Modifications in adenoviral coat fiber proteins and transcriptional regulatory sequences enhance transgene expression.
OBJECTIVE
To characterize the adenoviral properties required to enhance intracellular transgene expression for gene therapy.


METHODS
Primary human fibroblasts and macrophages were infected with standard replication-defective adenoviruses, adenoviral vectors containing modified fiber coat proteins expressing Arg-Gly-Asp (RGD) or heparin sulfate binding moieties, or a tetracycline-regulatable transgene transcription system. Each of these vectors expressed the beta-galactosidase gene (beta-Gal), which was quantified by flow cytometry. Ankle joints from rats with adjuvant induced arthritis were transduced intraarticularly with each of the vectors and B-Gal expression was quantified by flow cytometry.


RESULTS
Primary human fibroblasts and macrophages displayed marked increases in transgene expression from both modified fiber protein vectors and from the tetracycline-regulatable vector, compared to an unmodified vector expressing the transgene from the cytomegalovirus promoter/enhancer. In the rat model, the modified fiber protein vectors and the tetracycline-regulatable vector system also displayed increased transgene expression in inflamed rat joints.


CONCLUSIONS
Adenovirus attachment and uptake by cells and promoter strength limit transgene expression from conventional adenoviral vectors in models of rheumatoid arthritis.
Division of Rheumatology, Northwestern University Medical School, Chicago, Illinois 60611, USA.
Harris
H
Perlman
Hongtao
H
Liu
Constantinos
C
Georganas
James M
JM
Woods
M Asif
MA
Amin
Alisa E
AE
Koch
Thomas
T
Wickham
Imre
I
Kovesdi
Toshiaki
T
Mano
Kenneth
K
Walsh
Richard M
RM
Pope
D000256:Adenoviridae / Q000201:enzymology / Q000235:genetics*; D000818:Animals; D001169:Arthritis, Experimental / Q000473:pathology / Q000628:therapy; D036022:Capsid Proteins / Q000235:genetics*; D002478:Cells, Cultured; D005260:Female; D005347:Fibroblasts / Q000821:virology; D015967:Gene Expression Regulation, Viral; D018014:Gene Transfer Techniques; D005809:Genes, Regulator / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D006614:Hindlimb / Q000473:pathology; D008264:Macrophages / Q000821:virology; D051381:Rats; D011917:Rats, Inbred Lew; D011995:Recombination, Genetic; D014158:Transcription, Genetic; D019076:Transgenes; D001616:beta-Galactosidase / Q000235:genetics / Q000378:metabolism
D036022:Capsid Proteins; D001616:beta-Galactosidase