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. 1997 Aug;71(8):5932–5941. doi: 10.1128/jvi.71.8.5932-5941.1997

Transduction by adeno-associated virus vectors in the rabbit airway: efficiency, persistence, and readministration.

C L Halbert 1, T A Standaert 1, M L Aitken 1, I E Alexander 1, D W Russell 1, A D Miller 1
PMCID: PMC191849  PMID: 9223483

Abstract

The ability of recombinant adeno-associated virus (AAV) vectors to integrate into the host genome and to transduce nondividing cells makes them attractive as vehicles for gene delivery. In this study, we assessed the ability of several AAV vectors to transduce airway cells in rabbits by measuring marker gene expression. AAV vectors that transferred either a beta-galactosidase (beta-gal) or a human placental alkaline phosphatase (AP) gene were delivered to one lobe of the rabbit lung by use of a balloon catheter placed under fluoroscopic guidance. We observed vector-encoded beta-gal or AP staining almost exclusively in the epithelial and smooth muscle cells in the bronchus at the region of balloon placement. The overall efficiency of transduction in the balloon-treated bronchial epithelium was low but reached 20% in some areas. The majority of the staining was in ciliated cells but was also observed in basal cells and airway smooth muscle cells. We observed an 80-fold decrease in marker-positive epithelial cells during the 60-day period after vector infusion, whereas the number of marker-positive smooth muscle cells stayed constant. Although treatment with the topoisomerase inhibitor etoposide dramatically enhanced AAV transduction in primary airway epithelial cells in culture, treatment of rabbits did not improve transduction rates in the airway. Vector readministration failed to produce additional transduction events, which correlated with the appearance of neutralizing antibodies. These results indicate that both readministration and immune modulation will be required in the use of AAV vectors for gene therapy to the airway epithelium.

