Genetically modified adenoviral vector with the protein transduction domain of Tat improves gene transfer to CAR-deficient cells

Abstract

The transduction efficiency of Ad (adenovirus) depends, to some extent, on the expression level of CAR (coxsackievirus and Ad receptor) of a target cell. The low level of CAR on the cell surface is a potential barrier to efficient gene transfer. To overcome this problem, PTD.AdeGFP (where eGFP is enhanced green fluorescent protein) was constructed by modifying the HI loop of Ad5 (Ad type 5) fibre with the Tat (trans-activating) PTD (protein transduction domain) derived from HIV. The present study showed that PTD.AdeGFP significantly improved gene transfer to multiple cell types deficient in expression of CAR. The improvement in gene transfer was not the result of charge-directed binding between the virus and the cell surface. Although PTD.AdeGFP formed aggregates, it infected target cells in a manner different from AdeGFP aggregates precipitated by calcium phosphate. In addition, PTD.AdeGFP was able to transduce target cells in a dynamin-independent pathway. The results provide some new clues as to how PTD.AdeGFP infects target cells. This new vector would be valuable in gene-function analysis and for gene therapy in cancer.

A novel vector for a rapid generation of fiber-mutant adenovirus based on one step ligation and quick screening of positive clones

“…Electron microscopy study on the obtained fiber modified adenoviruses demonstrated typical dispersedly distributed adenoviral particles except for Ad5HI-Tat PTD virus. The particles of the latter formed aggregates (data not shown), which was consistent with our previous study (Liu et al, 2009).…”

“…Synthetic small peptide RGD, Tat PTD, NGR or poly-lysine (pK7) has been used to modify the fiber knob genetically by other groups to improve the tropsim of adenovirus (Bouri et al, 1999;Wickham et al, 1997;Dmitriev et al, 1998;Han et al, 2007;Liu et al, 2009;Mizuguchi et al, 2005;Zheng et al, 2007). In addition, a peptide SIK-VAV was recently showed to be able to bind to alpha integrin and chemically modified adenoviral vector using SIKVAV peptide could increase gene transfer to alpha integrin positive cells (Stevenson et al, 2007).…”

A targeting peptide improves adenovirus-mediated transduction of a glioblastoma cell line

“…A number of strategies have been developed, primarily based on the use of bispecific antibodies (25,26), creation of fiber-chimeric Ad (13)(14)(15)(16) and direct incorporation into viral capsid of peptide analogues specific for a tumor-associated receptor (17)(18)(19)(20)(21). The latter has been preferred for cancer cellspecific Ad retargeting, in which finding new cancer targeting peptides is crucial.…”

Noncovalently Associated cell-penetrating Peptides for Gene Delivery Applications

“…Retroviruses, such as HIV and murine leukemia virus (MLV), have their protein capsid enclosed in a lipid envelope while adenovirus and adeno-associated virus are non-enveloped viruses with only a protein capsid and envelope protein on their surface. In both cases, the outer layer of the virus has a negative surface charge at physiological pH [119] and is responsible for initiating infection. Proteins responsible for the binding of the virus with the cell membrane are located on the virus capsid or embedded in the lipid envelope of the virus.…”