Adenovirus vector-induced expression of the C-X-C chemokine IP-10 is mediated through capsid-dependent activation of NF-kappa B

The use of adenovirus vectors for gene therapy has been limited by well-def ined cellular and humoral immune responses, We have previously shown that a denovirus vectors rapidly induce the expression of the C-X-C chemokine, int erferon-inducible protein 10 (IP-IO), in vivo, Various first-generation, ty pe 5 adenovirus vectors, including adCMV beta gal and W-psoralen-inactivate d adenovirus, equally induced the expression of IP-10 mRNA as early as 3 h following infection in mouse renal epithelial cells (REC), Luciferase repor ter experiments using deletional mutants of the murine IP-10 5'-flanking re gion revealed that transcriptional activation of the IP 10 promoter by adCM V beta gal was dependent on the -161- to -96-bp region upstream of the tran scription start site. In electrophoretic mobility shift assays, adCMV beta gal, adCMV-GFP, FG140, and transcription-defective adenovirus induced prote in binding to oligonucleotides containing a consensus sequence for NF-kappa B at position -113 of the IP-10 promoter. Supershift assays confirmed an i ncrease in binding activity of NF-kappa B p65 but not p50 or cRel in REC ce lls infected with various replication-deficient adenoviruses, Coinfection o f REC cells with adCMV beta gal and an adenoviral vector expressing I kappa B alpha resulted in suppression of adCMV beta gal-induced expression of IP -10 at 6 and 16 h, further strengthening the conclusion that adenovirus-ind uced activation of IP 10 is dependent on NF-kappa B, The induction of IP-10 appeared to be direct because infection with adenovirus vectors failed to induce the expression of the potent IP-IO stimulators, interferon gamma and tumor necrosis factor alpha. Together, these findings demonstrate that ade novirus vectors directly induce the expression of IP-IO through capsid depe ndent activation of NF-kappa B.