Mechanism Of Non-coding RNA In Paclitaxel Therapy For Non-small Cell Lung Cancer

Background:Paclitaxel?PTX?is one of the first-line chemotherapy drugs for advanced non-small cell lung cancer?NSCLC?.The long-chain non-coding RNA?IncRNA?maternal expression gene 3?Maternal expression gene 3,MEG3?is a recognized tumor suppressor.CircRNA hsa_circ₀002874 has been reported to be associated with doxorubicin resistance in breast cancer MCF-7 cells.Other studies have shown that some ncRNA may be regulated by chemotherapeutic agents such as cisplatin and PTX.Objective:To investigate the effects of PTX on the expression of non-small cell lung cancer A549 cell line MEG3,hsa_circ₀002874 and its downstream molecular pathways;to study the association of related molecular pathways with phenotypic changes such as apoptosis;to explore the new anti-tumor mechanism of PTX.Methods:In order to better clarify the role of ncRNA in this study and its role in PTX administration,IncRNA MEG3 was identified by literature review and pre-experimental robustness verification;circRNA hsa_circ₀002874 was screened by 18 highly conserved circRNAs from reported gene chip results.Out.The MTT assay is used to assess the metabolic activity of the cells.Quantitative polymerase chain reaction?qPCR?is used to detect the amount of molecule to be tested.Western blot?WB?and immunofluorescence?IF?were used to detect the level of protein to be tested.siRNAs are used to down-regulate lncRNA,circRNA or miRNA expression;pCDNA plasmid transfection is used to up-regulate IncRNA or circRNA expression;mimic is used to up-regulate miRNA expression.Alien nuclear analysis was analyzed by DAPI staining.Flow cytometry?FCM?was used to analyze apoptosis.Plate cloning experiments were performed to analyze the growth state of the cells.The fluorescent probe H2DCF-DA was used to detect intracellular reactive oxygen species?ROS?.The P53 protein inhibitor PTFa is used to inhibit intracellular P53 activity.Dual luciferase reporter assays are used to detect specific binding of miRNA to target messenger RNA?mRNA?.Results:PTX significantly inhibits the proliferation of non-small cell lung cancer cells.PTX increased the expression of MEG3 and P53.Down-regulation of MEG3 reduces P53 protein expression and attenuates PTX-induced cytotoxicity;at the same time,up-regulation of MEG3 increases P53 expression and induced cell death.PTX increases P53 protein expression via the hsa_circ₀002874-miR1273f-MDM2-P53 pathway.The dual luciferase reporter assay confirmed that miR1273f specifically binds to MDM2 mRNA.Down-regulation of hsa_circ₀002874 increased miR1273f expression,thereby reducing MDM2 expression and increasing P53 protein expression;at the same time,up-regulation of hsa_circ₀002874 reduced miR1273f expression,thereby increasing MDM2 expression and decreasing P53 protein expression.Down-regulation of miR1273f increased MDM2 expression and decreased P53 protein expression;at the same time,up-regulation of miR1273f reduced MDM2 expression and increased P53 protein expression.In addition,inhibitors of P53 did not affect upstream MEG3,hsa_circ₀002874 and miR1273f expression.Conclusion:Our results indicate that ncRNA is a novel anti-tumor mechanism of PTX.The MEG3-P53 pathway and the hsa_circ₀002874-miR1273f-MDM2-P53 pathway are involved in PTX-induced apoptosis of A549 cells.This reflects that ncRNA can play a key regulatory role in the treatment,prognosis and chemotherapy drug resistance of NSCLC.