Abstract

The long term goal is to develop methods for clinical application of gene therapy to human neurodegenerative diseases. This proposal focuses on ex vivo and in vivo gene therapy to provide the potent neurotrophic factor, GDNF, to dopamine (DA) neurons in the Parkinsonian (PD) brain model. Herpes simplex amplicon (HSV), adenoviral (Ad) and retroviral (RV) vectors will be made for GDNF , a frameshift mutant GDNF as a control, and the cellular marker gene, nuclear localized LacZnl. Vectors will be made containing each of 3 cellular promoters, the chicken B-actin constitutive, the neuronal promoter, NSE, or the astrocyte promoter, GFAP, or viral promoters. Promoter specificity, strength and duration of expression in the context of HSV, Ad, and RV vectors will be compared in primary cultures of neurons and glia, and in vivo in rat brain. Vectors will be used for in vivo and ex vivo gene therapy in well characterized rat models of PD to study effects of GDNF gene therapy on damaged DA neurons in the adult brain and fetal DA neurons grafted into adult brain. A battery of behavioral tests of DA-dependent striatal functions will be used to compare efficacy of the different approaches. The effects of gene therapy with GDNF will also be compared to those of injecting recombinant GDNF into the brain. In addition, HSV vectors harboring the neuroprotective proto-oncogene, bcl-2, under control of the tyrosine hydroxylase promoter will be made and used in vivo to determine whether expression of bcl-2 in DA neurons an adult brain rescues them from damage induced by neurotoxin MPP+. Methods in molecular biology, immunocytochemistry, neuromorphometry, primary neuronal culture, Elisa assay, bioassay, rodent microsurgery and behavioral testing will be applied. These studies are directly relevant to clinical intervention in humans suffering from PD and to clinical trials utilizing fetal grafts. More generally, they promise to increase our knowledge on the potential of gene therapy to treat neurodegenerative diseases and methods for obtaining long term gene transgene expression in CNS from viral vectors.

  Funding Agency

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS031957-03
Application #
2750874
Study Section
Neurology B Subcommittee 2 (NEUB)
Program Officer
Murphy, Diane
Project Start
1996-09-01
Project End
2000-03-31
Budget Start
1998-08-01
Budget End
2000-03-31
Support Year
3
Fiscal Year
1998
Total Cost
Indirect Cost

  Institution

Name
Children's Memorial Hospital (Chicago)
Department
Type
DUNS #
City
Chicago
State
IL
Country
United States
Zip Code
60611

  Publications

Khodr, Christina E; Becerra, Amanda; Han, Ye et al. (2014) Targeting alpha-synuclein with a microRNA-embedded silencing vector in the rat substantia nigra: positive and negative effects. Brain Res 1550:47-60
Khodr, Christina E; Pedapati, Jyothi; Han, Ye et al. (2012) Inclusion of a portion of the native SNCA 3'UTR reduces toxicity of human S129A SNCA on striatal-projecting dopamine neurons in rat substantia nigra. Dev Neurobiol 72:906-17
Khodr, Christina E; Sapru, Mohan K; Pedapati, Jyothi et al. (2011) An ?-synuclein AAV gene silencing vector ameliorates a behavioral deficit in a rat model of Parkinson's disease, but displays toxicity in dopamine neurons. Brain Res 1395:94-107
Han, Y; Chang, Q A; Virag, T et al. (2010) Lack of humoral immune response to the tetracycline (Tet) activator in rats injected intracranially with Tet-off rAAV vectors. Gene Ther 17:616-25
Ebert, Allison D; Hann, Hoo Jae; Bohn, Martha C (2008) Progressive degeneration of dopamine neurons in 6-hydroxydopamine rat model of Parkinson's disease does not involve activation of caspase-9 and caspase-3. J Neurosci Res 86:317-25
Sapru, Mohan K; Yates, Jonathan W; Hogan, Shea et al. (2006) Silencing of human alpha-synuclein in vitro and in rat brain using lentiviral-mediated RNAi. Exp Neurol 198:382-90
Ebert, Allison D; Chen, Feng; He, Xiaolong et al. (2005) A tetracycline-regulated adenovirus encoding dominant-negative caspase-9 is regulated in rat brain and protects against neurotoxin-induced cell death in vitro, but not in vivo. Exp Neurol 191 Suppl 1:S80-94
Jiang, L; Rampalli, S; George, D et al. (2004) Tight regulation from a single tet-off rAAV vector as demonstrated by flow cytometry and quantitative, real-time PCR. Gene Ther 11:1057-67
Chen, Feng; Chang, Roger; Trivedi, Marcus et al. (2003) Caspase proteolysis of desmin produces a dominant-negative inhibitor of intermediate filaments and promotes apoptosis. J Biol Chem 278:6848-53
Gamblin, T Chris; Chen, Feng; Zambrano, Angara et al. (2003) Caspase cleavage of tau: linking amyloid and neurofibrillary tangles in Alzheimer's disease. Proc Natl Acad Sci U S A 100:10032-7

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