Enhanced Gene Delivery by Avidin-Displaying Baculovirus

Abstract

Flexible alteration of virus surface properties would be beneficial for enhanced and targeted gene delivery. A useful approach could be based on a high-affinity receptor-ligand pair, such as avidin and biotin. In this study, we have constructed an avidin-displaying baculovirus, Baavi. Avidin display was expected to enhance cell transduction due to the high positive charge of avidin in physiological pH and to provide a binding site for covering the virus with desired biotinylated ligands. Successful incorporation of avidin on the virus envelope was detected by immunoblotting and electron microscopy. Multiple biotin-binding sites per virus were detected with fluorescence-correlation spectroscopy and tight biotin binding was observed using an optical biosensor, IAsys. Baavi showed a 5-fold increase in transduction efficiency in rat malignant glioma cells (BT4C) and a 26-fold increase in rabbit aortic smooth muscle (RAASMC) cells compared to wild-type baculovirus. Enhanced transduction was also observed with biotinylated target cells. Biotinylated epidermal growth factor (EGF) enabled specific targeting of the virus with high efficiency to EGF receptor-expressing (SKOV-3) cells. An additional advantage of the avidin display was demonstrated with biotinylated paramagnetic particles, which enabled magnetic targeting. Altogether, we show that avidin display is a rapid and versatile method to improve viral properties for gene delivery.

Magnetic resonance imaging of viral particle biodistribution in vivo

“…This finding is in agreement with the previous work 15 and results from in vivo MRI study. 20 Furthermore, neither apparent gross cytotoxic effects nor hydrocephalus was associated with the injection of Baavi and bUSPIO in vivo on histology sections or MRI.…”

“…The results are in general agreement with our previous results using gold and larger iron oxide particles. 15 Transduction assays in HepG2 cell lines show that bUSPIO coating of the avidin-displaying baculovirus (Baavi) increased both transduction efficiency ( Figure 2a) and transgene expression level (Figure 2b). …”

Enhanced angiogenesis and reduced infarct size by vascular endothelial growth factor D is not translated to behavioral outcome in a rat model of ischemic stroke

“…All entry clones were completely sequenced to verify the sequences before cloning of VEGF-D ∆N∆C into a pBVboostFGII expression vector using the BVboost system LR-reaction (Laitinen et al 2005). Avidin displaying baculoviruses (Räty et al 2004) encoding for VEGF-D ∆N∆C (BAAVI-VEGF-D ∆N∆C ) were generated by using an improved Bac-to-Bac method (Airenne et al 2003). For the Baculo-lacZ virus, a nuclear targeted β-galactosidase (β nt -Gal) cassette with a CMV promoter was inserted into the StuI site of transfer vector pFASTBac1 (pFB) (Gibco BRL, Life Technologies), generating plasmid pFBCMV-β nt (Airenne et al 2000).…”

(Strept)avidin-displaying lentiviruses as versatile tools for targeting and dual imaging of gene delivery

“…Construction of avidin-and streptavidin-displaying vectors began by the removal of gp64 gene from the Baavi vector. 11 The VSV-G TM/CTD (VSV-GED) 18 domain was amplified by PCR using the following primers: forward 5 0 -GGGGTGATACTGGGCTATCC AA-3 0 , reverse 5 0 -AGATCTTTACTTTCCAAGTCGGT TCA-3 0 and ligated to SmaI (New England Biolabs Inc.) digested vector giving rise to AVD-VSV-GED baculovirus vector. Tetrameric avidin was replaced with streptavidin sequence resulting in SA-VSV-GED vector.…”

“…hHF gene (pENTR 221-Ferritin; RZPD German Resource Center for Genome Research, Berlin, Germany) and LacZ 11 were cloned to the third-generation lentivirus transfer plasmid pBOB-CAG under CAG (chicken betaactin promoter with CMV-IE enhancer) promoter, using Figure 5 Longitudinal results of T2*-weighted gradient echo magnetic resonance images in vivo. Rat brain was imaged 3-211 days after injection of (a) 111 In-labeled SA/GP64-hHF lentivirus, (b) SA/GP64-LacZ or (c) 111 In-labeled multi-Lys-DTPA chelate alone.…”