Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Research Article
  • Published:

Genetic manipulations of adenovirus type 5 fiber resulting in liver tropism attenuation

Gene Therapy volume 10, pages 153–162 (2003)Cite this article

Abstract

The development of genetically modified adenoviral vectors capable of specifically transducing a given cell population requires the addition and functional presentation of particular tropism determinants within the virus capsid, together with the abrogation of the molecular determinants that dictate their natural tropism in vivo. The human adenovirus serotype 5 (Ad5) first attaches to the cell surface following high-affinity binding of the C-terminal knob of the fiber capsid protein to the coxsackie and adenovirus receptor (CAR). Here we have assessed whether genetic shortening of the fiber shaft (virus BS1), or replacing the Ad5 fiber shaft and knob with their Ad3 counterparts (virus DB6), could cripple this interaction in vitro and in vivo. A 10-fold decrease in the binding of the modified capsids to soluble CAR was evidenced, which correlated with a similar reduction of their ability to transduce CAR-positive cells in vitro. The ability of BS1 to interact with cellular integrins was also impaired, suggesting that the penton base and the short-shafted fiber when embedded in the capsid preclude each other from efficiently interacting with their cognate cell surface receptors (CAR and integrins respectively). BS1 and DB6 intravenous injections in mice further supported a profound impairment of the ability of the capsid-modified viruses to transduce the liver as demonstrated by a 10-fold reduction of intracellular viral DNA and transgene expression. Interestingly enough, the host humoral response was also specifically weakened in BS1- and DB6-inoculated animals. Taken together, these observations indicate that (i) fiber shortening and (ii) pseudo-typing of Ad5-based vectors with the shaft and knob from non-CAR-binding serotypes constitute two promising strategies to successfully attenuate their native tropism in vitro and most importantly in vivo.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5

Similar content being viewed by others

References

  1. Wickham TJ . Targeting adenovirus. Gene Ther 2000; 7: 110–114.

    Article  CAS  PubMed  Google Scholar 

  2. Borgland SL et al. Adenovirus vector-induced expression of the C-X-C chemokine IP-10 is mediated through capsid-dependent activation of NF-kappaB. J Virol 2000; 74: 3941–3497.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Bruder JT, Kovesdi I . Adenovirus infection stimulates the Raf/MAPK signaling pathway and induces interleukin-8 expression. J Virol 1997; 71: 398–404.

    CAS  PubMed  PubMed Central  Google Scholar 

  4. Muruve DA, Barnes MJ, Stillman IE, Libermann TA . Adenoviral gene therapy leads to rapid induction of multiple chemokines and acute neutrophil-dependent hepatic injury in vivo. Hum Gene Ther 1999; 10: 965–976.

    Article  CAS  PubMed  Google Scholar 

  5. Inoue S et al. Transfer of heme oxygenase 1 cDNA by a replication-deficient adenovirus enhances interleukin 10 production from alveolar macrophages that attenuates lipopolysaccharide-induced acute lung injury in mice. Hum Gene Ther 2001; 12: 967–979.

    Article  CAS  PubMed  Google Scholar 

  6. Tibbles LA et al. Activation of p38 and ERK signaling during adenovirus vector cell entry lead to expression of the C-X-C chemokine IP-10. J Virol 2002; 76: 1559–1568.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Higginbotham JN, Seth P, Blaese RM, Ramsey WJ . The release of inflammatory cytokines from human peripheral blood mononuclear cells in vitro following exposure to adenovirus variants and capsid. Hum Gene Ther 2002; 13: 129–141.

    Article  CAS  PubMed  Google Scholar 

  8. Jooss K, Yang Y, Fisher KJ, Wilson JM . Transduction of dendritic cells by DNA Viral vectors directs the immune response to transgene products in muscle fibers. J Virol 1998; 72: 4212–4277.

    CAS  PubMed  PubMed Central  Google Scholar 

  9. Fechner H et al. Expression of coxsackie adenovirus receptor and alphav-integrin does not correlate with adenovector targeting in vivo indicating anatomical vector barriers. Gene Ther 1999; 6: 1520–1535.

    Article  CAS  PubMed  Google Scholar 

  10. Herz J, Gerard RD . Adenovirus-mediated transfer of low density lipoprotein receptor gene acutely accelerates cholesterol clearance in normal mice. Proc Natl Acad Sci USA 1993; 90: 2812–2816.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  11. Kass-Eisler A et al. The impact of developmental stage, route of administration and the immune system on adenovirus-mediated gene transfer. Gene Ther 1994; 1: 395–402.

