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Synthesis of adenoviral targeting molecules by intein-mediated protein ligation

Gene Therapy volume 10, pages 1362–1369 (2003)Cite this article

Abstract

Adenoviral vectors infect cells through the binding of capsid proteins to cell-surface receptors. The ubiquitous expression of adenoviral receptors in human tissues represents an obstacle toward the development of systemically deliverable vectors for cancer therapy, since effective therapy may require delivery to specific sites. For these reasons, major efforts are directed toward the elimination of the native tropism combined with identification of ligands that bind to tumor-specific cell-surface proteins. Highthroughput technologies have identified potential targeting ligands, which need to be evaluated for their ability to retarget adenovirus to alternative receptors. Here, we present a strategy that permits the routine analysis of adenoviral targeting ligands. We use intein-mediated protein ligation as a means to produce functional biological molecules, that is, adenoviral targeting molecules that function as adapters between cellular receptors and the adenovirus fiber protein. We demonstrate the versatility of the present system by conjugating targeting ligands that differ in size and nature including an apolipoprotein E synthetic peptide, the basic fibroblast growth factor and folic acid. The resulting adenoviral targeting molecules mediate adenoviral gene delivery in cells that express the corresponding receptor.

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Acknowledgements

We thank Joe de los Angeles (Novartis Pharmaceutical Corporation, Summit, NJ, USA) for providing us with structural representation of CAR D1 (PDB accession code: 1F5W) and folic acid (PDB accession code: 1CD2) and Inca Ghosh (New England Biolabs, Inc., Berverly, MA, USA) for helpful advice on the IMPACTTM-TWIN system. We thank Michele Kaloss (GTI, Gaithersburg, MD, USA) for plasmid sequencing and Jun Xu for helpful technical assistance in the initiation of these studies. We are most grateful to Gene Liau and Theodore Smith (GTI, Gaithersburg, MD, USA) for critical reading of this manuscript.

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  1. Genetic Therapy, Inc., A Novartis Company, Gaithersburg, MD, USA

    O Nyanguile, C Dancik, J Blakemore, K Mulgrew, M Kaleko & S C Stevenson

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  1. O Nyanguile
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  2. C Dancik
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  3. J Blakemore
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  6. S C Stevenson
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Correspondence to S C Stevenson.

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Nyanguile, O., Dancik, C., Blakemore, J. et al. Synthesis of adenoviral targeting molecules by intein-mediated protein ligation. Gene Ther 10, 1362–1369 (2003). https://doi.org/10.1038/sj.gt.3301989

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