Genetic retargeting of adenovirus: novel strategy employing 'deknobbing' of the fiber.
In: https://hal.science/hal-00116716 ; 2006, 2006
report
Zugriff:
BACKGROUND: We studied the ability of adenovirus type 5 (Ad5) to encapsidate new cellular ligands carried by their fibers to yield functional retargeted vectors for gene therapy. Recombinant Ad5 fibers containing shaft repeats 1 to 7 and an extrinsic trimerization motif, and terminated by its native knob or amino acid motifs containing RGD, have been rescued into infectious virions. METHODS: Polypeptide ligands of cell surface molecules, including single-chain antibodies or epidermal growth factor, were cloned into recombinant fibers. Phenotypic analysis of fiber constructs and rescuing into the Ad5 genome were performed. Recombinant viruses were characterized with reference to fiber content, growth rate and infectivity. RESULTS: A major limiting factor for recovering viable recombinant Ad5 carrying fiber-fused polypeptide ligands was apparently the ability of the ligand to fold correctly within the cellular cytoplasm. This constraint has previously not been systematically evaluated in the literature. Phenotypic analysis of the fiber-ligand fusions showed that their degree of cytoplasmic solubility correlated with their ability to yield viable Ad5 vectors. Our results suggested that the fiber manipulations diminish virus growth rate, probably through different, opposing effects: (i) the reduced shaft length increases fiber solubility in the absence of the knob but (ii) diminishes virus entry, and (iii) the absence of the knob alters the overall protein composition of the virion and decreases its fiber copy number. CONCLUSIONS: Based on our findings, cytoplasmic solubility and cytoplasmic ligand reactivity of fiber-ligand fusion proteins are the best prediction criterion for viability and recovery of genetically retargeted Ad vectors. Copyright 2002 John Wiley & Sons, Ltd.
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Genetic retargeting of adenovirus: novel strategy employing 'deknobbing' of the fiber.
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Autor/in / Beteiligte Person: | Magnusson, Mk ; Hong, Ss ; Henning, P. ; Boulanger, P. ; Lindholm, L. ; Department of Medical Microbiology and Immunology Göteborg ; Göteborgs Universitet = University of Gothenburg (GU) ; Got-A-Gene, AB ; Stena Center 1B ; Virologie et pathogenèse virale (VPV) ; Université Claude Bernard Lyon 1 (UCBL) ; Université de Lyon-Université de Lyon-Centre National de la Recherche Scientifique (CNRS) ; The work in Gothenburg (MKM, PH, LL) was supported by the European Community (contract no. QLK3-CT-1999-01262), the Swedish Medical Research Council (grant no. K98-06X-12624- 01), the Swedish Cancer Society (grant no. 0512-B99-12XCC), the Inga Britt and Arne Lundberg Foundation (grant no. 205/ 96), and by grants from Got-A-Gene AB, Gothenburg, Sweden. The work in Lyon (SSH, PB) was financially supported by the French Ministe`re de la Recherche et de la Technologie (PRFMMIP AO-98), and the French Foundation for Cystic Fibrosis (Vaincre la Mucoviscidose). Expert technical contributions from Elisabeth Pettersson and Laure Franqueville are gratefully acknowledged. Reinder Bolhuis is thanked for the generous supply of the clones for the scFv G250 and the scTCR. |
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Zeitschrift: | https://hal.science/hal-00116716 ; 2006, 2006 |
Veröffentlichung: | HAL CCSD, 2006 |
Medientyp: | report |
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