Lentiviral Vectors for Gene Delivery to the Nervous System - Gene Delivery and Therapy for Neurological Disorders - page 23

Chapter 2

Lentiviral Vectors for Gene Delivery to the Nervous System

Ioanna Eleftheriadou and Nicholas D. Mazarakis

Abstract

The effi cient management and development of therapeutic strategies for disorders of the nervous system
still remains a major medical challenge. Gene therapy for the nervous system diseases is particularly chal-
lenging due to the post-mitotic nature of neuronal cells and the restricted accessibility of the brain itself.
Viral vectors based on lentiviruses are particularly attractive vehicles, routinely used in developing gene-
based therapies to treat neurological diseases. Due to their unique properties, which allow them to trans-
duce most nervous system cell types, maintaining strong, and long-term transgene expression, they present
a versatile and powerful tool for many research and gene therapy applications. Lentiviral vectors pseudo-
typed with envelope glycoproteins derived from various viruses, such as VSV and rabies have been shown
to be able to genetically modify cells with good effi ciency and broad tropism. This chapter discusses lenti-
viral vectors properties and applications in gene therapy for neurodegenerative diseases, presenting some
of the recent progress in this fi eld. We also present the materials and methods necessary to generate high-
titer lentiviral vectors. Methods and applications involving lentiviral production are frequently changing.
Here we describe the current protocols used and optimized in our laboratories that allow us to produce
high-titer lentiviral vector preparations for both in vitro and in vivo applications. Full detailed protocols
describe here step-by-step the lentiviral vector production from DNA preparation, culturing of 293T pro-
ducer cells to transfection for viral production, and titration of the lentiviral vector preparations. We fur-
ther present standard in vitro transduction experiments and the in vivo applications for which these vectors
are used in our research. In the in vivo application section surgical details are presented.

Key words Lentiviral vectors, Gene Therapy, Neurodegenerative Diseases, Lentiviral vector production,

In vivo delivery of lentiviral vectors

1 Introduction

Gene therapy is the introduction of a functional gene, the alterna-
tion or removal of a defective one within an individual's cells and
biological tissues to provide a therapeutic advantage [ 1 ]. Human
Gene Therapy utilizing viral vector systems has gained consider-
able attention over the past two decades. It is considered one of
the most promising approaches developed for disorders that up to
date lack effective treatment, including both acquired and inher-
ited diseases [ 2 ]. This has a promising therapeutic advantage
compared to traditional medicine by achieving long-lasting

1.1 Gene Therapy