title: Inhibition of hepatitis B virus and induction of hepatoma cell apoptosis by ASGPR-directed delivery of shRNAs.
abstract: Hepatitis B virus (HBV) infection is a worldwide liver disease and nearly 25% of chronic HBV infections terminate in hepatocellular carcinoma (HCC). Currently, there is no effective therapy to inhibit HBV replication and to eliminate hepatoma cells, making it highly desired to develop novel therapies for these two stages of the HBV-caused detrimental disease. Recently, short hairpin RNA (shRNA) has emerged as a potential therapy for virus-infected disease and cancer. Here, we have generated a shRNA, pGenesil-siHBV4, which effectively inhibits HBV replication in the human hepatoma cell line HepG2.2.15. The inhibitory effects of pGenesil-siHBV4 are manifested by the decrease of both the HBV mRNA level and the protein levels of the secreted HBV surface antigen (HBsAg) and HBV e antigen (HBeAg), and by the reduction of secreted HBV DNA. Using mouse hydrodynamic tail vein injection, we demonstrate that pGenesil-siHBV4 is effective in inhibiting HBV replication in vivo. Because survivin plays a key role in cancer cell escape from apoptosis, we further generated pGenesil-siSurvivin, a survivin-silencing shRNA, and showed its effect of triggering apoptosis of HBV-containing hepatoma cells. To develop targeted shRNA therapy, we have identified that as a specific binder of the asialoglycoprotein receptor (ASGPR), jetPEI-Hepatocyte delivers pGenesil-siHBV4 and pGenesil-siSurvivin specifically to hepatocytes, not other types of cells. Finally, co-transfection of pGenesil-siHBV4 and pGenesil-siSurvivin exerts synergistic effects in inducing hepatoma cell apoptosis, a novel approach to eliminate hepatoma by downregulating survivin via multiple mechanisms. The application of these novel shRNAs with the jetPEI-Hepatocyte targeting strategy demonstrates the proof-of-principle for a promising approach to inhibit HBV replication and eliminate hepatoma cells with high specificity.
authors.affiliation: Department of Immunology, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, People's Republic of China.
authors.forename: Jingwei
authors.initials: J
authors.lastname: Ma
authors.forename: Chunmei
authors.initials: C
authors.lastname: Huang
authors.forename: Xinxin
authors.initials: X
authors.lastname: Yao
authors.forename: Chuan
authors.initials: C
authors.lastname: Shi
authors.forename: Lifang
authors.initials: L
authors.lastname: Sun
authors.forename: Lu
authors.initials: L
authors.lastname: Yuan
authors.forename: Ping
authors.initials: P
authors.lastname: Lei
authors.forename: Huifen
authors.initials: H
authors.lastname: Zhu
authors.forename: Hongbo
authors.initials: H
authors.lastname: Liu
authors.forename: Xiongwen
authors.initials: X
authors.lastname: Wu
authors.forename: Qin
authors.initials: Q
authors.lastname: Ning
authors.forename: Chun
authors.initials: C
authors.lastname: Zhou
authors.forename: Guanxin
authors.initials: G
authors.lastname: Shen
mesh_terms: D000818:Animals; D017209:Apoptosis; D037263:Asialoglycoprotein Receptor / Q000235:genetics / Q000378:metabolism*; D006528:Carcinoma, Hepatocellular / Q000209:etiology / Q000235:genetics / Q000628:therapy*; D004279:DNA, Viral / Q000037:antagonists & inhibitors / Q000096:biosynthesis; D015972:Gene Expression Regulation, Neoplastic; D005822:Genetic Vectors; D006514:Hepatitis B Surface Antigens / Q000235:genetics; D006513:Hepatitis B e Antigens / Q000235:genetics; D006515:Hepatitis B virus / Q000235:genetics / Q000254:growth & development*; D019694:Hepatitis B, Chronic / Q000150:complications / Q000235:genetics / Q000628:therapy*; D006801:Humans; D051034:Inhibitor of Apoptosis Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D007275:Injections, Intravenous; D008099:Liver / Q000473:pathology / Q000821:virology; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D058990:Molecular Targeted Therapy; D009928:Organ Specificity; D034741:RNA, Small Interfering / Q000235:genetics / Q000627:therapeutic use*; D012097:Repressor Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D000077022:Survivin; D014162:Transfection
chemical_list: D037263:Asialoglycoprotein Receptor; C117749:Birc5 protein, mouse; D004279:DNA, Viral; D006514:Hepatitis B Surface Antigens; D006513:Hepatitis B e Antigens; D051034:Inhibitor of Apoptosis Proteins; D034741:RNA, Small Interfering; D012097:Repressor Proteins; D000077022:Survivin