Optimized Delivery of C3 Transferase by Lentiviral Vectors forCNS Injury Therapy Open Access

Laur, Olga (2009)

Permanent URL: https://etd.library.emory.edu/concern/etds/9z9030052?locale=en


Identifying targets for effective treatment of neurodegenerative diseases, brain and spinal cord injuries is one of the fundamental goals of modern neuroscience. These pose tremendous challenges and burdens on our society, especially as life expectancy increases.

In the past decades, through the elucidation of several inhibitory pathways governing the non-permissive nature of the central nervous system, RhoA GTPase has emerged as an important inhibitory signal convergence point, and therefore an attractive therapeutic target. Investigators in previous studies have successfully demonstrated axonal regeneration both in vitro and in vivo by RhoA inhibition using various forms of C3 transferase from Clostridiumbotulinum, a potent RhoA inhibitor. However, the poor permeability of this enzyme together with its short-term expression limited the success of such strategies.

In this study we have explored a novel axonal regeneration approach, based on the delivery of C3 transferase into the central nervous system utilizing FIV or HIV lentiviral vectors. Such strategies will potentially overcome limitations associated with direct C3 enzyme delivery, and lead to its constitutive, sustained cellular expression. Consequently, this will lead to continuous axonal regeneration necessary for complete neuronal recovery.

We were able to develop several C3-containing lentiviral plasmids that can be used for lentiviral vector production. Moreover, we assessed the C3 expression levels and functionality of these plasmids to determine their readiness for lentiviral production. Achieving constitutive, sustained production of C3 transferase through plasmid development and subsequent lentiviral generation is an important milestone in exploring the full regenerative and therapeutic potentials of RhoA inhibition.

Table of Contents

Introduction 1

Methods 10

Results 16

Discussion 30

References 42

Tables and Figures

Table 1 49

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