GSBS Dissertations and Theses

Date of Completion

8-5-2014

Document Type

Doctoral Dissertation

Academic Program

Interdisciplinary Graduate Program

Department

Neurology

First Thesis Advisor

Miguel Sena-Esteves, Ph.D.

Keywords

Dependovirus, G(M1) Ganglioside, GM1 Gangliosidosis, Genetic Vectors, Lysosomal Storage Diseases, beta-Galactosidase

Subjects

Dissertations, UMMS; Dependovirus; G(M1) Ganglioside; Gangliosidosis, GM1; Genetic Vectors; Lysosomal Storage Diseases; beta-Galactosidase

Abstract

GM1 gangliosidosis is a lysosomal storage disorder caused by a deficiency in the catabolizing enzyme β-galactosidase (βgal). This leads to accumulation of GM1-ganglioside (GM1) in the lysosome inducing ER stress and cell death. GM1 gangliosidosis is primarily a disorder of the central nervous system (CNS) with peripheral organ involvement. In this work we report two major findings, 1) systemic treatment of GM1 gangliosidosis with an adenoassociated virus (AAV9) encoding mouse-βgal (mβgal) in a GM1 gangliosidosis mouse model (βGal-/-), and 2) an investigation into an intracranial injection of a therapeutic AAVrh8 encoding mβgal. Systemic treatment of GM1 gangliosidosis with AAV9 resulted in a moderate expression of enzyme in the CNS, reduction of GM1 storage, significant retention of motor function and a significant increase in lifespan. Interestingly, the therapeutic effect was more robust in females. Intracranial injections of AAVrh8 vector expressing high levels of βgal resulted in enzyme spread throughout the brain, significant retention of motor function and a significant increase in lifespan. Histological alterations were also found at the injection site in both βGal-/- and normal animals. We constructed a series of vectors with a range of decreasing enzyme expression levels to investigate the cause for the unanticipated result. Microarrays were performed on the injection site and we showed that a lower expressing AAVrh8-mβgal vector mitigated the negative response. Intracranial injection of this newly developed vector was shown to clear lysosomal storage throughout the CNS of βGal-/- mice. Taken together, these studies indicate that a combined systemic and fine-tuned intracranial approach may be the most effective in clearing lysosomal storage completely in the CNS while providing therapeutic benefit to the periphery.

Comments

This dissertation includes 6 videos that are referenced in Chapter II. See Additional Files available below.

DOI

10.13028/M26K64

Rights and Permissions

Copyright is held by the author, with all rights reserved.

CMW - V1 M 3e11 258d.wmv (28548 kB)
Male animals near humane endpoint of untreated βGal-/- mice. βGal-/- + 3e11 vg at 258 days, βGal+/+ untreated at 258 days and βGal-/- + PBS at 221 days.

CMW - V2 F 3e11 260d.wmv (22605 kB)
Female animals near humane endpoint of untreated βGal-/- mice. βGal-/- + 3e11 vg at 260 days, βGal+/+ untreated at 260 days and βGal-/- + PBS at 236 days.

CMW - V3 M 1e11 263d.wmv (45746 kB)
Male animals near humane endpoint of untreated βGal-/- mice βGal-/- + 1e11 vg at 249 days, βGal+/- untreated at 264 days and βGal-/- + PBS at 263 days.

CMW - V4 F 1e11 264d.wmv (28754 kB)
Female animals near humane endpoint of untreated βGal-/- mice. βGal-/- + 3e11 vg at 258 days, βGal+/+ untreated at 278 days and βGal-/- + PBS at 264 days.

CMW - V5 F 3e11 590d.wmv (22865 kB)
Treated female βGal-/- + 3e11 vg at 590 days.

CMW - V6 F 3E11 566d.wmv (22999 kB)
Treated female βGal-/- + 3e11 vg at 563 days.

Share

COinS
 
 

To view the content in your browser, please download Adobe Reader or, alternately,
you may Download the file to your hard drive.

NOTE: The latest versions of Adobe Reader do not support viewing PDF files within Firefox on Mac OS and if you are using a modern (Intel) Mac, there is no official plugin for viewing PDF files within the browser window.