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Cited by 105Virus-based gene therapy researchCystic Fibrosis Research AdvancesRNA Interference and Gene Delivery

A shortened adeno-associated virus expression cassette for CFTR gene transfer to cystic fibrosis airway epithelia

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Ostedgaard, Lynda S., Rokhlina, Tatiana, Karp, Philip H., Lashmit, Philip, Afione, Sandra, Schmidt, Michael, Zabner, Joseph, Stinski, Mark F. +2 more·2005·Proceedings of the National Academy of Sciences
DOI 10.1073/pnas.0409845102PMC PMC549485
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Abstract

Adeno-associated viruses (AAVs) such as AAV5 that transduce airway epithelia from the apical surface are attractive vectors for gene transfer in cystic fibrosis (CF). However, their utility in CF has been limited because packaging of the insert becomes inefficient when its length exceeds ≈4,900–5,000 bp. To partially circumvent this size constraint, we previously developed a CF transmembrane conductance regulator (CFTR) transgene that deleted a portion of the R domain (CFTRΔR). In this study, we focused on shortening the other elements in the AAV expression cassette. We found that portions of the CMV immediate/early ( CMVie ) enhancer/promoter could be deleted without abolishing activity. We also tested various intervening sequences, poly(A) signals, and an intron to develop an expression cassette that meets the size restrictions imposed by AAV. We then packaged these shortened elements with the CFTRΔR transgene into AAV5 and applied them to the apical surface of differentiated CF airway epithelia. Two to 4 weeks later, the AAV5 vectors partially corrected the CF Cl – transport defect. These results demonstrate that a single AAV vector can complement the CF defect in differentiated airway epithelia and thereby further the development of effective CF gene transfer.

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  • Addition of SV40 intron confers about twofold enhancement of AAV transgene expression in lung carcinoma cells.

    “Moreover, addition of SV40 intron confers about twofold enhancement of AAV transgene expression in lung carcinoma cells [25].”

    From Designing and optimizing AAV-mediated gene therapy for neurodegenerative diseases: from bench to bedside (2024) · cites this paper as [25]

    The Results and Discussion report increased β-gal activity with an SV40 intron in a full-length promoter construct and with the 113CMV promoter, but no increase with the 173CMV promoter. The paper does not state an approximately twofold increase, and it says the intron was not included in the AAV expression cassette. Materials and Methods describe the intron tests as plasmid-transfection experiments in cell lines, rather than tests of intron-containing AAV vectors. (LLM, conf=0.88)

    Experimental — accuracy not yet measured

  • A short synthetic polyadenylation signal was successfully used in a previous study.

    “Because a short synthetic polyadenylation signal was successfully used in a previous study [22], we also generated AAV expression cassettes that have a synthetic polyadenylation signal alone (CW3A), or together with a single (CW3SA) or tandem upstream element of the SV40 late polyadenylation signal (CW3SSA).”

    From Optimization of AAV expression cassettes to improve packaging capacity and transgene expression in neurons (2014) · cites this paper as [22]

  • Transduction of airway epithelial cells with an rAAV vector containing a truncated CFTR expression cassette was sufficient to reverse, at least partially, the cystic fibrosis chloride transport defect in these cells.

    “Transduction of airway epithelial cells with an rAAV vector containing this expression cassette was sufficient to reverse, at least partially, the cystic fibrosis chloride transport defect in these cells. 35”

    From Expressing Transgenes That Exceed the Packaging Capacity of Adeno-Associated Virus Capsids (2016) · cites this paper as [35]

  • The CMV promoter can be shortened to 173 bp without apparent loss in expression levels, at least in certain cell types.

