Highly Efficient Retrograde Gene Transfer into Motor Neurons by a Lentiviral Vector Pseudotyped with Fusion Glycoprotein

Miyabi Hirano,Kazuto Kobayashi,Shigeki Kato,Tomoaki Okada,Kenta Kobayashi,Hiroyuki Yaginuma,Joseph Charles Glorioso

doi:10.1371/journal.pone.0075896

Abstract

The development of gene therapy techniques to introduce transgenes that promote neuronal survival and protection provides effective therapeutic approaches for neurological and neurodegenerative diseases. Intramuscular injection of adenoviral and adeno-associated viral vectors, as well as lentiviral vectors pseudotyped with rabies virus glycoprotein (RV-G), permits gene delivery into motor neurons in animal models for motor neuron diseases. Recently, we developed a vector with highly efficient retrograde gene transfer (HiRet) by pseudotyping a human immunodeficiency virus type 1 (HIV-1)-based vector with fusion glycoprotein B type (FuG-B) or a variant of FuG-B (FuG-B2), in which the cytoplasmic domain of RV-G was replaced by the corresponding part of vesicular stomatitis virus glycoprotein (VSV-G). We have also developed another vector showing neuron-specific retrograde gene transfer (NeuRet) with fusion glycoprotein C type, in which the short C-terminal segment of the extracellular domain and transmembrane/cytoplasmic domains of RV-G was substituted with the corresponding regions of VSV-G. These two vectors afford the high efficiency of retrograde gene transfer into different neuronal populations in the brain. Here we investigated the efficiency of the HiRet (with FuG-B2) and NeuRet vectors for retrograde gene transfer into motor neurons in the spinal cord and hindbrain in mice after intramuscular injection and compared it with the efficiency of the RV-G pseudotype of the HIV-1-based vector. The main highlight of our results is that the HiRet vector shows the most efficient retrograde gene transfer into both spinal cord and hindbrain motor neurons, offering its promising use as a gene therapeutic approach for the treatment of motor neuron diseases.

Highlights

Motor neuron diseases, including amyotrophic lateral sclerosis and spinal muscular atrophy, are characterized by progressive muscle weakness and paralysis resulting from degeneration of motor neurons in the spinal cord and brain [1,2,3]
We developed a vector with highly efficient retrograde gene transfer (HiRet) in the central nervous system by pseudotyping an human immunodeficiency virus type 1 (HIV-1)-based vector with fusion glycoprotein B type (FuG-B), in which the cytoplasmic domain of rabies virus glycoprotein (RV-G) was replaced by the corresponding part of vesicular stomatitis virus glycoprotein (VSV-G) [29]
We developed a novel type of vector showing neuron-specific retrograde gene transfer (NeuRet) by pseudotyping the HIV-1 vector with fusion glycoprotein C type (FuG-C), in which the short C-terminal segment of the extracellular domain and transmembrane/cytoplasmic domains of RV-G were substituted with the corresponding regions of VSV-G [31]. (The structure of viral fusion glycoproteins is schematically illustrated in Figure 1.) Both neuronal and glial cells around the injection site are transduced by the HiRet vector, whereas only neuronal cells around this site are transduced by the NeuRet vector [29,31]

Summary

Introduction

Motor neuron diseases, including amyotrophic lateral sclerosis and spinal muscular atrophy, are characterized by progressive muscle weakness and paralysis resulting from degeneration of motor neurons in the spinal cord and brain [1,2,3]. The development of gene therapy technique to introduce transgenes that promote neuronal survival and protection into motor neurons should be an effective approach for the treatment of motor neuron diseases. The intramuscular injection of an adenoviral vector enables the delivery of transgenes into motor neurons [8]. This adenovirusmediated transfer of genes involved in neuronal survival and protection, such as brain-derived neurotrophic factor, ciliary neurotrophic factor, glial cell line-derived neurotrophic factor, and neuronal apoptosis inhibitory protein, prevents motor neuron death in axotomy-induced injury models [9,10,11,12,13].

Methods

Results

Conclusion