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Double genetic modification of adenovirus fiber with RGD polylysine motifs significantly enhances gene transfer to isolated human pancreatic islets.

Publication ,  Journal Article
Contreras, JL; Wu, H; Smyth, CA; Eckstein, CP; Young, CJ; Seki, T; Bilbao, G; Curiel, DT; Eckhoff, DE
Published in: Transplantation
July 15, 2003

BACKGROUND: New strategies for improving durable functional islet mass will be instrumental in facilitating islet transplantation as a cure for type 1 diabetes mellitus. The ability to transfer immunoregulatory or cytoprotective genes into pancreatic islets may enhance survival. Adenoviral vectors (Ad5) have been used widely to deliver therapeutic genes to different tissues. Limitations associated with the use of Ad5 for gene therapy are related to the reliance of the virus on the presence of its primary receptor, the transient nature of the transgene expression, and the immediate inflammatory and immune response elicited by the infection. Because the arginine-glycine-aspartame (RGD) and polylysine (pK7) motifs have been shown to enhance Ad5 infection through an Ad5 receptor-independent pathway, we hypothesized that they could act additively to improve infectivity and reduce toxicity to isolated human pancreatic islets (IHPI). METHODS: Hand-picked IHPI were infected with nonmodified Ad5, single-modified Ad5 with RGD (Ad5RGD) or pK7 (ad5pK7), and Ad5RGDpK7. Transfection efficiency was evaluated by green fluorescent protein and luciferase expression. Apoptosis was assessed using a quantitative assay, activation of caspase 3 by a colorimetric assay, nuclear factor (NF)-kappaB nuclear translocation using a promoter-luciferase NF-kappaB responsive construct, regulated on activation normal T-cell expressed and secreted (RANTES) by enzyme-linked immunosorbent assay. In vivo functionality was evaluated after transplantation into diabetic nonobese diabetic severe combined immunodeficiency mice. RESULTS: Compared with unmodified and singly-modified Ad5 vectors, Ad5RGDpK7 demonstrated the highest infectivity. After the infection of IHPI with adenoviral vectors using the minimal dose required to infect greater than 80% of the islet cells (Ad5, 500 viral particles [VP]/cell; Ad5RGD and Ad5pK7, 10 VP/cell; Ad5RGDpK7, 0.1 VP/cell), islets infected with Ad5RGDpK7 presented a significant reduction in apoptosis, NF-kappaB nuclear translocation, RANTES expression, and higher glucose disposal rate; reduced Ad5-driven specific Th1 and antibody response were also observed. CONCLUSIONS: Ad5RGDpK7 exhibited higher transfection efficiency, allowing a significant reduction in the viral dose required to infect greater than 80% of the islet cells. The reduction in the viral dose was associated with reduced toxicity, inflammation, and immune responses related to Ad5 infection. This strategy may thus be used to successfully modify isolated pancreatic islets.

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Published In

Transplantation

DOI

ISSN

0041-1337

Publication Date

July 15, 2003

Volume

76

Issue

1

Start / End Page

252 / 261

Location

United States

Related Subject Headings

  • Transplantation, Heterologous
  • Tissue Donors
  • Surgery
  • Polylysine
  • Oligopeptides
  • Mice, SCID
  • Mice, Inbred NOD
  • Mice, Inbred C57BL
  • Mice
  • Male
 

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Contreras, J. L., Wu, H., Smyth, C. A., Eckstein, C. P., Young, C. J., Seki, T., … Eckhoff, D. E. (2003). Double genetic modification of adenovirus fiber with RGD polylysine motifs significantly enhances gene transfer to isolated human pancreatic islets. Transplantation, 76(1), 252–261. https://doi.org/10.1097/01.TP.0000066361.02042.CA
Contreras, Juan L., Hongju Wu, Cheryl A. Smyth, Christopher P. Eckstein, Carlton J. Young, Toshiro Seki, Guadalupe Bilbao, David T. Curiel, and Devin E. Eckhoff. “Double genetic modification of adenovirus fiber with RGD polylysine motifs significantly enhances gene transfer to isolated human pancreatic islets.Transplantation 76, no. 1 (July 15, 2003): 252–61. https://doi.org/10.1097/01.TP.0000066361.02042.CA.
Contreras JL, Wu H, Smyth CA, Eckstein CP, Young CJ, Seki T, et al. Double genetic modification of adenovirus fiber with RGD polylysine motifs significantly enhances gene transfer to isolated human pancreatic islets. Transplantation. 2003 Jul 15;76(1):252–61.
Contreras, Juan L., et al. “Double genetic modification of adenovirus fiber with RGD polylysine motifs significantly enhances gene transfer to isolated human pancreatic islets.Transplantation, vol. 76, no. 1, July 2003, pp. 252–61. Pubmed, doi:10.1097/01.TP.0000066361.02042.CA.
Contreras JL, Wu H, Smyth CA, Eckstein CP, Young CJ, Seki T, Bilbao G, Curiel DT, Eckhoff DE. Double genetic modification of adenovirus fiber with RGD polylysine motifs significantly enhances gene transfer to isolated human pancreatic islets. Transplantation. 2003 Jul 15;76(1):252–261.

Published In

Transplantation

DOI

ISSN

0041-1337

Publication Date

July 15, 2003

Volume

76

Issue

1

Start / End Page

252 / 261

Location

United States

Related Subject Headings

  • Transplantation, Heterologous
  • Tissue Donors
  • Surgery
  • Polylysine
  • Oligopeptides
  • Mice, SCID
  • Mice, Inbred NOD
  • Mice, Inbred C57BL
  • Mice
  • Male