| Objective:This study aims to explore the role and mechanism of GLS1 in ferroptosis of pancreatic cancer,and successfully synthesized a new nanomaterial to combine GLS1therapeutic targets with ferroptosis,providing new ideas and insights for the targeted therapy of pancreatic cancer.Methods:（1）The correlation between GLS1 and Epithelial-Mesenchymal Transition of pancreatic cancer was analyzed based on CCLE database and GEO database.（2）Western blot and q PCR were used to interfere GLS1 expression in Panc1cells and to verify its interfering efficiency,clarifying the role of AMPK-mTOR pathway in GLS1-mediated ferroptosis resistance of pancreatic cancer.（3）The content of malondialdehyde（MDA）,ferric divalent ion(Fe2+)and lipid ROS in the cells was determined to verify the effect of GLS1 removal on the sensitivity of pancreatic cancer cells to ferroptosis and further clarify the role of AMPK-MTOR and GLS1 in inducing ferroptosis resistance in pancreatic cancer.（4）To evaluate the effect of GLS1 and AMPK-mTOR pathway on ferroptosis of pancreatic cancer in mice with pancreatic cancer.（5）Construction of PA@Au loaded with GLS1 inhibitor and material characterization analysis.（6）The cytotoxicity and cell uptake rate of m PPt-JHU were measured by CCK-8assay and laser confocal.（7）The sensitization effect of m PPt-JHU on ferroptosis caused by Erastin was verified in vitro by Western blot assay and cell malondialdehyde（MDA）assay.（8）The sensitization effect of m PPt-JHU on ferroptosis caused by Erastin was verified in vivo by subcutaneous tumor model of pancreatic cancer in mice.Results:（1）CCLE database and GEO database data showed that GLS1 was negatively correlated with epithelial type indicator E-cadherin,and positively correlated with mesenchymal type indicator Snailer,V-imentin,N-cadherin,ZEB1 and MMP family,and surface GLS1 promoted mesenchymal transformation of tumors.（2）After the addition of Erastin in pancreatic cancer cells,the GPX4 and GLS1 protein level decreased with the occurrence of ferroptosis.（3）Knockdown GLS1 expression in pancreatic cancer cells made tumor cells show increased sensitivity to ferroptosis（4）Knockdown GLS1 expression in pancreatic cancer cells madethe phosphorylation level of AMPK increased and that of mTOR decreased,but after adding AMPK inhibitor and mTOR agonist to pancreatic cancer cells that interfere with GLS1expression can inhibit the trend of increased susceptibility to ferroptosis.（5）Mice with intraperitoneal metastatic tumor of pancreatic carcinoma demonstrated that GLS1-AMPK-mTOR plays an important role in ferroptosis resistance of pancreatic cancer cells in vivo（6）The molecular probe（m PPt-JHU）was constructed to load GLS1 inhibitor JHU-083,which has high drug loading efficacy,stable physical and chemical properties,good biocompatibility and cell community efficacy.In vitro and vivo results showed that m PPt-JHU effectively promoted erastin-induced ferroptosis in pancreatic cancer cells.Conclusion:The regulation of AMPK-mTOR pathway by GLS1 can enhance the resistance of pancreatic cancer cells to ferroptosis.m PPt-JHU can effectively load GLS1inhibitor and realize ferroptosis sensitization in vitro and vivo.This study provides a new idea and direction for the treatment of pancreatic cancer. |
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