Studies of Efficacy and Liver Toxicity Related to Adeno-associated Virus-mediated RNA Interference

Overview

Journal Hum Gene Ther

Specialties Genetics
Pharmacology

Date 2013 Jul 9

PMID 23829557

Citations 13

Authors

Cheng-Pu Sun

Tzu-Hui Wu

Chun-Chi Chen

Ping-Yi Wu

Yao-Ming Shih

Koichi Tsuneyama

Mi-Hua Tao

Affiliations

Soon will be listed here.

Abstract

Adeno-associated virus (AAV)-mediated RNA interference shows promise as a therapy for chronic hepatitis B virus (HBV) infection, but its low efficacy and hepatotoxicity pose major challenges. We have generated AAV vectors containing different promoters and a panel of HBV-specific short hairpin RNAs (shRNAs) to investigate factors that contribute to the efficacy and pathogenesis of AAV-mediated RNA interference. HBV transgenic mice injected with high doses of AAV vectors containing the U6 promoter produced abundant shRNAs, transiently inhibited HBV, but induced severe hepatotoxicity. Sustained HBV suppression without liver toxicity can be achieved by lowering the dose of AAV-U6 vectors. AAVs containing the weaker H1 promoter did not cause liver injury, but their therapeutic efficacy was highly dependent on the sequence of the shRNA. Mice treated with the toxic U6-promoter-driven shRNA showed little change in hepatic microRNA levels, but a dramatic increase in hepatic leukocytes and inflammatory cytokines and chemokines. Hepatotoxicity was completely absent in immunodeficient mice and significantly alleviated in wild-type mice depleted of macrophages and granulocytes, suggesting that host inflammatory responses are the major cause of liver injury induced by the overexpressed shRNAs from AAV-U6 vectors. Our results demonstrate that selection of a highly potent shRNA and control its expression level is critical to achieve sustained HBV suppression without inducing inflammatory side effects.

Citing Articles

Engineering viral vectors for acoustically targeted gene delivery.

Li H, Harb M, Heath J, Trippett J, Shapiro M, Szablowski J Nat Commun. 2024; 15(1):4924.

PMID: 38858354 PMC: 11164914. DOI: 10.1038/s41467-024-48974-y.

An RNA Interference/Adeno-Associated Virus Vector-Based Combinatorial Gene Therapy Approach Against Hepatitis E Virus.

Zhang C, Freistaedter A, Schmelas C, Gunkel M, Dao Thi V, Grimm D Hepatol Commun. 2021; 6(4):878-888.

PMID: 34719133 PMC: 8948557. DOI: 10.1002/hep4.1842.

Unresolved Issues in RNA Therapeutics in Vascular Diseases With a Focus on Aneurysm Disease.

Schellinger I, Dannert A, Mattern K, Raaz U, Tsao P Front Cardiovasc Med. 2021; 8:571076.

PMID: 33937351 PMC: 8081859. DOI: 10.3389/fcvm.2021.571076.

Efficacy, accumulation, and transcriptional profile of anti-HIV shRNAs expressed from human U6, 7SK, and H1 promoters.

Goguen R, Del Corpo O, Malard C, Daher A, Alpuche-Lazcano S, Chen M Mol Ther Nucleic Acids. 2021; 23:1020-1034.

PMID: 33614248 PMC: 7868930. DOI: 10.1016/j.omtn.2020.12.022.

Adipose Tissue: An Emerging Target for Adeno-associated Viral Vectors.

Bates R, Huang W, Cao L Mol Ther Methods Clin Dev. 2020; 19:236-249.

PMID: 33102616 PMC: 7566077. DOI: 10.1016/j.omtm.2020.09.009.

References

Chang C, Lo C, Shih Y, Chen Y, Wu P, Tsuneyama K . Treatment of hepatocellular carcinoma with adeno-associated virus encoding interleukin-15 superagonist. Hum Gene Ther. 2010; 21(5):611-21. DOI: 10.1089/hum.2009.187. View

Morrissey D, Blanchard K, Shaw L, Jensen K, Lockridge J, Dickinson B . Activity of stabilized short interfering RNA in a mouse model of hepatitis B virus replication. Hepatology. 2005; 41(6):1349-56. DOI: 10.1002/hep.20702. View

Klein C, Bock C, Wedemeyer H, Wustefeld T, Locarnini S, Dienes H . Inhibition of hepatitis B virus replication in vivo by nucleoside analogues and siRNA. Gastroenterology. 2003; 125(1):9-18. DOI: 10.1016/s0016-5085(03)00720-0. View

Chen C, Yang H, Su J, Jen C, You S, Lu S . Risk of hepatocellular carcinoma across a biological gradient of serum hepatitis B virus DNA level. JAMA. 2006; 295(1):65-73. DOI: 10.1001/jama.295.1.65. View

