Research On Antiglioma Drugs Targeting Novel Inhibitors Of EGFR And CXCR4

Objection: Multiforme Glioblastomas(GBMs)is one of the most aggressive and fatal malignant tumors in the brain.There is short of efficient therapeutic regime at present and of great significance to find a new type treatment methods.Molecular targeted therapy has better selectivity and specificity than traditional therapy,which is one of the hot issues in the field of tumor therapy.EGFRv III is expressed specifically in tumor cells but not in normal tissues.Especially high expression in GBMs patients,30% of patients have EGFRv III gene mutation,which is an ideal target for GBMs targeted therapy.Some drugs have been used to target EGFR and achieved some clinical effects in present,however,they have little effect in GBMs treatment.How to overcome the primary or secondary drug resistance of the targeted treatment is still a larger challenge because of its complex mechanism.TNF-JNK-Axl-ERK signal pathway is involved in the role of EGFR in tumor growth and migration.Therefore,the project will design a double target combination therapy plan for the related effect of EGFR and TNT in intracellular signal transduction.The therapeutic effect of the combined regimen was evaluated by molecular and cellular biology in vitro and transplanted tumor model of nude mice in vivo,and its mechanism of anti glioma was studied at the molecular level.The successful implementation of this project is expected to provide new ideas for GBMs molecular targeted combination therapy,which has important theoretical significance and clinical practical value.Methods:U87 glioma cells be transfected EGFR gene were cultured in vitro and treated with EGFR inhibitor,TNF inhibitor and two combinations respectively.The combined effects of EGFR inhibitor and TNF inhibitor were detected in GBMs xenotransplantation model of U87 nude mice.Cells viability were detected by MTT.EGFR,p38,ERK and JNK were checked up by Western blot,TNF m RNA was measured by q RT-PCR and TNF? by ELISA respectively.The resistance of U87 cells to EGFR inhibitors was evaluated by si RNA interference.Result:(1)U87 cells expressed EGFR were resistant to EGFR inhibitors.TNF was activated after EGFR inhibition.(2)Inhibition of TNF can enhance the inhibitory effect of glioma cells on EGFR in vitro.(3)Inhibition of TNF can induce the inhibition of EGFR in vivo.(4)In the mechanism,inhibition of TNF can significantly reduce the activity of MAPKs,which is manifested by increased expression of p38,ERK and JNK phosphorylation,result in the the activation of JNK-Axl-ERK signaling pathway induced by TNF was inhibited.Conclusion: The results of this study confirmed that EGFR and TNF inhibitor have synergistic inhibition effect on tumor proliferation and invasion.The results of this study confirmed that EGFR inhibitor combined with TNF inhibitor has potential clinical significance in the treatment of GBMs patients with EGFR positive.Objective: Chemokine is one of superfamily cytokines,chemokine CXCL12 is known as so called stromal cell-derived factor-1(CXCL chemokine 12,CXCL12).CXCR4 is an unique receptor of CXCL12 which highly conserved and G protein coupled.CXCL12-CXCR4 axis plays crucial roles in the migration and invasion of malignant tumors.The expression of CXCL12-CXCR4 increases with the increase of malignant degree of glioma.Molecular targeted therapy for CXCR4 can effectively inhibit the formation of tumor neovascularization and reduce the proliferation and migration of tumor cells.In this study,the crystal structure of CXCL12 with the computer-aided drug design to design a new small molecule CXCR4 inhibitor with the basic functional structure of CXCR4 inhibitors and discard the chemical structure of toxic reactions.The therapeutic effect of the small molecule inhibitor was evaluated by molecular and cellular biology in vitro,and the mechanism of its anti glioma effect was examined at the molecular level.The successful implementation of this project may be expected to provide a new drug for GBM molecular targeted therapy,which has important theoretical significance and clinical practical value.Methods: To develop a novel CXCR4 inhibitor(CPZ1344)for CXCL12 containing pyridinamide scaffold.The inhibitory effect of CPZ1344 on tumor cell U87?U251?SHG44 was evaluated with MTT method,evaluate apoptosis with flow cytometry and TUNEL experiment,cell cycle was detected by flow cytometry,and the ability of tumor angiogenesis with CPZ1344 was evaluated by tubule formation test.The expression of CXCR4,p-Akt and p-m TOR were detected by Western blot.Results:(1)CXCR4 was highly expressed in glioblastoma.(2)CPZ1344 can effectively inhibit the proliferation and invasion of U87 cells and induce apoptosis.(3)CPZ1344 induced G1 phase cell cycle arrest.(4)CPZ1344 blocked part of CXCR4signaling pathway,and the level of CXCR4 was lower with CPZ1344 intervention,and the expression levels of p-Akt and p-m TOR were also significantly reduced.(5)CPZ1344 has strong antiangiogenic activity in vitro,which significantly inhibits the formation of human umbilical vein endothelial cells(HUVCE)tubules.Conclusion: The results of the research confirm that CPZ1344 has significant anti tumor effect on the GBM overexpressed CXCR4.CPZ1344 may be developed into a drug to treat glioma.