Study On Pharmacological Effect Of Elemene Extract From Curcuma Wenyujin On 5-FU Resistance Of Colorectal Cancer

Objective:Colorectal cancer is a common malignant tumour that occurs in the mucosal epithelium,with the fourth highest incidence and third highest mortality rate of malignant tumours worldwide.Currently,chemotherapy based on 5-fluorouracil（5-FU）and other drugs is the main regimen for the clinical treatment of advanced colorectal cancer,but during the treatment process,tumour cells become resistant to the chemical drugs resulting in unsatisfactory treatment results for cancer patients,which is one of the main reasons for the clinical failure of cancer patients.Therefore,developing effective new anti-cancer drugs or exploring combinations between chemotherapeutic drugs will help in the treatment of cancer.Elemene is a potent anticancer active ingredient extracted from Chinese herbs such as Curcuma wenyujin.Studies have shown thatβ-elemene is the main active ingredient in elemene,and can be involved in regulating tumour cell resistance through multiple targets and channels.However,the mechanism of action ofβ-elemene in 5-FU resistance in colorectal cancer is still unclear.In this study,we will establish colorectal cancer 5-FU resistant cell lines and investigate the biological function differences of colorectal cancer 5-FU resistant cells,focusing on 1)the molecular mechanism ofβ-elemicene in reversing 5-FU resistance in colorectal cancer cells by regulating iron death;2)the molecular mechanism of m⁶A methylation modification in regulating iron death and its role in reversing 5-FU resistance in colorectal cancer cells byβ-elemicene.Methods:（1）The colorectal cancer cell lines HCT116 and Lo Vo cell lines were used as the parents to construct 5-FU resistant cell lines HCT116FR and Lo Vo FR,respectively;the sensitivity of colorectal cancer cells to 5-FU was detected by CCK-8assay,flow cytometry,cell invasion and cell scratch assay;（2）the sensitivity of colorectal cancer cells to 5-FU was detected by CCK-8 assay,flow cytometry,flat dish monoclonal formation and animal assays to analyze the role ofβ-elemene in 5-FU resistance in colorectal cancer cells;（3）The mechanism of action ofβ-elemene in reversing 5-FU resistance in colorectal cancer cells through the regulation of iron death was investigated by Western Blot and q RT-PCR.（4）Through transfection of si RNA,Western Blot and q RT-PCR,the molecular mechanism of m⁶A methylation to regulate iron death and its mechanism ofβ-elemene in reversing 5-FU resistance of colorectal cancer cells were studied.Results:（1）The results of biological function studies showed that the IC₅₀ of resistant cells to 5-FU was significantly enhanced compared with parental cells;the proportion of apoptosis in 5-FU-treated resistant cells was significantly reduced;the migration invasion ability of resistant cells was significantly increased.（2）β-elemene significantly increased ROS and ferric ion levels and decreased intracellular GSH levels in drug-resistant cells;both Fer-1 and Lipo-1,iron death inhibitors,significantly alleviated the inhibitory effect ofβ-elemene on drug-resistant cells;（3）flow cytometry results showed that drug-resistant cells overexpressing p53 were less sensitive toβ-elemene,and p53 significantly upregulated GPX4,SLC7A11,and p21 protein expression;（4）the results of flow cytometry,Western Blot,intracellular ROS and iron ion concentration assays showed that knockdown of m⁶A demethyltransferase FTO significantly reduced the apoptotic proportion,intracellular ROS concentration and iron ion concentration in colorectal cancer resistant cells,and FTO was regulating the iron death-related signaling protein expression.Conclusion:β-elemene reverses 5-FU resistance in colorectal cancer cells through iron death;p53 plays an inhibitory role inβ-elemene reversal of 5-FU resistance in colorectal cancer;desmethyltransferase FTO regulates iron death through the p53/GPX4 pathway and is involved inβ-elemene reversal of 5-FU resistance in colorectal cancer.