    CAS  PubMed  Google Scholar 

  12. Huard J et al. The route of administration is a major determinant of the transduction efficiency of rat tissues by adenoviral recombinants. Gene Ther 1995; 2: 107–115.

    CAS  PubMed  Google Scholar 

  13. Bergelson JM et al. The murine CAR homolog is a receptor for coxsackie B viruses and adenoviruses. J Virol 1998; 72: 415–419.

    CAS  PubMed  PubMed Central  Google Scholar 

  14. Tomko RP, Xu R, Philipson L . HCAR and MCAR: The human and mouse cellular receptors for subgroup C adenoviruses and group B coxsackieviruses. Proc Natl Acad Sci USA 1997; 94: 3352–3356.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Wickham TJ, Mathias P, Cheresh DA, Nemerow GR . Integrins alpha v beta 3 and alpha v beta 5 promote adenovirus internalization but not virus attachment. Cell 1993; 73: 309–319.

    Article  CAS  PubMed  Google Scholar 

  16. Wickham TJ, Filardo EJ, Cheresh DA, Nemerow GR . Integrin alpha v beta 5 selectively promotes adenovirus mediated cell membrane permeabilization. J Cell Biol 1994; 127: 257–264.

    Article  CAS  PubMed  Google Scholar 

  17. Rux JJ, Burnett RM . Type-specific epitope locations revealed by X-ray crystallographic study of adenovirus type 5 hexon. Mol Ther 2000; 1: 18–30.

    Article  CAS  PubMed  Google Scholar 

  18. Bewley MC et al. Structural analysis of the mechanism of adenovirus binding to its human cellular receptor, CAR. Science 1999; 286: 1579–1583.

    Article  CAS  PubMed  Google Scholar 

  19. Roelvink PW et al. Identification of a conserved receptor-binding site on the fiber proteins of CAR-recognizing adenoviridae. Science 1999; 286: 1568–1571.

    Article  CAS  PubMed  Google Scholar 

  20. van Raaij MJ, Louis N, Chroboczek J, Cusack S . Structure of the human adenovirus serotype 2 fiber head domain at 1.5 A resolution. Virology 1999; 262: 333–343.

    Article  CAS  PubMed  Google Scholar 

  21. Durmort C et al. Structure of the fiber head of ad3, a non-car-binding serotype of adenovirus. Virology 2001; 285: 302–312.

    Article  CAS  PubMed  Google Scholar 

  22. Chiu CY, Mathias P, Nemerow GR, Stewart PL . Structure of adenovirus complexed with its internalization receptor alpha vbeta 5 integrin. J Virol 1999; 73: 6759–6768.

    CAS  PubMed  PubMed Central  Google Scholar 

  23. Wickham T et al. Increased in vitro and in vivo gene transfer by adenovirus vectors containing chimeric fiber proteins. J Virol 1997; 71: 8221–8229.

    CAS  PubMed  PubMed Central  Google Scholar 

  24. Vigne E et al. RGD inclusion in the hexon monomer provides adenovirus type 5-based vectors with a fiber knob-independent pathway for infection. J Virol 1999; 73: 5156–5161.

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Krasnykh V et al. Characterization of an adenovirus vector containing a heterologous peptide epitope in the HI loop of the fiber knob. J Virol 1998; 72: 1844–1852.

    CAS  PubMed  PubMed Central  Google Scholar 

  26. Dmitriev I et al. An adenovirus vector with genetically modified fibers demonstrates expanded tropism via utilization of a coxsackievirus and adenovirus receptor-independent cell entry mechanism. J Virol 1998; 72: 9706–9713.

    CAS  PubMed  PubMed Central  Google Scholar 

  27. Xia H, Anderson B, Mao Q, Davidson BL . Recombinant human adenovirus: targeting to the human transferrin receptor improves gene transfer to brain microcapillary endothelium. J Virol 2000; 74: 11359–11366.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  28. Legrand V et al. Fiberless recombinant adenoviruses: virus maturation and infectivity in the absence of fiber. J Virol 1999; 73: 907–919.

    CAS  PubMed  PubMed Central  Google Scholar 

  29. Shayakhmetov DM, Lieber A . Dependence of adenovirus infectivity on length of the fiber shaft domain. J Virol 2000; 74: 10 274–10 286.

    Article  Google Scholar 

  30. Stevenson S, Rollence M, Marshall-Neff J, McClelland A . Selective targeting of human cells by a chimeric adenovirus vector containing a modified fiber protein. J Virol 1997; 71: 4782–4790.

    CAS  PubMed  PubMed Central  Google Scholar 

  31. Von Seggern DJ et al. A helper-independent adenovirus vector with E1, E3, and fiber deleted: structure and infectivity of fiberless particles. J Virol 1999; 73: 1601–1608.

