GOLPH3 Regulates Glycometabolism Reprogram To Affect The Stemness Of Tumor Cells In Chemotherapy Resistance Of Breast Cancer

Objective: Breast cancer is the most common malignant tumor in women.Chemotherapy is the most important treatment and adjuvant therapy except surgical excision.However,due to tumor heterogeneity,breast cancer is prone to relapse,metastasis and drug resistance after treatment.The extracorporeal peripheral membrane protein GOLPH3 has been found to be highly expressed in various cancers and to predict poor prognosis.It has been reported in the literature that the increased expression of GOLPH3 can repair the DNA damage of cancer cells after the tumor cells are killed by chemotherapy drugs,leading to increased survival and promoting the generation of tumor drug resistance.In addition,tumor stem cells are considered to be very important factors in promoting drug resistance and recurrence of cancer,and they generally obtain their energy needs through glycolysis.Therefore,this study aims to explore the effects of GOLPH3 on glucose metabolism and cell dryness of tumor cells and its relationship with drug resistance of breast cancer cells,so as to find a new therapeutic strategy for preventing drug resistance of breast cancer.Methods: In this study,the effects of Golph3-induced changes in glucose metabolism and cell dryness on drug resistance of tumor cells were investigated using both in vitro and in vivo experiments.In this study,the constructed drug-resistant breast cancer cell lines were used to knock down GOLPH3 to observe the effect of GOLPH3 on cell activity and the changes of glucose uptake rate and lactic acid release after low expression of GOLPH3.At the same time,bioinformatics analyzed the changes of related genes in the GOLPH3 low expression group and the control group.In addition,q PCR and WB were used to verify the changes of key enzymes of glucose metabolism and cell dryness genes.After that,HK2 overexpression plasmid was constructed and transfected into drug-resistant MCF7 cells with low expression of GOLPH3.q PCR and WB were used to observe the changes of genes related to cell dryness.After HK2 was overexpressed,the number of CD44+ cells in the overexpressed group and the control group was detected by flow cytometry.In this study,breast cancer cells with low expression of GOLPH3 were injected into mouse fat pad to construct breast cancer transplant tumor model.Mice in KD+DXT group and NC+DXT group were injected intravenically once a week with docetaxel at the dosage of 15mg/kg.The changes of transplanted tumor were observed and recorded,and the effects of GOLPH3 on drugs and tumors were evaluated.In addition,energy metabolite analysis was performed on the obtained tumor samples to observe the pathways that may be affected.The expression of HK2 in tumor samples obtained by different treatments was determined by immunofluorescence assay.Results: Experiments have shown that the cell viability of breast cancer resistant cells decreased after knocking down GOLPH3.Lactic acid release and glucose absorption decreased in aerobic conditions.The results of biogenic analysis showed that the stem and metabolism-related pathways were significantly enriched,and HK2 and CD44 were significantly decreased in the low expression group of GOLPH3.The results of q PCR and WB showed that the expressions of HK2,a key glycolysis enzyme,and CD44,were decreased in the Golph3-knockdown resistant cells,and the expressions of HK2 and CD44 were increased after the overexpression of HK2,a key glycolysis enzyme,indicating that the dryness of breast cancer cells depended on glycolysis and was regulated by GOLPH3.Flow cytometry showed that the number of CD44 positive cells increased significantly,indicating that the dry capacity of tumor cells was positively correlated with glucose metabolism.In vivo animal experiments have also proved the above conclusion,the state and tumor size of mice with low expression of GOLPH3 are significantly inhibited,and when GOLPH3 is knocked down in combination with drugs,the therapeutic effect of mouse breast cancer is significantly enhanced,and the drug resistance is reduced.The results of energy metabolite targeting analysis also showed that the glucose metabolism pathway,lactic acid and glutamine were significantly reduced in the tumor tissue of mice with GOPLH3 knockdown.Immunofluorescence also showed that the red fluorescence representing HK2 was significantly reduced,and GOLPH3 was positively correlated with the metabolism of sugar in the body.Conclusion: The results show that GOLPH3 affects the key enzyme of glucose metabolism HK2,leading to the reprogramming of glucose metabolism in vivo,promoting the survival of breast cancer stem cells CSCs and mediating drug resistance of breast cancer.