    “it has been reported that the CMV promoter can be shortened to 173 bp without apparent loss in expression levels-at least in certain cell types. 35”

    From Expressing Transgenes That Exceed the Packaging Capacity of Adeno-Associated Virus Capsids (2016) · cites this paper as [35]

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Research works printed but not linked: 46

  1. [1]Welsh, M. J., Ramsey, B. W., Accurso, F. & Cutting, G. R. (2001) in The Metabolic and Molecular Basis of Inherited Disease, eds. Scriver, C. R., Beaudet, A. L., Sly, W. S., Valle, D., Childs, B. & Vogelstein, B. (McGraw–Hill, New York), pp. 5121–5189.
  2. [2]Flotte, T. R. (1999) Curr. Opin. Mol. Ther. 1, 510–516. [PubMed] [Google Scholar]
  3. [3]Welsh, M. J. (1999) J. Clin. Invest. 104, 1165–1166. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. [4]Davies, J. C., Geddes, D. M. & Alton, E. W. (2001) J. Gene Med. 3, 409–417. [DOI] [PubMed] [Google Scholar]
  5. [5]Driskell, R. A. & Engelhardt, J. F. (2003) Annu. Rev. Physiol. 65, 585–612. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. [6]High, K. A. (2001) Ann. N.Y. Acad. Sci. 953, 64–74. [DOI] [PubMed] [Google Scholar]
  7. [7]Grimm, D. & Kay, M. A. (2003) Curr. Gene Ther. 3, 281–304. [DOI] [PubMed] [Google Scholar]
  8. [8]Conlon, T. J. & Flotte, T. R. (2004) Expert Opin. Biol. Ther. 4, 1093–1101. [DOI] [PubMed] [Google Scholar]
  9. [9]Halbert, C. L. & Miller, A. D. (2004) Methods Mol. Biol. 246, 201–212. [DOI] [PubMed] [Google Scholar]
  10. [10]McCarty, D. M., Young, S. M. & Samulski, R. J. (2004) Annu. Rev. Genet. 38, 819–845. [DOI] [PubMed] [Google Scholar]
  11. [11]Zabner, J., Seiler, M., Walters, R., Kotin, R. M., Fulgeras, W., Davidson, B. L. & Chiorini, J. A. (2000) J. Virol. 74, 3852–3858. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. [12]Walters, R. W., Yi, S. M., Keshavjee, S., Brown, K. E., Welsh, M. J., Chiorini, J. A. & Zabner, J. (2001) J. Biol. Chem. 276, 20610–20616. [DOI] [PubMed] [Google Scholar]
  13. [13]Di Pasquale, G., Davidson, B. L., Stein, C. S., Martins, I., Scudiero, D., Monks, A. & Chiorini, J. A. (2003) Nat. Med. 9, 1306–1312. [DOI] [PubMed] [Google Scholar]
  14. [14]Dong, J. Y., Fan, P. D. & Frizzell, R. A. (1996) Hum. Gene Ther. 7, 2101–2112. [DOI] [PubMed] [Google Scholar]
  15. [15]Flotte, T. R., Afione, S. A., Solow, R., Drumm, M. L., Markakis, D., Guggino, W. B., Zeitlin, P. L. & Carter, B. J. (1993) J. Biol. Chem. 268, 3781–3790. [PubMed] [Google Scholar]
  16. [16]Zhang, L., Wang, D., Fischer, H., Fan, P. D., Widdicombe, J. H., Kan, Y. W. & Dong, J. Y. (1998) Proc. Natl. Acad. Sci. USA 95, 10158–10163. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. [17]Riordan, J. R., Rommens, J. M., Kerem, B. S., Alon, N., Rozmahel, R., Grzelczak, Z., Zielenski, J., Lok, S., Plavsic, N., Chou, J. L., et al. (1989) Science 245, 1066–1073. [DOI] [PubMed] [Google Scholar]
  18. [18]Wang, D., Fischer, H., Zhang, L., Fan, P., Ding, R. X. & Dong, J. (1999) Gene Ther. 6, 667–675. [DOI] [PubMed] [Google Scholar]
  19. [19]Zhang, L. N., Karp, P., Gerard, C. J., Pastor, E., Laux, D., Munson, K., Yan, Z., Liu, X., Godwin, S., Thomas, C. P., et al. (2004) Mol. Ther. 10, 990–1002. [DOI] [PubMed] [Google Scholar]
  20. [20]Nakai, H., Storm, T. A. & Kay, M. A. (2000) Nat. Biotechnol. 18, 527–532. [DOI] [PubMed] [Google Scholar]
  21. [21]Yan, Z., Zhang, Y., Duan, D. & Engelhardt, J. F. (2000) Proc. Natl. Acad. Sci. USA 97, 6716–6721. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. [22]Halbert, C. L., Allen, J. M. & Miller, A. D. (2002) Nat. Biotechnol. 20, 697–701. [DOI] [PubMed] [Google Scholar]
  23. [23]Pergolizzi, R. G., Ropper, A. E., Dragos, R., Reid, A. C., Nakayama, K., Tan, Y., Ehteshami, J. R., Coleman, S. H., Silver, R. B., Hackett, N. R., et al. (2003) Mol. Ther. 8, 999–1008. [DOI] [PubMed] [Google Scholar]