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Selected References

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  1. Alexander I. E., Russell D. W., Miller A. D. DNA-damaging agents greatly increase the transduction of nondividing cells by adeno-associated virus vectors. J Virol. 1994 Dec;68(12):8282–8287. doi: 10.1128/jvi.68.12.8282-8287.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Alexander I. E., Russell D. W., Spence A. M., Miller A. D. Effects of gamma irradiation on the transduction of dividing and nondividing cells in brain and muscle of rats by adeno-associated virus vectors. Hum Gene Ther. 1996 May 1;7(7):841–850. doi: 10.1089/hum.1996.7.7-841. [DOI] [PubMed] [Google Scholar]
  3. Alton E. W., Middleton P. G., Caplen N. J., Smith S. N., Steel D. M., Munkonge F. M., Jeffery P. K., Geddes D. M., Hart S. L., Williamson R. Non-invasive liposome-mediated gene delivery can correct the ion transport defect in cystic fibrosis mutant mice. Nat Genet. 1993 Oct;5(2):135–142. doi: 10.1038/ng1093-135. [DOI] [PubMed] [Google Scholar]
  4. Berger J., Howard A. D., Gerber L., Cullen B. R., Udenfriend S. Expression of active, membrane-bound human placental alkaline phosphatase by transfected simian cells. Proc Natl Acad Sci U S A. 1987 Jul;84(14):4885–4889. doi: 10.1073/pnas.84.14.4885. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Boissy R., Astell C. R. An Escherichia coli recBCsbcBrecF host permits the deletion-resistant propagation of plasmid clones containing the 5'-terminal palindrome of minute virus of mice. Gene. 1985;35(1-2):179–185. doi: 10.1016/0378-1119(85)90170-2. [DOI] [PubMed] [Google Scholar]
  6. Clowes M. M., Lynch C. M., Miller A. D., Miller D. G., Osborne W. R., Clowes A. W. Long-term biological response of injured rat carotid artery seeded with smooth muscle cells expressing retrovirally introduced human genes. J Clin Invest. 1994 Feb;93(2):644–651. doi: 10.1172/JCI117016. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Drumm M. L., Pope H. A., Cliff W. H., Rommens J. M., Marvin S. A., Tsui L. C., Collins F. S., Frizzell R. A., Wilson J. M. Correction of the cystic fibrosis defect in vitro by retrovirus-mediated gene transfer. Cell. 1990 Sep 21;62(6):1227–1233. doi: 10.1016/0092-8674(90)90398-x. [DOI] [PubMed] [Google Scholar]
  8. Engelhardt J. F., Allen E. D., Wilson J. M. Reconstitution of tracheal grafts with a genetically modified epithelium. Proc Natl Acad Sci U S A. 1991 Dec 15;88(24):11192–11196. doi: 10.1073/pnas.88.24.11192. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Ferrari F. K., Samulski T., Shenk T., Samulski R. J. Second-strand synthesis is a rate-limiting step for efficient transduction by recombinant adeno-associated virus vectors. J Virol. 1996 May;70(5):3227–3234. doi: 10.1128/jvi.70.5.3227-3234.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Fire A., Harrison S. W., Dixon D. A modular set of lacZ fusion vectors for studying gene expression in Caenorhabditis elegans. Gene. 1990 Sep 14;93(2):189–198. doi: 10.1016/0378-1119(90)90224-f. [DOI] [PubMed] [Google Scholar]
  11. Fisher-Adams G., Wong K. K., Jr, Podsakoff G., Forman S. J., Chatterjee S. Integration of adeno-associated virus vectors in CD34+ human hematopoietic progenitor cells after transduction. Blood. 1996 Jul 15;88(2):492–504. [PubMed] [Google Scholar]
  12. Fisher K. J., Gao G. P., Weitzman M. D., DeMatteo R., Burda J. F., Wilson J. M. Transduction with recombinant adeno-associated virus for gene therapy is limited by leading-strand synthesis. J Virol. 1996 Jan;70(1):520–532. doi: 10.1128/jvi.70.1.520-532.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Flotte T. R., Afione S. A., Conrad C., McGrath S. A., Solow R., Oka H., Zeitlin P. L., Guggino W. B., Carter B. J. Stable in vivo expression of the cystic fibrosis transmembrane conductance regulator with an adeno-associated virus vector. Proc Natl Acad Sci U S A. 1993 Nov 15;90(22):10613–10617. doi: 10.1073/pnas.90.22.10613. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Flotte T. R., Solow R., Owens R. A., Afione S., Zeitlin P. L., Carter B. J. Gene expression from adeno-associated virus vectors in airway epithelial cells. Am J Respir Cell Mol Biol. 1992 Sep;7(3):349–356. doi: 10.1165/ajrcmb/7.3.349. [DOI] [PubMed] [Google Scholar]
