Abstract
The replication-incompetent adenovirus (Ad) vector is one of the most promising vectors for gene therapy; however, systemic administration of Ad vectors results in severe hepatotoxicities, partly due to the leaky expression of Ad genes in the liver. Here we show that nuclear factor-kappa B (NF-κB) mediates the leaky expression of Ad genes from the Ad vector genome, and that the inhibition of NF-κB leads to the suppression of Ad gene expression and hepatotoxicities following transduction with Ad vectors. Activation of NF-κB by recombinant tumor necrosis factor (TNF)-α significantly enhanced the leaky expression of Ad genes. More than 50% suppression of the Ad gene expression was found by inhibitors of NF-κB signaling and siRNA-mediated knockdown of NF-κB. Similar results were found when cells were infected with wild-type Ad. Compared with a conventional Ad vector, an Ad vector expressing a dominant-negative IκBα (Adv-CADNIκBα), which is a negative regulator of NF-κB, mediated approximately 70% suppression of the leaky expression of Ad genes in the liver. Adv-CADNIκBα did not induce apparent hepatotoxicities. These results indicate that inhibition of NF-κB leads to suppression of Ad vector-mediated tissue damages via not only suppression of inflammatory responses but also reduction in the leaky expression of Ad genes.
Highlights
Genes by inserting microRNA-targeted sequences in the Ad vector genome[9]
Similar results were found in H1299 cells (Figure S2). These results indicated that nuclear factor-kappa B (NF-κ B) was essential for Ad gene expression and replication of wild-type Ad (WT-Ad), and that NF-κ B induced the leaky expression of the Ad genes following transduction with Ad vectors
The results showed that NF-κ B promoted Ad gene expression following treatment with WT-Ad and an Ad vector (Figs 2–4 and 6)
Summary
Problems remain with all of these novel Ad vectors, including the complexity of the Ad vector preparation and some remaining Ad vector-induced hepatotoxicities. To overcome these drawbacks, the mechanisms of the E1A gene-independent leaky expression of Ad genes should be clarified. Transduction with Ad vectors causes activation of a number of transcriptional factors. Injection with recombinant Ad vectors immediately induces production of inflammatory cytokines associated with innate immune responses[14,15], suggesting that several transcriptional factors would be activated in the innate immune responses. NF-κ B is a ubiquitous transcriptional factor which promotes the expression of a large number of genes, those from gene families associated with host immune responses[16]. An Ad vector expressing a dominant-negative Iκ Bα (Adv-CADNIκ Bα ), which is a negative regulator of NF-κ B, mediated the significant suppression of the leaky expression of Ad genes, with the result that no apparent hepatotoxicities were induced
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