title: Gene therapy targeting hepatocellular carcinoma by a dual-regulated oncolytic adenovirus harboring the focal adhesion kinase shRNA.
abstract: Cancer targeting gene-viro-therapy (CTGVT) approach has become a hotspot and a trend in the field of cancer biotherapy and oncolytic adenovirus is an ideal vector to carry the targeting genes. In this study, we used human telomerase reverse transcriptase (hTERT) promoter to control the adenovirus early region 1a (E1A) and the human α-fetoprotein (AFP) promoter integrated with hypoxia response element (HRE) to control the adenovirus early region 1b (E1B). Then the novel double-regulated adenovirus Ad-hTERT-HREAF (named SG505) was engineered. The short-hairpin RNA against focal adhesion kinase (FAK) was inserted into SG505 and thus forming Ad-hTERT-HREAF-shRNA (called SG505‑siFAK). Then various oncolytic adenoviruses were examined to verify whether they could suppress liver cancer cells selectively and efficiently both in vitro and in vivo. Both replicative and replication-defective adenoviruses carrying FAK-shRNA significantly inhibited the expression of FAK in Hep3B and SMMC-7721 cell lines and efficiently suppressed the growth of liver cancer cell lines with minor effect to normal cells. Furthermore, the recombined oncolytic adenoviruses, SG505-siFAK, SG505-EGFP and SG505 were able to selectively propagate in AFP-positive liver cancer cells in vitro and the SG505-siFAK efficiently suppressed the expression of FAK. SG505-siFAK showed the most potent tumor inhibition capability among the three recombined adenovirus with IC50 levels of 0.092±0.009 and 0.424±0.414 pfu/cell in the Hep3B and HepG2 cell line, respectively. Animal experiment further confirmed that SG505-siFAK achieved the most significant tumor inhibition of Hep3B liver cancer xenografted growth by intratumoral injection comparing to the intravenous injection among the three recombined viruses. Immunohistochemical results indicated that FAK expression was downregulated significantly in the tumors treated with SG505-siFAK. The dual-regulated oncolytic adenovirus SG505-siFAK was proven to inhibit the growth of liver cancer cells selectively and efficiently, therefore SG505-siFAK could be a potential agent for future clinical trials of hepatocellular carcinoma.
authors.affiliation: Department of General Surgery, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai 200233, P.R. China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Gao
authors.affiliation: Department of General Surgery, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai 200233, P.R. China.
authors.forename: Yayun
authors.initials: Y
authors.lastname: Zhu
authors.affiliation: Department of General Surgery, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai 200233, P.R. China.
authors.forename: Xinyu
authors.initials: X
authors.lastname: Huang
authors.affiliation: Department of General Surgery, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai 200233, P.R. China.
authors.forename: Kaixing
authors.initials: K
authors.lastname: Ai
authors.affiliation: Department of General Surgery, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai 200233, P.R. China.
authors.forename: Qi
authors.initials: Q
authors.lastname: Zheng
authors.affiliation: Department of General Surgery, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai 200233, P.R. China.
authors.forename: Zhou
authors.initials: Z
authors.lastname: Yuan
mesh_terms: D000256:Adenoviridae / Q000235:genetics* / Q000502:physiology; D017498:Adenovirus E1 Proteins / Q000235:genetics / Q000378:metabolism; D000818:Animals; D006528:Carcinoma, Hepatocellular / Q000378:metabolism / Q000628:therapy*; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D051417:Focal Adhesion Kinase 1 / Q000378:metabolism*; D005822:Genetic Vectors / Q000008:administration & dosage* / Q000494:pharmacology; D057809:HEK293 Cells; D056945:Hep G2 Cells; D006801:Humans; D015552:Injections, Intralesional; D007275:Injections, Intravenous; D008113:Liver Neoplasms / Q000378:metabolism / Q000628:therapy*; D051379:Mice; D050130:Oncolytic Virotherapy / Q000379:methods; D050504:Oncolytic Viruses / Q000235:genetics / Q000502:physiology; D000067492:RNAi Therapeutics / Q000379:methods*; D011994:Recombinant Proteins / Q000235:genetics / Q000494:pharmacology; D019098:Telomerase / Q000235:genetics; D023041:Xenograft Model Antitumor Assays
chemical_list: D017498:Adenovirus E1 Proteins; D011994:Recombinant Proteins; D051417:Focal Adhesion Kinase 1; C496050:PTK2 protein, human; C509186:TERT protein, human; D019098:Telomerase