Glaucoma, a progressive optic neuropathy usually involving elevated intraocular pressure, is a leading cause of blindness worldwide. The incidence increases with age and is high among certain ethnic groups. Although treatments are available, they are not ideal. Glaucoma also occurs in the absence of increased intraocular pressure and current treatments are not effective for these patients. Given these problems, improved strategies for therapy are needed. Glaucoma is a prime candidate for gene therapy approaches. The pathology of glaucoma involves structures throughout the eye including the trabecular meshwork (TM), ciliary body epithelium (CE), ciliary muscle (CM), and retinal ganglion cells (RGC) providing several candidate targets for gene delivery. Numerous gene therapy approaches for glaucoma are feasible without knowing the specific disease related gene. For example, the TM or CM could be targeted to increase outflow, ciliary body epithelial cells could be targeted to reduce fluid production, and neuroprotective strategies could be applied to prevent cell death. Progress in glaucoma gene therapy is severely hampered by a lack of appropriate promoters to express the transgenes. Tissue specific promoters are not yet available nor are any promoters known to express for considerable lengths of time in relevant cells. We have designed a novel system based on Herpes simplex virus (HSV) amplicon vectors that will allow us to isolate potential gene therapy promoters under in vivo conditions. The overall goal is to isolate promoters that are suitable for ocular gene therapy.
The specific aims are:
Specific Aim 1 : We will test the hypothesis that one or more promoters suitable for use in gene therapy in the TM can be isolated using our HSV amplicon system. We will do this by using chromatin immunoprecipitation to isolate expressing DNA from primary TM cells, cloning the selected DNA into our promoterless GFP amplicon vector, and selecting GFP expressing cells from rat anterior chamber cells following transduction with the promoter library amplicons.
Specific Aim 2 : We will test the hypothesis that a specific set of transcriptional regulatory proteins exist in the trabecular meshwork by combining transcription factor profiling of TM cells with bioinformatic analysis of promoters from the library isolated in Aim 1 and the promoters of genes identified by others (EST and microarray databases).
| Liu, Xuyang; Brandt, Curtis R; Rasmussen, Carol A et al. (2007) Ocular drug delivery: molecules, cells, and genes. Can J Ophthalmol 42:447-54 |
