Gene delivery to airway epithelial cells in vivo: a direct comparison of apical and basolateral transduction strategies using pseudotyped lentivirus vectors.

Lentivirus vectors are being investigated as gene delivery vehicles for cystic fibrosis airway gene therapy. Vesicular stomatitis virus G glycoprotein (VSV-G)-pseudotyped vectors transduce airway epithelia via receptors that are located predominantly on the basolateral surface of the airway epithelium. Effective transduction with VSV-G-pseudotyped vectors requires the use of a pre-treatment that disrupts epithelial tight junctions, allowing access to these basolateral receptors. In contrast, it has been reported that apically targeted lentiviral vectors allow efficient gene transfer in the absence of any pre-treatment. In a direct comparison of transduction by a VSV-G-pseudotyped vector, in combination with a pre-treatment with lysophosphatidylcholine (LPC), and the same vector pseudotyped with the apically targeted baculovirus GP64 envelope (without any pre-treatment), the GP64 vector was found to be significantly less efficient. However, when a pre-treatment with LPC was used the level of transduction with the GP64-pseudotyped lentiviral vector was not significantly different to that resulting from the VSV-G-pseudotyped vector. The cell types transduced with each vector were essentially the same, with the majority of cells transduced being respiratory (ciliated cells). However, unlike the VSV-G-pseudotyped vector, which results in persisting gene expression, transduction with the GP64-pseudotyped vector resulted in gene expression that declined to undetectable levels over six months, whether or not an LPC pre-treatment was used.

Kremer KL ，Dunning KR ，Parsons DW ，Anson DS ... -  《JOURNAL OF GENE MEDICINE》

Lentiviral vectors pseudotyped with baculovirus gp64 efficiently transduce mouse cells in vivo and show tropism restriction against hematopoietic cell types in vitro.

Schauber CA ，Tuerk MJ ，Pacheco CD ，Escarpe PA ，Veres G ... -  《GENE THERAPY》

Involvement of glycosaminoglycans in vesicular stomatitis virus G glycoprotein pseudotyped lentiviral vector-mediated gene transfer into airway epithelial cells.

The involvement of surface molecules in HIV-1-derived lentivirus (LV)-mediated transduction of airway epithelial cells has not been studied so far. The present study aimed to evaluate the role of glycosaminoglycans (GAGs) in gene transfer mediated by a third generation vesicular stomatitis virus G glycoprotein (VSV-G) pseudotyped LV vector in an in vitro model of polarized airway epithelial cells. Human bronchial (16HBE-S1) and tracheal (CFT1-C2) epithelial cells were grown either on plastic or on filters and transduced with the LV vector polypurine tract (PPT)-green fluoresecent protein (GFP). Zonula Occludens (ZO)-1, a marker of tight junction, and GAG localization were assessed by cytofluorimetry and confocal microscopy. Soluble GAGs and removal of cell surface GAGs were used to affect LV-mediated transduction. Extensive optimization of experimental parameters (presence of polybrene during the infection, the incubation time in the presence of LV particles, period of time intercurring between infection and gene expression analysis) was carried out in plastic-adherent cells. Polybrene resulted to be cytotoxic and was not further used. In CFT1-C2 polarized cells, EGTA treatment determined a 20% decrease in transepithelial resistance, a diminished ZO-1 localization at the tight junction location and a 31% increase in GFP positive cells. Heparane sulfate was distributed evenly on the cell surface. Heparin and soluble chondroitin sulfate A and B inhibited LV-mediated transduction in a dose-dependent fashion. These results were confirmed upon enzymatic removal of GAGs from the cell surface. Taken together, these results show that GAGs are involved in VSV-G LV transduction of airway epithelial cells.

Copreni E ，Castellani S ，Palmieri L ，Penzo M ，Conese M ... -  《-》

Lentiviral vectors pseudotyped with glycoproteins from Ross River and vesicular stomatitis viruses: variable transduction related to cell type and culture conditions.

Kahl CA ，Pollok K ，Haneline LS ，Cornetta K ... -  《MOLECULAR THERAPY》

Pseudotyped human lentiviral vector-mediated gene transfer to airway epithelia in vivo.

Johnson LG ，Olsen JC ，Naldini L ，Boucher RC ... -  《GENE THERAPY》