CNE-1and CNE-2were1.26and1.37respectively.5) Furthermore, we verified the similar results in the tumor xenograft model. These resultsindicated that LHA might serve as an attractive therapeutic target for NPC treatment.Conclusion: In conclusion, this study provides evidence for LDH-A inhibition in NPCtreatment alone or combined with irradiation both in vitro and vivo, and indicates thattargeting glycolysis may be an effective strategy for NPC therapy. Further studies arerequired to explore whether inhibition of other key enzymes in the glycolytic pathway havesimilar effects as the inhibition of LDH-A and still more efforts are needed to developnovel high effective inhibitors of LDH-A enzyme in the future.