title: Conjugate-based targeting of recombinant adeno-associated virus type 2 vectors by using avidin-linked ligands.
abstract: The development of targeted vectors, capable of tissue-specific transduction, remains one of the important aspects of vector modification for gene therapy applications. Recombinant adeno-associated virus type 2 (rAAV-2)-based vectors are nonpathogenic, have relatively low immunogenicity, and are capable of long-term transgene expression. AAV-2 vectors bind primarily to heparan sulfate proteoglycan (HSPG), a receptor that is present in many tissues and cell types. Because of the widespread expression of HSPG on many tissues, targeted transduction in vivo appears to be limited with AAV-2 vectors. Thus, development of strategies to achieve transductional targeting will have a profound benefit in the future application of these vectors. We report here a novel conjugate-based targeting method to enhance tissue-specific transduction of AAV-2-based vectors. The present report utilized a high-affinity biotin-avidin interaction as a molecular bridge to cross-link purified targeting ligands, produced genetically as fusion proteins to core-streptavidin, in a prokaryotic expression system. Conjugation of the bispecific targeting protein to the vector was achieved by biotinylating purified rAAV-2 without abolishing the capsid structure, internalization, and subsequent transgene expression. The tropism-modified vectors, targeted via epidermal growth factor receptor (EGFR) or fibroblast growth factor 1alpha receptor (FGFR1alpha), resulted in a significant increase in transduction efficiency of EGFR-positive SKOV3.ip1 cells and FGFR1alpha-positive M07e cells, respectively. Further optimization of this method of targeting should enhance the potential of AAV-2 vectors in ex vivo and in vivo gene therapy and may form the basis for developing targeting methods for other AAV serotype capsids.
authors.affiliation: Department of Pathology, LHRB 513, The University of Alabama at Birmingham, 701 19th Street South, Birmingham, AL 35294, USA. sponnazh@path.uab.edu
authors.forename: Selvarangan
authors.initials: S
authors.lastname: Ponnazhagan
authors.forename: Gandham
authors.initials: G
authors.lastname: Mahendra
authors.forename: Sanjay
authors.initials: S
authors.lastname: Kumar
authors.forename: John A
authors.initials: JA
authors.lastname: Thompson
authors.forename: Mark
authors.initials: M
authors.lastname: Castillas
mesh_terms: D019921:Biotinylation; D002460:Cell Line; D000229:Dependovirus / Q000145:classification / Q000235:genetics*; D066246:ErbB Receptors / Q000378:metabolism*; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000235:genetics*; D019812:Heparan Sulfate Proteoglycans / Q000378:metabolism; D006801:Humans; D008024:Ligands; D020794:Receptor Protein-Tyrosine Kinases / Q000378:metabolism*; D051496:Receptor, Fibroblast Growth Factor, Type 1; D017468:Receptors, Fibroblast Growth Factor / Q000378:metabolism*; D011995:Recombination, Genetic; D019809:Streptavidin / Q000378:metabolism*; D014161:Transduction, Genetic
chemical_list: D019812:Heparan Sulfate Proteoglycans; D008024:Ligands; D017468:Receptors, Fibroblast Growth Factor; D019809:Streptavidin; D066246:ErbB Receptors; C496348:FGFR1 protein, human; D020794:Receptor Protein-Tyrosine Kinases; D051496:Receptor, Fibroblast Growth Factor, Type 1