The Effect Of FNC On The Proliferation And Metastasis In Non-small Cell Lung Cancer H460 Cell

Background and Objective:Lung cancer is the most common malignancy with the highest morbidity and mortality in the world.The latest data shows that lung cancer accounts for about13%of all new cancer cases and 26%of death cases related cancer.Non-small cell lung cancer?NSCLC?is the main type of lung cancer,accounting for about 80-85%of all lung cancer cases.Chemotherapy is the main method for the treatment of patients with non-small cell lung cancer.However,two-thirds of NSCLC patients have entered the advanced stage at the time of diagnosis,although great progress has been made in the treatment strategy,the prognosis of NSCLC patients is still poor,with the 5-year survival rate less than 15%.Therefore,it is urgent to find an anti-non-small cell lung cancer drug with better efficacy and little side effect.FNC?2'-deoxy-2'-?-fluoro-4'azide-nucleoside?,a novel nucleoside analogue containing fluorine,which was designed and synthesized by the School of Chemistry and Molecular Engineering of Zhengzhou university,and had been applied for the national invention patent,the patent number is ZL201010506595.X.Previous experimental results indicated that FNC had potential therapeutic effect on a variety of tumor cells,and had obvious proliferation inhibition effect on non-small cell lung cancer cell line H460,IC₅₀ value is 0.64?M.In this study,human non-small cell lung cancer cells H460 was selected to study the effect of FNC on the proliferation and metastasis in non-small cell lung cancer cells through in vivo and in vitro experiments.Methods:1.The H460 cells with scratch were treated with 0.00,0.04,0.16,0.64?mol·l^-1 FNC,the cell migration area was measured and photographed with an inverted microscope at 0 h,24 h,48 h,72 h.2.H460 cells were treated with 0.00,0.04,0.16,0.64?mol·l^-1 FNC,the effect of FNC on the invasiveness of non-small cell lung cancer H460 cells was detected by Transwell assay.3.H460 cells were treated with 0.00,0.04,0.16,0.64?mol·l^-11 FNC for 48h,the mRNA expression of DNMT1,DNMT3A,DNMT3B and RAR?were detected by Reverse Transcription-Polymerase Chain Reaction?RT-PCR?.4.H460 cells were treated with 0.00,0.04,0.16,0.64?mol·l^-1 FNC for 48h,the expression of E-cadherin,VEGF,MMP-2,MMP-9 protein were detected by Western Blot.5.H460 cells were injected subcutaneously into the right dorsal flank of BALB/c nu/nu mice.mice were then divided into five groups?n=6?according to the mean tumor volume.FNC were given at 2,4,8 mg/kg once a day.Gemcitabine was used as a positive control drug three days a time,negative control group was given 0.9%normal saline once a day.After 11 days,mice was euthanized,tumors were excised,weighed,photographed and fixed in 4%polyoxymethylene,then the expression of CD31 was analyzed by immunohistochemistry.Results:1.H460 cells were treated with FNC for 24 h,wound scratch healing area between each group was no significant difference;H460 cells were treated with FNC for 48 h,72 h,the wound healing area decreased significantly,with dose-response manner following FNC treatment,FNC significantly inhibited the migration of H460 cells.2.FNC significantly inhibited the invasion of H460 cells,with dose-response manner following FNC treatment.3.The mRNA expression level of DNMT1 and DNMT3A were no significant difference;the mRNA expression level of DNMT3B in H460 cells significantly reduced and expression of RAR?significantly increased.4.The expression of E-cadherin protein increased,and the expression of VEGF,MMP-2 and MMP-9 protein significantly decreased.5.FNC significantly inhibited the growth of tumor xenografts in nude mice.When compared with untreated control mice after 11 days treatment,anti-tumor rates for Gemcitabine?30 mg/kg?,FNC?2 mg/kg?,FNC?4 mg/kg?,and FNC?8 mg/kg?were62.44%,27.02%,61.80%,and 76.31%,respectively.Meanwhile FNC treatment significantly reduced the expression of CD31.Conclusions:1.FNC can significantly inhibit the proliferation of NSCLC cells in tumor-bearing mice.2.FNC can inhibit tumor angiogenesis and tumor cell invasion and migration.3.FNC may play ananti-tumor effect in non-small cell lung cancer H460 cell by acting on multiple targets.4.Inhibition of DNMT3B expression is one of the important mechanisms of FNC inhibiting proliferation and metastasis of NSCLC.