CAR-dependent and CAR-independent pathways of adenovirus vector-mediated gene transfer and expression in human fibroblasts.

Publication Type	Academic Article
Authors	Hidaka C, Milano E, Leopold P, Bergelson J, Hackett N, Finberg R, Wickham T, Kovesdi I, Roelvink P, Crystal R
Journal	J Clin Invest
Volume	103
Issue	4
Pagination	579-87
Date Published	02/01/1999
ISSN	0021-9738
Keywords	Adenoviruses, Human, Capsid Proteins, Gene Transfer Techniques, Genetic Vectors, Receptors, Virus
Abstract	Primary fibroblasts are not efficiently transduced by subgroup C adenovirus (Ad) vectors because they express low levels of the high-affinity Coxsackie virus and adenovirus receptor (CAR). In the present study, we have used primary human dermal fibroblasts as a model to explore strategies by which Ad vectors can be designed to enter cells deficient in CAR. Using an Ad vector expressing the human CAR cDNA (AdCAR) at high multiplicity of infection, primary fibroblasts were converted from being CAR deficient to CAR sufficient. Efficiency of subsequent gene transfer by standard Ad5-based vectors and Ad5-based vectors with alterations in penton and fiber was evaluated. Marked enhancement of binding and transgene expression by standard Ad5 vectors was achieved in CAR-sufficient fibroblasts. Expression by AdDeltaRGDbetagal, an Ad5-based vector lacking the arginine-glycine-aspartate (RGD) alphaV integrin recognition site from its penton base, was achieved in CAR-sufficient, but not CAR-deficient, cells. Fiber-altered Ad5-based vectors, including (a) AdF(pK7)betagal (bearing seven lysines on the end of fiber) (b) AdF(RGD)betagal (bearing a high-affinity RGD sequence on the end of fiber), and (c) AdF9sK betagal (bearing a short fiber and Ad9 knob), demonstrated enhanced gene transfer in CAR-deficient fibroblasts, with no further enhancement in CAR-sufficient fibroblasts. Together, these observations demonstrate that CAR deficiency on Ad targets can be circumvented either by supplying CAR or by modifying the Ad fiber to bind to other cell-surface receptors.
DOI	10.1172/JCI5309
PubMed ID	10021467
PubMed Central ID	PMC408101