The Low Dose Dox Regulates Oncolytic Adenovirus By TGF-?/Smad3 Signaling Pathway To Inhibit Growth In The HCC

Currently,the incidence of liver cancer ranks the fifth in the world of which mortality rate ranks third.About 600,000 people have died each year from liver cancer.Therefore,the study of the treatment of liver cancer has its great significance.TGF-?/Smad3 signaling pathway plays an important role in the development and migration of HCC.TGF-? can promote the formation of liver fibrosis,and the liver fibrosis to liver cancer.What's more,we found that the expression of carcinoembryonic antigen(CEA)in liver cancer cells is regulated by TGF-?/Smad3 signaling pathway.Therefore,to improve the targeting and safety of the oncolytic adenovirus,we have deleted the E1 B 55 kDa gene of the oncolytic adenovirus and using the CEA promoter instead the E1 A promoter.Simultaneously,the oncolytic adenovirus CD55-TRAIL-IETD-MnSOD(CD55-TMn)have been constructed by the oncolytic adenovirus CD55 carrying the TRAIL and MnSOD double genes.Moreover,the using of low dose Dox can promote the oncolytic adenovirus CD55-TMn replication and expression of carrying antitumor gene,which can enhance CD55-TMn of inhibitory effect on the HCC.In this study,CD55-EGFP was used to infect the HCC that had been after treatment of TGF-?/Smad3 signaling pathway p-smad3 inhibitor SIS3,TGF-?receptor type I / II inhibitor LY2109761 and low-dose Dox,and the fluorescence intensity of EGFP protein was detected by fluorescence microscopy and the expression of E1 A protein,smad3 protein and p-smad3 protein was detected by Western Blot.The results showed that low dose of Dox could increase the proliferation and replication of oncolytic adenovirus CD55-TMn and promote the expression of antitumor gene.To detect the safety of the low dose Dox,the migration and invasion of the HCC were detected by Western Blot,QPCR,wound healing assay and cell invasion assay.The results showed that low dose Dox promoted smad3 phosphorylation and up-regulated TGF-?/Smad3 signaling pathway,but had no effect on promoting the migration and invasion of the hepatoma cells.The inhibitory growth effect of the combination treatment of the oncolytic adenovirus CD55-TMn and the low dose Dox in the HCC were detected by MTT assay,Hoechst 33342 staining,Western Blot and animal models.The results showed that the combined treatment more effectively inhibited the growth of liver cancer cells than either single treatment of the oncolytic adenovirus CD55-TMn or the low dose Dox.In summary,the constructed oncolytic adenovirus CD55-TMn with CEA promoter can target the TGF-?/Smad3 signaling pathway involved in the development of HCC and the invasion and migration of HCC,which can more effectively inhibit and kill the tumor under the treatment of low dose Dox.Therefore,CD55-TMn combined with the low dose Dox could not only effectively inhibit and kill the HCC,but also reduce the use of chemotherapeutic drugs,which was expected to become a drug that effectively inhibits liver cancer by targeting the TGF-?/Smad3 signaling pathway.