  24. [24]Ostedgaard, L. S., Zabner, J., Vermeer, D. W., Rokhlina, T., Karp, P. H., Stecenko, A. A., Randak, C. & Welsh, M. J. (2002) Proc. Natl. Acad. Sci. USA 99, 3093–3098. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. [25]Stinski, M. F. & Roehr, T. J. (1985) J. Virol. 55, 431–441. [DOI] [PMC free article] [PubMed] [Google Scholar]
  26. [26]Levitt, N., Briggs, D., Gil, A. & Proudfoot, N. J. (1989) Genes Dev. 3, 1019–1025. [DOI] [PubMed] [Google Scholar]
  27. [27]Chiorini, J. A., Kim, F., Yang, L. & Kotin, R. M. (1999) J. Virol. 73, 1309–1319. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. [28]Chiorini, J. A., Yang, L., Liu, Y., Safer, B. & Kotin, R. M. (1997) J. Virol. 71, 6823–6833. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. [29]Kaludov, N., Brown, K. E., Walters, R. W., Zabner, J. & Chiorini, J. A. (2001) J. Virol. 75, 6884–6893. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. [30]Karp, P. H., Moninger, T., Weber, S. P., Nesselhauf, T. S., Launspach, J., Zabner, J. & Welsh, M. J. (2002) in Epithelial Cell Culture Protocols, ed. Wise, C. (Humana, Totowa, NJ), Vol. 188, pp. 115–137. [DOI] [PubMed] [Google Scholar]
  31. [31]Lee, J. H. & Welsh, M. J. (1999) Gene Ther. 6, 676–682. [DOI] [PubMed] [Google Scholar]
  32. [32]Stinski, M. F. (1999) in Gene Expression Systems: Using Nature for the Art of Expression, eds. Fernandez, J. M. & Holffler, J. P. (Academic, San Diego), pp. 211–233.
  33. [33]Yew, N. S., Wysokenski, D. M., Wang, K. X., Ziegler, R. J., Marshall, J., McNeilly, D., Cherry, M., Osburn, W. & Cheng, S. H. (1997) Hum. Gene Ther. 8, 575–584. [DOI] [PubMed] [Google Scholar]
  34. [34]Jiang, C., O'Connor, S. P., Armentano, D., Berthelette, P. B., Schiavi, S. C., Jefferson, D. M., Smith, A. E., Wadsworth, S. C. & Cheng, S. H. (1996) Am. J. Physiol. 271, L527–L537. [DOI] [PubMed] [Google Scholar]
  35. [35]Zabner, J., Wadsworth, S. C., Smith, A. E. & Welsh, M. J. (1996) Gene Ther. 3, 458–465. [PubMed] [Google Scholar]
  36. [36]Nott, A., Meislin, S. H. & Moore, M. J. (2003) RNA 9, 607–617. [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. [37]Okayama, H. & Berg, P. (1983) Mol. Cell. Biol. 3, 280–289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  38. [38]Kozak, M. (1986) Proc. Natl. Acad. Sci. USA 83, 2850–2854. [DOI] [PMC free article] [PubMed] [Google Scholar]
  39. [39]Konstan, M. W. & Davis, P. B. (2002) Adv. Drug Deliv. Rev. 54, 1409–1423. [DOI] [PubMed] [Google Scholar]
  40. [40]Ritter, T., Brandt, C., Prösch, S., Vergopoulos, A., Vogt, K., Kolls, J. & Volk, H. D. (2000) Cytokine 12, 1163–1170. [DOI] [PubMed] [Google Scholar]
  41. [41]Stein, J., Volk, H. D., Liebenthal, C., Kruger, D. H. & Prosch, S. (1993) J. Gen. Virol. 74, 2333–2338. [DOI] [PubMed] [Google Scholar]
  42. [42]Halbert, C. L., Allen, J. M. & Miller, A. D. (2001) J. Virol. 75, 6615–6624. [DOI] [PMC free article] [PubMed] [Google Scholar]
  43. [43]Gao, G., Alvira, M. R., Somanathan, S., Lu, Y., Vandenberghe, L. H., Rux, J. J., Caldcedo, R., Sanmiguel, J., Abbas, Z. & Wilson, J. M. (2003) Proc. Natl. Acad. Sci. USA 100, 6081–6086. [DOI] [PMC free article] [PubMed] [Google Scholar]
  44. [44]Schmidt, M., Katano, H., Bossis, I. & Chiorini, J. A. (2004) J. Virol. 78, 6509–6516. [DOI] [PMC free article] [PubMed] [Google Scholar]
  45. [45]Trapnell, B. C., Chu, C. S., Paakko, P. K., Banks, T. C., Yoshimura, K., Ferrans, V. J., Chernick, M. S. & Crystal, R. G. (1991) Proc. Natl. Acad. Sci. USA 88, 6565–6569. [DOI] [PMC free article] [PubMed] [Google Scholar]
  46. [46]Chu, C. S., Trapnell, B. C., Curristin, S. M., Cutting, G. R. & Crystal, R. G. (1992) J. Clin. Invest. 90, 785–790. [DOI] [PMC free article] [PubMed] [Google Scholar]

