Improved transduction of primary murine hepatocytes by recombinant adeno-associated virus 2 vectors in vivo

L. Zhong, W. Li, Z. Yang, L. Chen, Y. Li, K. Qing, K. A. Weigel-Kelley, M. C. Yoder, W. Shou, A. Srivastava

Research output: Contribution to journalArticle

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Abstract

Adeno-associated virus 2 (AAV) vectors are currently in use in Phase I/II clinical trials for gene therapy of cystic fibrosis and hemophilia B. Although 100% of murine hepatocytes can be targeted by AAV vectors, the transgene expression is limited to ∼5% of hepatocytes. Since the viral genome is a single-stranded DNA, and single strands of both polarities are encapsidated with equal frequency, it has been suggested that failure to undergo DNA strand-annealing accounts for the lack of efficient transgene expression. We and others, on the other hand, have proposed that failure to undergo viral second-strand DNA synthesis attributes to the observed low efficiency of transgene expression. We have previously documented that a cellular protein, designated FKBP52, when present in phosphorylated forms, inhibits the viral second-strand DNA synthesis, and consequently, limits transgene expression in nonhepatic cells, whereas unphosphorylated forms of FKBP52 have no effect. To further evaluate whether phosphorylated FKBP52 is also involved in regulating AAV-mediated transgene expression in murine hepatocytes, we generated transgenic mice overexpressing the cellular T-cell protein tyrosine phosphatase (TC-PTP) protein, known to catalyze dephosphorylation of FKBP52, as well as mice deficient in FKBP52. We demonstrate here that dephosphorylation of FKBP52 in TC-PTP transgenic (TC-PTP-TG) mice, and removal of FKBP52 in FKBP52-knockout (FKBP52-KO) mice results in efficient transduction of murine hepatocytes following tail-vein injection of recombinant AAV vectors. We also document efficient viral second-strand DNA synthesis in hepatocytes from both TC-PTP-TG and FKBP52-KO mice. Thus, our data strongly support the contention that the viral second-strand DNA synthesis, rather than DNA strand-annealing, is the rate-limiting step in the efficient transduction of hepatocytes, which should have implications in the optimal use of recombinant AAV vectors in human gene therapy.

Original languageEnglish (US)
Pages (from-to)1165-1169
Number of pages5
JournalGene Therapy
Volume11
Issue number14
DOIs
StatePublished - Jul 1 2004

Fingerprint

Dependovirus
Hepatocytes
Non-Receptor Type 2 Protein Tyrosine Phosphatase
Transgenes
DNA
Knockout Mice
Genetic Therapy
Transgenic Mice
Hemophilia B
tacrolimus binding protein 4
Phase II Clinical Trials
Clinical Trials, Phase I
Viral Genome
Single-Stranded DNA
Cystic Fibrosis
Tail
Veins
Proteins
Injections

Keywords

  • AAV vectors
  • Gene expression
  • Gene transfer
  • Liver cells

ASJC Scopus subject areas

  • Genetics

Cite this

Zhong, L., Li, W., Yang, Z., Chen, L., Li, Y., Qing, K., ... Srivastava, A. (2004). Improved transduction of primary murine hepatocytes by recombinant adeno-associated virus 2 vectors in vivo. Gene Therapy, 11(14), 1165-1169. https://doi.org/10.1038/sj.gt.3302283

Improved transduction of primary murine hepatocytes by recombinant adeno-associated virus 2 vectors in vivo. / Zhong, L.; Li, W.; Yang, Z.; Chen, L.; Li, Y.; Qing, K.; Weigel-Kelley, K. A.; Yoder, M. C.; Shou, W.; Srivastava, A.

In: Gene Therapy, Vol. 11, No. 14, 01.07.2004, p. 1165-1169.

Research output: Contribution to journalArticle

Zhong, L, Li, W, Yang, Z, Chen, L, Li, Y, Qing, K, Weigel-Kelley, KA, Yoder, MC, Shou, W & Srivastava, A 2004, 'Improved transduction of primary murine hepatocytes by recombinant adeno-associated virus 2 vectors in vivo', Gene Therapy, vol. 11, no. 14, pp. 1165-1169. https://doi.org/10.1038/sj.gt.3302283
Zhong, L. ; Li, W. ; Yang, Z. ; Chen, L. ; Li, Y. ; Qing, K. ; Weigel-Kelley, K. A. ; Yoder, M. C. ; Shou, W. ; Srivastava, A. / Improved transduction of primary murine hepatocytes by recombinant adeno-associated virus 2 vectors in vivo. In: Gene Therapy. 2004 ; Vol. 11, No. 14. pp. 1165-1169.
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