Effects Of Podophyllic Acid[4-[2,2,6,6-tetramethylpiperdinooxyl]]Hydrozide On Potential Metastasis Of Tca8113 Cells In Vitro

Objective:Podophyllic acid[4-[2,2,6,6-tetrame thylpiperdinooxyl]]hydrozide(GP-1),one of spin-labeled derivatives of podophyllotoxin,which have more effective antitumor activity and lower toxicity than podophyllotoxin.It has been reported that GP-1 could significantly inhibit the growth of leukemia L1210 cells in vitro and the mouse implanted S-180 and HepA tumors in vivo,and also could induce SGC-7901 cells apoptosis.However,the effect of GP-1 on Tca8113 cells’proliferation and metastasis has not been reported.In this study,we investigated the effects of GP1 on the potential metastasis of Tca8113 cells in vitro.Methods:The viability of Tca8113,A549,HepG2,BCG823 and EJ cells was assessed using Sulforhodamine B(SRB)assay.Tca8113 cells cycle was tested by flow cytometry(FCM).Apoptosis of Tca8113 cells was determined by Fluorescence microscope with DAPI staining.The abilities of Tca8]13 cells migration and invasion were evaluated by Transwell chamber migration assay.Adhesion abilities of Tca8113 cells were done through cell-matrix adhesion assay.TIMP-2,PAI-1 and PTEN mRNA level were determined by RT-PCR.The protein expression levels of Akt,p-Akt,IKK and P-IKK were determined using Western blotting analysis.Results:GP-1 significantly inhibited the proliferation of five tumor cell lines in vitro,and the inhibition efficiency of GP-1 on Tca8113 cells was most potent.GP-1(0.1～0.4 ug/mL)obviously arrested Tca8113 cells in S phase,and a fraction of cells with hypodiploid DNA content representing apoptosis was detected using FCM.Further,GP-1 markedly induced Tca8113 cells apoptosis,and the apoptosis rate was 68.1%after 48 h treatment with GP-1 0.4 ug/mL.GP-1 also inhibited Tca8113 cells’ adhesion,migration and invasion,and the inhibitory rate was 73.58%,50.59%and 52.18%,respectively after 24 h treatment with GP-1 0.4 ug/mL.After Tca8113 cells were treated with GP-1 for 24 h,the TIMP-2 and PAI-1 mRNA levels were up-regulated and the protein expressions of p-Akt,p-IKK were decreased.Conclusion:GP-1 could inhibit the proliferation of five tumor cell lines in vitro,the inhibition efficiency of GP-1 on Tca8113 cells was most potent.GP-1 induced cell apoptosis and arrested Tca8113 cells in S phase,GP-1 has anti-metastasis potential of Tca8113 cells in vitro,and its mechanism may be related to up-regulation of TIMP-2 and PAI-1 mRNA expression and reduction of p-Akt and p-IKK protein expression.