Cited by

Sorted by citations

Authors
Ostedgaard, L. S., Rokhlina, T., Karp, P. H., Lashmit, P., Afione, S., Schmidt, M., Zabner, J., Stinski, M. F., Chiorini, J. A., & Welsh, M. J.
Affiliations
University of Iowa (US)
ORCID
Venue
Proceedings of the National Academy of Sciences
Publisher
National Academy of Sciences
Published
2005-02-09
Volume
102
Issue
8
Pages
2952-2957
ISSN
  • 0027-8424 (ISSN-L)
  • 1091-6490
DOI
10.1073/pnas.0409845102
PMC
PMC549485
OpenAlex
W2087358560
Cited by
105
References
3
Citations by year
2026: 3 · 2025: 8 · 2024: 4 · 2023: 8 · 2022: 1 · 2021: 2 · 2020: 7 · 2019: 8 · 2018: 1 · 2017: 5 · 2016: 3 · 2015: 7 · 2014: 4 · 2013: 1 · 2012: 6
FWCI
3.97
Type
journal article
Language
en
Open access
Locations
  • Proceedings of the National Academy of Sciences (publishedVersion)
  • PubMed Central (publishedVersion)
  • PubMed (publishedVersion)
Funders
  • Howard Hughes Medical Institute
  • Roy J. Carver Charitable Trust
  • Amgen
  • National Heart, Lung, and Blood Institute (HL-51670, P01HL051670)
  • National Institute of Allergy and Infectious Diseases (AI-13562, R01AI013562, R21AI013562)
  • National Institute of Diabetes and Digestive and Kidney Diseases (DK54759, P30DK054759)
  • National Institute of Dental and Craniofacial Research
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Virus-based gene therapy research
  • Health Sciences › Medicine › Pulmonary and Respiratory Medicine › Cystic Fibrosis Research Advances
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › RNA Interference and Gene Delivery
Keywords
cystic fibrosis, adeno-associated virus vectors, AAV5, airway epithelium, gene expression cassette, polyadenylation signal, introns
MeSH terms
Dependovirus, Cells, Cultured, Chlorides, Cystic Fibrosis, Enhancer Elements, Genetic, Epithelium, Genetic Vectors, Humans, Introns, Promoter Regions, Genetic, Trachea, Genetic Therapy, Ion Transport, Cystic Fibrosis Transmembrane Conductance Regulator, Gene Transfer, Horizontal

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