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MiR133b-mediated inhibition of EGFR-PTK pathway promotes rAAV2 transduction by facilitating intracellular trafficking and augmenting second-strand synthesis |
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| Authors: | Xiaoping Huang Xiao Wang Ling Li Qizhao Wang Wentao Xu Wenlin Wu Xiaolan Xie Yong Diao |
| Affiliation: | 1. College of Chemical Engineering and Materials Sciences, Quanzhou Normal University, Quanzhou, China;2. School of Medicine, Huaqiao University, Quanzhou, China Contribution: Methodology (equal), Validation (equal);3. School of Medicine, Huaqiao University, Quanzhou, China Contribution: Formal analysis (equal), Investigation (equal);4. School of Medicine, Huaqiao University, Quanzhou, China Contribution: Data curation (equal), Investigation (equal);5. College of Chemical Engineering and Materials Sciences, Quanzhou Normal University, Quanzhou, China Contribution: Methodology (equal), Software (equal);6. College of Chemical Engineering and Materials Sciences, Quanzhou Normal University, Quanzhou, China Contribution: Data curation (equal), Resources (equal);7. School of Medicine, Huaqiao University, Quanzhou, China |
| Abstract: | Recombinant adeno-associated virus (rAAV) is an extremely attractive vector in the in vivo delivery of gene therapy as it is safe and its genome is simple. However, challenges including low permissiveness to specific cells and restricted tissue specificity have hindered its clinical application. Based on the previous studies, epidermal growth factor receptor-protein tyrosine kinase (EGFR-PTK) negatively regulated rAAV transduction, and EGFR-positive cells were hardly permissive to rAAV transduction. We constructed a novel rAAV-miRNA133b vector, which co-expressed miRNA133b and transgene, and investigated its in vivo and in vitro transduction efficiency. Confocal microscopy, live-cell imaging, pharmacological reagents and labelled virion tracking were used to analyse the effect of miRNA133b on rAAV2 transduction and the underlying mechanisms. The results demonstrated that miRNA133b could promote rAAV2 transduction and the effects were limited to EGFR-positive cells. The increased transduction was found to be a direct result of decreased rAAV particles degradation in the cytoplasm and enhanced second-strand synthesis. ss-rAAV2-miRNA133b vector specifically increased rAAV2 transduction in EGFR-positive cells or tissues, while ss-rAAV2-Fluc-miRNA133b exerted an antitumor effect. rAAV-miRNA133b vector might emerge as a promising platform for delivering various transgene to treat EGFR-positive cell-related diseases, such as non-small-cell lung cancer. |
| Keywords: | EGFR-protein tyrosine kinase miRNA133b rAAV2 second-strand DNA synthesis ubiquitination |