    CAS  PubMed  PubMed Central  Google Scholar 

  32. Gall J, Kass-Eisler A, Leinwand L, Falck-Pedersen E . Adenovirus type 5 and 7 capsid chimera: fiber replacement alters receptor tropism without affecting primary immune neutralization epitopes. J Virol 1996; 70: 2116–2123.

    CAS  PubMed  PubMed Central  Google Scholar 

  33. Jakubczak JL et al. Adenovirus type 5 viral particles pseudotyped with mutagenized fiber proteins show diminished infectivity of coxsackie B–adenovirus receptor-bearing cells. J Virol 2001; 75: 2972–2981.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Leissner P et al. Influence of adenoviral fiber mutations on viral encapsidation, infectivity and in vivo tropism. Gene Ther 2001; 8: 49–57.

    Article  CAS  PubMed  Google Scholar 

  35. Alemany R, Curiel DT . CAR-binding ablation does not change biodistribution and toxicity of adenoviral vectors. Gene Ther 2001; 8: 1347–1353.

    Article  CAS  PubMed  Google Scholar 

  36. Einfeld DA et al. Reducing the native tropism of adenovirus vectors requires removal of both CAR and integrin interactions. J Virol 2001; 75: 11 284–11 291.

    Article  Google Scholar 

  37. van Beusechem VW et al. Recombinant adenovirus vectors with knobless fibers for targeted gene transfer. Gene Ther 2000; 7: 1940–1946.

    Article  CAS  PubMed  Google Scholar 

  38. Krasnykh V et al. Genetic targeting of an adenovirus vector via replacement of the fiber protein with the phage T4 fibritin. J Virol 2001; 75: 4176–4183.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Magnusson MK, Hong SS, Boulanger P, Lindholm L . Genetic retargeting of adenovirus: novel strategy employing "deknobbing" of the fiber. J Virol 2001; 75: 7280–7289.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  40. Roelvink PW et al. The Coxsackievirus–adenovirus receptor protein can function as a cellular attachment protein for adenovirus serotypes from subgroups A, C, D, E, and F. J Virol 1998; 72: 7909–7915.

    CAS  PubMed  PubMed Central  Google Scholar 

  41. Chroboczek J, Ruigrok RW, Cusack S . Adenovirus fiber. In: Doerfler W, Böhm P (eds). Current Topics of Microbiology Immunology. Berlin: Springer-Verlag, 1995; pp. 163–200.

    Google Scholar 

  42. Roelvink P, Kovesdi I, Wickham T . Comparative analysis of adenovirus fiber–cell interaction: adenovirus type 2 (Ad2) and Ad9 utilize the same cellular fiber receptor but use different binding strategies for attachment. J Virol 1996; 70: 7614–7621.

    CAS  PubMed  PubMed Central  Google Scholar 

  43. Zabner J et al. A chimeric type 2 adenovirus vector with a type 17 fiber enhances gene transfer to human airway epithelia. J Virol 1999; 73: 8689–8695.

    CAS  PubMed  PubMed Central  Google Scholar 

  44. Kirby I et al. Adenovirus type 9 fiber knob binds to the Coxsackie B virus-adenovirus receptor (CAR) with lower affinity than fiber knobs of other CAR-binding adenovirus serotypes. J Virol 2001; 75: 7210–7214.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  45. van Raaij MJ, Mitraki A, Lavigne G, Cusack S . A triple beta-spiral in the adenovirus fibre shaft reveals a new structural motif for a fibrous protein. Nature 1999; 401: 935–938.

    Article  CAS  PubMed  Google Scholar 

  46. Crouzet J et al. Recombinational construction in Escherichia coli of infectious adenoviral genomes. Proc Natl Acad Sci USA 1997; 94: 1414–1419.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  47. Douglas JT et al. Targeted gene delivery by tropism-modified adenoviral vectors. Nat Biotechnol 1996; 14: 1574–1578.

    Article  CAS  PubMed  Google Scholar 

  48. Dedieu J et al. Long-term gene delivery into the livers of immunocompetent mice with E1/E4-defective adenoviruses. J Virol 1997; 71: 4626–5161.

    CAS  PubMed  PubMed Central  Google Scholar 

  49. Blanche F et al. An improved anion-exchange HPLC method for the detection and purification of adenoviral particles. Gene Ther 2000; 7: 1055–1062.

    Article  CAS  PubMed  Google Scholar 

  50. Krasnykh V, Mikheeva G, Douglas J, Curiel D . Generation of recombinant adenovirus vectors with modified fibers for altering viral tropism. J Virol 1996; 70: 6839–6846.

