Long-term expression is essential for virtually all gene therapy treatments of neurodegenerative disorders. Helper virus-free Herpes Simplex Virus (HSV-1) vectors, lentivirus vectors, or adeno- associated virus vectors can support recombinant gene expression for 6 to 14 months. Thus, significant improvements in long-term expression are required for human gene therapy. Our approach is to view the large, ~150 kb, genome of an HSV-1 vector as a minichromosome. We used boundary elements to create a euchromatin-like domain, and enhancers to turn on expression from specific promoters within this domain. During the previous grant period, we identified specific enhancers and promoters that support long- term expression. We isolated an upstream enhancer from the tyrosine hydroxylase promoter that supports long-term expression. We developed HSV-1 vectors that support inducible, long-term expression. We identified promoters that support glutamatergic or GABAergic neuron-specific, long- term expression. We developed targeted gene transfer to specific types of neurons, with long-term expression. We used these advances to support development of specific gene therapies. First, we corrected the rat model of Parkinson's disease by coexpressing three DA biosynthetic enzymes and a vesicular monoamine transporter (4-gene-vector). Expression in GABAergic striatal neurons was observed for 14 months. The 4-gene-vector supported higher levels of behavioral correction, higher levels of DA, and only this vector supported regulated release of DA. Second, we showed that genetic activation of protein kinase C (PKC) pathways in small groups of rat postrhinal cortex neurons enhances learning of visual object discriminations. Moreover, in aged rats, we showed that activation of PKC pathways in small groups of hippocampal dentate granule neurons corrects deficits in spatial learning. The goal of this proposal is to elucidate specific mechanisms that support long-term expression. The first specific aim will isolate and characterize specific enhancers, and their cognate enhancer binding proteins, that support long-term expression. The second specific aim will characterize the chromosomal state of HSV-1 vectors that support long-term expression. The third specific aim will overexpress specific enhancer binding proteins, or chromatin modifying enzymes, to improve long- term expression. Long-term recombinant gene expression is essential for virtually all gene therapy treatments of neurological diseases. During the previous grant period, we identified specific promoters that support long-term expression from Herpes Simplex Virus (HSV-1) vectors. This proposal will elucidate specific mechanisms that support long-term expression.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
3R01NS045855-06S1
Application #
7913100
Study Section
Special Emphasis Panel (ZRG1-BDCN-Y (06))
Program Officer
Tagle, Danilo A
Project Start
2003-04-15
Project End
2010-01-31
Budget Start
2009-09-01
Budget End
2010-01-31
Support Year
6
Fiscal Year
2009
Total Cost
$11,470
Indirect Cost
Name
Harvard University
Department
Neurology
Type
Schools of Medicine
DUNS #
047006379
City
Boston
State
MA
Country
United States
Zip Code
02115
Zhang, Guo-Rong; Zhao, Hua; Cook, Nathan et al. (2017) Characteristic and intermingled neocortical circuits encode different visual object discriminations. Behav Brain Res 331:261-275
Zhang, Guo-Rong; Zhao, Hua; Abdul-Muneer, P M et al. (2015) Neurons can be labeled with unique hues by helper virus-free HSV-1 vectors expressing Brainbow. J Neurosci Methods 240:77-88
Zhang, Guo-rong; Zhao, Hua; Cao, Haiyan et al. (2012) Targeted gene transfer of different genes to presynaptic and postsynaptic neocortical neurons connected by a glutamatergic synapse. Brain Res 1473:173-84
Zhang, Guo-Rong; Zhao, Hua; Choi, Eui M et al. (2012) CaMKII, MAPK, and CREB are coactivated in identified neurons in a neocortical circuit required for performing visual shape discriminations. Hippocampus 22:2276-89
Zhang, Guo-Rong; Zhao, Hua; Cao, Haiyan et al. (2012) Overexpression of either lysine-specific demethylase-1 or CLOCK, but not Co-Rest, improves long-term expression from a modified neurofilament promoter, in a helper virus-free HSV-1 vector system. Brain Res 1436:157-67
Cao, Haiyan; Zhang, Guo-rong; Geller, Alfred I (2011) Antibody-mediated targeted gene transfer of helper virus-free HSV-1 vectors to rat neocortical neurons that contain either NMDA receptor 2B or 2A subunits. Brain Res 1415:127-35
Zhang, Guo-rong; Li, Xu; Cao, Haiyan et al. (2011) The vesicular glutamate transporter-1 upstream promoter and first intron each support glutamatergic-specific expression in rat postrhinal cortex. Brain Res 1377:1-12
Zhang, Guo-Rong; Zhao, Hua; Li, Xu et al. (2011) A 16 bp upstream sequence from the rat tyrosine hydroxylase promoter supports long-term expression from a neurofilament promoter, in a helper virus-free HSV-1 vector system. Brain Res 1415:109-18
Zhang, Guo-rong; Cao, Haiyan; Kong, Lingxin et al. (2010) Identified circuit in rat postrhinal cortex encodes essential information for performing specific visual shape discriminations. Proc Natl Acad Sci U S A 107:14478-83
Zhang, Guo-rong; Geller, Alfred I (2010) A helper virus-free HSV-1 vector containing the vesicular glutamate transporter-1 promoter supports expression preferentially in VGLUT1-containing glutamatergic neurons. Brain Res 1331:12-9

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