An D, Qin F, Auyeung V, Mao S, Kung S, Baltimore D . Optimization and functional effects of stable short hairpin RNA expression in primary human lymphocytes via lentiviral vectors. Mol Ther. 2006; 14(4):494-504. PMC: 2562632. DOI: 10.1016/j.ymthe.2006.05.015. View

Grimm D, Streetz K, Jopling C, Storm T, Pandey K, Davis C . Fatality in mice due to oversaturation of cellular microRNA/short hairpin RNA pathways. Nature. 2006; 441(7092):537-41. DOI: 10.1038/nature04791. View

Ehlert E, Eggers R, Niclou S, Verhaagen J . Cellular toxicity following application of adeno-associated viral vector-mediated RNA interference in the nervous system. BMC Neurosci. 2010; 11:20. PMC: 2841193. DOI: 10.1186/1471-2202-11-20. View

van der Laan L, Wang Y, Tilanus H, Janssen H, Pan Q . AAV-mediated gene therapy for liver diseases: the prime candidate for clinical application?. Expert Opin Biol Ther. 2011; 11(3):315-27. DOI: 10.1517/14712598.2011.548799. View

Giladi H, Ketzinel-Gilad M, Rivkin L, Felig Y, Nussbaum O, Galun E . Small interfering RNA inhibits hepatitis B virus replication in mice. Mol Ther. 2003; 8(5):769-76. DOI: 10.1016/s1525-0016(03)00244-2. View

10.

Iloeje U, Yang H, Su J, Jen C, You S, Chen C . Predicting cirrhosis risk based on the level of circulating hepatitis B viral load. Gastroenterology. 2006; 130(3):678-86. DOI: 10.1053/j.gastro.2005.11.016. View

11.

Couto L, High K . Viral vector-mediated RNA interference. Curr Opin Pharmacol. 2010; 10(5):534-42. DOI: 10.1016/j.coph.2010.06.007. View

12.

Gu S, Jin L, Zhang F, Huang Y, Grimm D, Rossi J . Thermodynamic stability of small hairpin RNAs highly influences the loading process of different mammalian Argonautes. Proc Natl Acad Sci U S A. 2011; 108(22):9208-13. PMC: 3107324. DOI: 10.1073/pnas.1018023108. View

13.

Chen C, Chang C, Sun C, Yu C, Wu P, Jeng K . Use of RNA interference to modulate liver adenoma development in a murine model transgenic for hepatitis B virus. Gene Ther. 2011; 19(1):25-33. DOI: 10.1038/gt.2011.60. View

14.

Chen C, Ko T, Ma H, Wu H, Xiao X, Li J . Long-term inhibition of hepatitis B virus in transgenic mice by double-stranded adeno-associated virus 8-delivered short hairpin RNA. Gene Ther. 2006; 14(1):11-9. DOI: 10.1038/sj.gt.3302846. View

15.

McBride J, Boudreau R, Harper S, Staber P, Monteys A, Martins I . Artificial miRNAs mitigate shRNA-mediated toxicity in the brain: implications for the therapeutic development of RNAi. Proc Natl Acad Sci U S A. 2008; 105(15):5868-73. PMC: 2311380. DOI: 10.1073/pnas.0801775105. View

16.

Kwon H, Lok A . Hepatitis B therapy. Nat Rev Gastroenterol Hepatol. 2011; 8(5):275-84. DOI: 10.1038/nrgastro.2011.33. View

17.

Shlomai A, Shaul Y . Inhibition of hepatitis B virus expression and replication by RNA interference. Hepatology. 2003; 37(4):764-70. DOI: 10.1053/jhep.2003.50146. View

18.

Shultz L, Lyons B, Burzenski L, Gott B, Chen X, Chaleff S . Human lymphoid and myeloid cell development in NOD/LtSz-scid IL2R gamma null mice engrafted with mobilized human hemopoietic stem cells. J Immunol. 2005; 174(10):6477-89. DOI: 10.4049/jimmunol.174.10.6477. View

19.

Morrissey D, Lockridge J, Shaw L, Blanchard K, Jensen K, Breen W . Potent and persistent in vivo anti-HBV activity of chemically modified siRNAs. Nat Biotechnol. 2005; 23(8):1002-7. DOI: 10.1038/nbt1122. View

20.

Khodr C, Sapru M, Pedapati J, Han Y, West N, Kells A . An α-synuclein AAV gene silencing vector ameliorates a behavioral deficit in a rat model of Parkinson's disease, but displays toxicity in dopamine neurons. Brain Res. 2011; 1395:94-107. PMC: 3105182. DOI: 10.1016/j.brainres.2011.04.036. View