    CAS  PubMed  PubMed Central  Google Scholar 

  51. Katagiri YU et al. Non-RGD domains of osteopontin promote cell adhesion without involving alpha v integrins. J Cell Biochem 1996; 62: 123–131.

    Article  CAS  PubMed  Google Scholar 

  52. Hidaka C et al. CAR-dependent and CAR-independent pathways of adenovirus vector-mediated gene transfer and expression in human fibroblasts. J Clin Invest 1999; 103: 579–587.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Wu E et al. A 50-kDa membrane protein mediates sialic acid-independent binding and infection of conjunctival cells by adenovirus type 37. Virology 2001; 279: 78–89.

    Article  CAS  PubMed  Google Scholar 

  54. Chiu CY et al. Structural analysis of a fiber-pseudotyped adenovirus with ocular tropism suggests differential modes of cell receptor interactions. J Virol 2001; 75: 5375–5380.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  55. Li Q et al. Assessment of recombinant adenoviral vectors for hepatic gene therapy. Hum Gene Ther 1993; 4: 403–409.

    Article  CAS  PubMed  Google Scholar 

  56. Morsy MA et al. Patient selection may affect gene therapy success. Dominant negative effects observed for ornithine transcarbamylase in mouse and human hepatocytes. J Clin Invest 1996; 97: 826–832.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Zinn KR et al. Imaging and tissue biodistribution of 99mTc-labeled adenovirus knob (serotype 5). Gene Ther 1998; 5: 798–808.

    Article  CAS  PubMed  Google Scholar 

  58. Gu DL et al. Fibroblast growth factor 2 retargeted adenovirus has redirected cellular tropism: evidence for reduced toxicity and enhanced antitumor activity in mice. Cancer Res 1999; 59: 2608–2614.

    CAS  PubMed  Google Scholar 

  59. Sarukhan A et al. Successful interference with cellular immune responses to immunogenic proteins encoded by recombinant viral vectors. J Virol 2001; 75: 269–277.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  60. Morelli AE et al. Recombinant adenovirus induces maturation of dendritic cells via an NF-kappaB-dependent pathway. J Virol 2000; 74: 9617–9628.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  61. Banchereau J, Steinman RM . Dendritic cells and the control of immunity. Nature 1998; 392: 245–252.

    Article  CAS  PubMed  Google Scholar 

  62. Weinberg DH, Ketner G . A cell line that supports the growth of a defective early region 4 deletion mutant of human adenovirus type 2. Proc Natl Acad Sci USA 1983; 80: 5383–5386.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  63. Graham FL, Prevec L . Methods for construction of adenovirus vectors. Mol Biotechnol 1995; 3: 207–220.

    Article  CAS  PubMed  Google Scholar 

  64. Benihoud K et al. Efficient, repeated adenovirus-mediated gene transfer in mice lacking both tumor necrosis factor alpha and lymphotoxin alpha. J Virol 1998; 72: 9514–9525.

    CAS  PubMed  PubMed Central  Google Scholar 

Download references

Acknowledgements

We thank Dr David Curiel for providing us with purified soluble CAR and Ad5 knob, as well as the 1D6-14 anti-fiber antibody. We thank IntroGene B.V. for accessing the PER.C6 packaging cell line. We are grateful to S Esselin for technical assistance, and to the entire staff of the animal facility at Institut Gustave Roussy.

Author information

Authors and Affiliations

  1. UMR1582 CNRS/IGR/Aventis, Institut Gustave Roussy, Villejuif, France

    E Vigne, J-F Dedieu, A Brie, A Gillardeaux, D Briot, K Benihoud, M Latta-Mahieu, M Perricaudet & P Yeh

  2. Gencell S.A., Vitry-sur-Seine, France

    E Vigne, J-F Dedieu, A Brie, A Gillardeaux, D Briot, M Latta-Mahieu & P Yeh

  3. Centre de réfe´rence IGR/Aventis/Gencell, Institut Gustave Roussy, Villejuif, 94805, Cedex, France

    P Saulnier

Authors
  1. E Vigne
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. J-F Dedieu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. A Brie
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. A Gillardeaux
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. D Briot
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. K Benihoud
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. M Latta-Mahieu
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. P Saulnier
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. M Perricaudet
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. P Yeh
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Vigne, E., Dedieu, JF., Brie, A. et al. Genetic manipulations of adenovirus type 5 fiber resulting in liver tropism attenuation. Gene Ther 10, 153–162 (2003). https://doi.org/10.1038/sj.gt.3301845

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.gt.3301845

Keywords

This article is cited by

Search

Advanced search

Quick links