  15. Goldman M. J., Wilson J. M. Expression of alpha v beta 5 integrin is necessary for efficient adenovirus-mediated gene transfer in the human airway. J Virol. 1995 Oct;69(10):5951–5958. doi: 10.1128/jvi.69.10.5951-5958.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Goodman S., Xiao X., Donahue R. E., Moulton A., Miller J., Walsh C., Young N. S., Samulski R. J., Nienhuis A. W. Recombinant adeno-associated virus-mediated gene transfer into hematopoietic progenitor cells. Blood. 1994 Sep 1;84(5):1492–1500. [PubMed] [Google Scholar]
  17. Graham F. L., Smiley J., Russell W. C., Nairn R. Characteristics of a human cell line transformed by DNA from human adenovirus type 5. J Gen Virol. 1977 Jul;36(1):59–74. doi: 10.1099/0022-1317-36-1-59. [DOI] [PubMed] [Google Scholar]
  18. Grubb B. R., Pickles R. J., Ye H., Yankaskas J. R., Vick R. N., Engelhardt J. F., Wilson J. M., Johnson L. G., Boucher R. C. Inefficient gene transfer by adenovirus vector to cystic fibrosis airway epithelia of mice and humans. Nature. 1994 Oct 27;371(6500):802–806. doi: 10.1038/371802a0. [DOI] [PubMed] [Google Scholar]
  19. Halbert C. L., Aitken M. L., Miller A. D. Retroviral vectors efficiently transduce basal and secretory airway epithelial cells in vitro resulting in persistent gene expression in organotypic culture. Hum Gene Ther. 1996 Oct 1;7(15):1871–1881. doi: 10.1089/hum.1996.7.15-1871. [DOI] [PubMed] [Google Scholar]
  20. Halbert C. L., Alexander I. E., Wolgamot G. M., Miller A. D. Adeno-associated virus vectors transduce primary cells much less efficiently than immortalized cells. J Virol. 1995 Mar;69(3):1473–1479. doi: 10.1128/jvi.69.3.1473-1479.1995. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Hermonat P. L., Muzyczka N. Use of adeno-associated virus as a mammalian DNA cloning vector: transduction of neomycin resistance into mammalian tissue culture cells. Proc Natl Acad Sci U S A. 1984 Oct;81(20):6466–6470. doi: 10.1073/pnas.81.20.6466. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Hyde S. C., Gill D. R., Higgins C. F., Trezise A. E., MacVinish L. J., Cuthbert A. W., Ratcliff R., Evans M. J., Colledge W. H. Correction of the ion transport defect in cystic fibrosis transgenic mice by gene therapy. Nature. 1993 Mar 18;362(6417):250–255. doi: 10.1038/362250a0. [DOI] [PubMed] [Google Scholar]
  23. Kaplitt M. G., Leone P., Samulski R. J., Xiao X., Pfaff D. W., O'Malley K. L., During M. J. Long-term gene expression and phenotypic correction using adeno-associated virus vectors in the mammalian brain. Nat Genet. 1994 Oct;8(2):148–154. doi: 10.1038/ng1094-148. [DOI] [PubMed] [Google Scholar]
  24. Kauffman S. L. Cell proliferation in the mammalian lung. Int Rev Exp Pathol. 1980;22:131–191. [PubMed] [Google Scholar]
  25. Koeberl D. D., Alexander I. E., Halbert C. L., Russell D. W., Miller A. D. Persistent expression of human clotting factor IX from mouse liver after intravenous injection of adeno-associated virus vectors. Proc Natl Acad Sci U S A. 1997 Feb 18;94(4):1426–1431. doi: 10.1073/pnas.94.4.1426. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. Kotin R. M., Siniscalco M., Samulski R. J., Zhu X. D., Hunter L., Laughlin C. A., McLaughlin S., Muzyczka N., Rocchi M., Berns K. I. Site-specific integration by adeno-associated virus. Proc Natl Acad Sci U S A. 1990 Mar;87(6):2211–2215. doi: 10.1073/pnas.87.6.2211. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Lebkowski J. S., McNally M. M., Okarma T. B., Lerch L. B. Adeno-associated virus: a vector system for efficient introduction and integration of DNA into a variety of mammalian cell types. Mol Cell Biol. 1988 Oct;8(10):3988–3996. doi: 10.1128/mcb.8.10.3988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. McLaughlin S. K., Collis P., Hermonat P. L., Muzyczka N. Adeno-associated virus general transduction vectors: analysis of proviral structures. J Virol. 