Kari Juhani Airenne

Baculoviruses exhibit restricted cell type specificity in rat brain: a comparison of baculovirus- and adenovirus-mediated intracerebral gene transfer in vivo.

Baculoviruses have recently been shown to be effective gene transfer vectors in mammalian cells. However, very little information is available about their target cell tropism in the central nervous system. We studied transduction efficiency, tropism and biodistribution of baculoviruses after local delivery to rat brain and compared their properties to adenoviruses. It was found that baculoviruses specifically transduced cuboid epithelium of the choroid plexus in ventricles and that the transduction efficiency was as high as 76±14%, whereas adenoviruses showed preference to corpus callosum glial cells and ventricular ependymal lining. Only a modest microglia response was seen after the baculovirus transduction whereas the adenovirus gene transfer led to a strong microglia response. Sensitive nested RT-PCR revealed transgene expression in the hindbrain and in ectopic organs including spleen, heart and lung, which indicates that some escape of both vectors occurs to ectopic organs after local gene transfer to the brain. We conclude that both baculovirus and adenovirus vectors can be used for local intracerebral gene therapy. The knowledge of the cell type specificity of the vectors may offer a possibility to achieve targeted gene delivery to distinct brain areas. Baculoviruses seem to be especially useful for the targeting of choroid plexus cells.

SPECT/CT imaging of baculovirus biodistribution in rat.

Non-invasive imaging provides essential information regarding the biodistribution of gene therapy vectors and it can also be used for the development of targeted vectors. In this study, we have utilized micro Single-photon emission computed tomography to visualize biodistribution of a 99mTc-polylys-ser-DTPA-biotin-labelled avidin-displaying baculovirus, Baavi, after intrafemoral (i.f.), intraperitoneal (i.p.), intramuscular (i.m.) and intracerebroventricular (i.c.v.) administration. The imaging results suggest that the virus can spread via the lymphatic network after different administration routes, also showing accumulation in the nasal area after systemic administration. Extensive expression in the kidneys and spleen was seen after i.p. administration, which was confirmed by reverse transcriptase-polymerase chain reaction and immunohistochemistry. Additionally, transduction of kidneys was seen with i.m. and i.f. administrations. We conclude that baculovirus may be beneficial for the treatment of kidney diseases after i.p. administration route.

Targeting of biotinylated compounds to its target tissue using a low-density lipoprotein receptor-avidin fusion protein

The very high binding affinity of avidin to biotin is one of the highest to occur in nature. We constructed a fusion protein composed of avidin and the endocytotic LDL receptor in order to target biotinylated molecules to cells of the desired tissues. In addition to the native avidin, charge-mutated and nonglycosylated avidins were utilized as part of the fusion proteins, in order to modify its properties. All of the fusion protein versions retained the biotin-binding capacity. Although the specificity was not increased, however, fusion proteins composed of natural avidin and nonglycosylated avidin bound most efficiently to the biotinylated ligands. Fluorescence microscopy and atomic force microscopy studies revealed the expression of the fusion protein on cell membranes, and demonstrated specific and high-affinity binding of biotin to the low-density lipoprotein receptor (LDLR)–avidin fusion protein in vitro. Additionally, systemically administered biotinylated ligand targeted with high specificity the intracerebral tumors of rats that were expressing fusion protein after the virus-mediated gene transfer. These results suggest that local gene transfer of the fusion protein to target tissues may offer a novel tool for the delivery of biotinylated molecules in vitro and in vivo for therapeutic and imaging purposes.