Autophagy Facilitates Metadherin-induced Chemotherapy Resistance Through The AMPK/ATG5 Pathway In Gastric Cancer

Objectives:Metadherin(MTDH)is overexpressed in some malignancies and enhances drug resistance;however,its role in gastric cancer(GC)and the underlying mechanisms remain largely unexplored.Here,we explore whether MTDH induces the GC drug resistance and the mechanism by which MTDH cation.Methods:We analysed the levels of MTDH in gastric cancer and adjacent normal gastric mucosal tissues by Real-time quantitative PCR(q-PCR)immunohistochemical staining.We also analysed the level of autophagy by western blot?confocal microscopy and transmission electron microscopy after MTDH knockdown and overexpression,and examined the fluorouracil(5-FU)resistance by CCK-8 and exzmined the Multidrug resistance gene(MDR1)by q-PCR and western blot after MTDH overexpression.Next,we inhibited autophagy by knockdown of autophagy related protein 5(ATG5),and examined the fluorouracil(5-FU)resistance by CCK-8 and exzmined the Multidrug resistance gene(MDR1)again by q-PCR and western blot after MTDH overexpression.AMPK knockdown siRNA and pathway inhibitor Compound C were applied to determine the molecular mechanisms of autophagy.Finally,the GC patient-derived xenograft(PDX)tumours were used to prove the 5-FU resistance.Results:We found that the expression of MTDH significantly increased in the gastric cancer specimens compared to the adjacent normal gastric mucosal tissues.Further study showed a positive correlation between the MTDH expression level and 5-FU resistance.Overexpression of MTDH in MKN45 cells increased the levels of P-gp and promoted 5-FU resistance,while inhibition of MTDH showed the opposite result.The inhibition of autophagy and overexpress MTDH at the same time decreased the levels of P-gp and inhibited 5-FU resistance,proved that MTDH promoting autophagy,then promote gastric cancer chemotherapy drug resistance.Moreover,MTDH induced AMPK phosphorylation,regulated ATG5 expression,and finally influenced autophagy,suggesting that MTDH may activate autophagy via AMPK/ATG5 signalling pathway.AMPK knockdown siRNA and pathway inhibitor Compound C validated the molecular mechanisms of autophagy.Conclusions:MTDH-stimulated cancer fluorouracil(5-FU)resistance may be mediated through autophagy activated by AMPK/ATG5 pathway in gastric cancer.And MTDH played a significant role on this process.Targeting this mediated route may help improve the efficacy of chemotherapy in gastric cancer.