1988 Jun;62(6):1963–1973. doi: 10.1128/jvi.62.6.1963-1973.1988. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Miller D. G., Adam M. A., Miller A. D. Gene transfer by retrovirus vectors occurs only in cells that are actively replicating at the time of infection. Mol Cell Biol. 1990 Aug;10(8):4239–4242. doi: 10.1128/mcb.10.8.4239. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Mizukami H., Young N. S., Brown K. E. Adeno-associated virus type 2 binds to a 150-kilodalton cell membrane glycoprotein. Virology. 1996 Mar 1;217(1):124–130. doi: 10.1006/viro.1996.0099. [DOI] [PubMed] [Google Scholar]
  31. Muzyczka N. Use of adeno-associated virus as a general transduction vector for mammalian cells. Curr Top Microbiol Immunol. 1992;158:97–129. doi: 10.1007/978-3-642-75608-5_5. [DOI] [PubMed] [Google Scholar]
  32. Plopper C. G., Halsebo J. E., Berger W. J., Sonstegard K. S., Nettesheim P. Distribution of nonciliated bronchiolar epithelial (Clara) cells in intra- and extrapulmonary airways of the rabbit. Exp Lung Res. 1983 Sep;5(2):79–98. doi: 10.3109/01902148309061506. [DOI] [PubMed] [Google Scholar]
  33. Rosenfeld M. A., Yoshimura K., Trapnell B. C., Yoneyama K., Rosenthal E. R., Dalemans W., Fukayama M., Bargon J., Stier L. E., Stratford-Perricaudet L. In vivo transfer of the human cystic fibrosis transmembrane conductance regulator gene to the airway epithelium. Cell. 1992 Jan 10;68(1):143–155. doi: 10.1016/0092-8674(92)90213-v. [DOI] [PubMed] [Google Scholar]
  34. Russell D. W., Alexander I. E., Miller A. D. DNA synthesis and topoisomerase inhibitors increase transduction by adeno-associated virus vectors. Proc Natl Acad Sci U S A. 1995 Jun 6;92(12):5719–5723. doi: 10.1073/pnas.92.12.5719. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Russell D. W., Miller A. D., Alexander I. E. Adeno-associated virus vectors preferentially transduce cells in S phase. Proc Natl Acad Sci U S A. 1994 Sep 13;91(19):8915–8919. doi: 10.1073/pnas.91.19.8915. [DOI] [PMC free article] [PubMed] [Google Scholar]
  36. Samulski R. J., Chang L. S., Shenk T. Helper-free stocks of recombinant adeno-associated viruses: normal integration does not require viral gene expression. J Virol. 1989 Sep;63(9):3822–3828. doi: 10.1128/jvi.63.9.3822-3828.1989. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Takahashi N., Kadota T., Kawano S., Ishikawa K., Kuroyanagi K., Hamajima Y., Ohta K., Ohta S., Kai S., Kohmura H. [Toxicity studies of VP 16-213 (I)--Acute toxicity in mice, rats and rabbits]. J Toxicol Sci. 1986 Apr;11 (Suppl 1):1–16. doi: 10.2131/jts.11.supplementi_1. [DOI] [PubMed] [Google Scholar]
  38. Xiao X., Li J., Samulski R. J. Efficient long-term gene transfer into muscle tissue of immunocompetent mice by adeno-associated virus vector. J Virol. 1996 Nov;70(11):8098–8108. doi: 10.1128/jvi.70.11.8098-8108.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. Yang Y., Trinchieri G., Wilson J. M. Recombinant IL-12 prevents formation of blocking IgA antibodies to recombinant adenovirus and allows repeated gene therapy to mouse lung. Nat Med. 1995 Sep;1(9):890–893. doi: 10.1038/nm0995-890. [DOI] [PubMed] [Google Scholar]
  40. Zabner J., Couture L. A., Gregory R. J., Graham S. M., Smith A. E., Welsh M. J. Adenovirus-mediated gene transfer transiently corrects the chloride transport defect in nasal epithelia of patients with cystic fibrosis. Cell. 1993 Oct 22;75(2):207–216. doi: 10.1016/0092-8674(93)80063-k. [DOI] [PubMed] [Google Scholar]
  41. Zeitlin P. L., Chu S., Conrad C., McVeigh U., Ferguson K., Flotte T. R., Guggino W. B. Alveolar stem cell transduction by an adeno-associated viral vector. Gene Ther. 1995 Nov;2(9):623–631. [PubMed] [Google Scholar]
  42. Zeitlin P. L., Lu L., Rhim J., Cutting G., Stetten G., Kieffer K. A., Craig R., Guggino W. B. A cystic fibrosis bronchial epithelial cell line: immortalization by adeno-12-SV40 infection. Am J Respir Cell Mol Biol. 1991 Apr;4(4):313–319. doi: 10.1165/ajrcmb/4.4.313. [DOI] [PubMed] [Google Scholar]

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