Targeting and Purification of Metabolically Biotinylated Baculovirus

Abstract

Targeting viral entry is one of the major goals in the development of vectors for gene therapy. Ideally, the coupling of each new targeting motif would not require changes in vector structure. To achieve this, we developed novel metabolically biotinylated baculoviral vectors by displaying a small biotin acceptor peptide (BAP) fused either to different sites in the baculovirus glycoprotein gp64 or to the transmembrane anchor of vesicular stomatitis virus G protein. Baculoviral particles were biotinylated during vector production by coexpression of Escherichia coli biotin ligase (BirA). The insertion of BAP at amino acid position 283 of gp64 resulted in the most efficient biotin display. Unlike vectors with lower biotin display, these vectors also showed improved transduction when retargeted to transferrin, epidermal growth factor, and CD46 receptors overexpressed on rat glioma and human ovarian carcinoma cells. Biotinylated baculoviral vectors could also be concentrated by one-step magnetic particle-based capture to reach titers up to 10(10) plaque-forming units/ml. These results demonstrate the utility of metabolically biotinylated baculovirus for vector targeting and viral purification applications.

How to avoid complement attack in baculovirus-mediated gene delivery

“…(see also van Oers, 2011). These approaches are more widely known as baculovirus display technology constituting a eukaryotic display platform for a wide range of applications such as drug discovery, vaccination or targeted gene therapy (Grabherr and Ernst, 2010;Hu et al, 2008;Kaikkonen et al, 2008;Raty et al, 2004). The most widely used technique for surface engineering makes use of the trimeric major baculoviral envelope glycoprotein GP64 as a fusion partner (Kadlec et al, 2008).…”

Baculovirus is an efficient vector for the transduction of the eye: comparison of baculovirus‐ and adenovirus‐mediated intravitreal vascular endothelial growth factor D gene transfer in the rabbit eye

“…Purification was carried out as described by Kaikkonen et al [24]. Virus preparations were tested for sterility and analysed for the absence of endotoxin and mycoplasma contaminations.…”

Quality control and analytical methods for baculovirus-based products

“…Making use of the singular characteristics of rBac, novel strategies for improved rBac recovery have been successfully developed such as: (1) a concentration step by cation-exchange chromatography using the electrostatic surface charge of rBac (Barsoum, 1999); and (2) concentration of biotinylated rBac vectors using magnetic beads (Kaikkonen et al, 2008). Despite these technological advances, a fully scaleable DSP for non-conjugated rBac has yet to be accomplished.…”