Challenges in monoclonal antibody-based therapies.

Therapeutic monoclonal antibodies (mAbs) are the fastest growing class of new therapeutic molecules. They hold great promises for the treatment of a variety of diseases, including chronic inflammatory diseases and cancer. However, the current manufacturing and purification processes cause limitations in the production capacity of therapeutic antibodies, leading to an increase in cost. Genetic delivery of therapeutic monoclonal antibodies by in vivo production offers a new potential solution to these problems. Firstly, therapeutic efficacy can be improved by maintaining stable therapeutic, non-toxic levels within the blood circulation over a long period of time. Repeated high-dose bolus injections could be avoided, thereby reducing the possibility of side-effects. Secondly, the high cost of manufacturing and purification of the therapeutic antibodies could be reduced, making an in vivo/ex vivo mAb gene transfer an economically viable and attractive option. In general, three approaches can be used for the stable long-term expression and secretion of therapeutic antibodies in vivo: 1) direct in vivo administration of integrating vectors carrying a mAb gene, 2) grafting of ex vivo genetically modified autologous cells, and 3) implantation of an encapsulated antibody producing heterologous or autologous cells. This paper describes the key factors and problems associated with the current antibody-based immunotherapies and reviews prospects for genetic in vivo delivery of therapeutic antibodies.

Improving safety of gene therapy.

Gene therapy has progressed from early clinical trials to first commercial gene therapy drugs. While there is a long history with the side-effects and adverse effects of pharmaceutical drugs, drugs based on gene delivery have presented new challenges for researchers, clinicians and regulatory authorities. On the path from early pre-clinical research to final commercial products, gene therapy tools and production methods have undergone tremendous changes to improve safety and efficacy. Deletion of adenovirus replication genes E1 and E3 has progressed to gutless adenoviruses with all viral genes removed; similarly evolution of lentiviral vectors has progressed from first generation viruses to safer third generation self-inactivating vectors. Improved chromatographic methods have eased the purification of viruses and delivery reservoirs, such as collagen or silicon collars for cardiovascular gene transfer have decreased systemic leakage of viruses; together with tissue-specific promoters and imaging of the biodistribution of viral particles, gene therapy specificity and safety can be improved even further. This review will introduce gene delivery vectors used in gene therapy and highlight key approaches used to improve their safety.

Enhanced gene delivery by avidin-displaying baculovirus.

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.

Baculovirus capsid display: a novel tool for transduction imaging.

Baculoviruses are enveloped insect viruses that can carry large quantities of foreign DNA in their genome. Baculoviruses have proved to be very promising gene therapy vectors but little is known about their transduction mechanisms in mammalian cells. We show in this study that Autographa californica multiple nuclear polyhedrosis virus capsid is compatible with the incorporation of desired proteins in large quantities. Fusions can be made to the N-terminus or C-terminus of the major capsid protein vp39 without compromising the viral titer or functionality. As an example of the baculovirus capsid display we show a tracking of the baculovirus transduction in mammalian cells by an enhanced green fluorescent protein (EGFP)-displaying virus. Our confocal and electron microscopy results suggest that the transduction block in mammalian cells is not in the endosomal escape, as previously proposed, but rather in the cytoplasmic transport or nuclear entry of the virus capsid. Our results also suggest that the EGFP-tagged virus can be used for visualization of the virus biodistribution in vivo. Furthermore, capsid-modified baculoviruses hold great promise for the nuclear and subcellular targeting of transgenes and as a novel peptide display system for a variety of eukaryotic applications.