This is a proposal to develop a retinal model of neurodegeneration that will ultimately enable much more rapid and efficient evaluation of chemical modifiers of disease than is currently possible. We are carrying out proof-of-concept experiments using the huntingtin (Htt) protein, which causes Huntington disease when it contains an elongated tract of glutamines. The significance of this work is high, because if we are successful we will show that it is possible to isolate a discrete portion of the CNS for chemical and genetic studies, using anatomy, physiology, and function. This could help bridge cellular studies of neurodegeneration, which can rapidly generate much data, with whole animal studies, which are expensive and time consuming. The retinal model allows independent measurement of function in each eye, enabling internal controls within an animal that vastly improve statistical power. In preliminary work, we have shown that it is possible to detect retinal degeneration in the R6/2 mouse model of HD. We have further demonstrated that this is associated with progressive loss of function in tests of retinal physiology (electroretinography). We have shown that it is possible to deliver a disease-modifying compound in tiny amounts using liposome-mediated drug delivery, and to observe a beneficial effect on retinal physiology. We have also developed a behavior assay that lets us independently measure vision in the right vs. left eye of an unrestrained mouse. Finally, we have shown that it is possible to transduce large numbers of retinal neurons after a single injection of a modified adeno-associated virus. This work puts us in an excellent position to test whether we can build a model of retinal degeneration based on expression of various Htt proteins with mutations of broad interest to the HD field.
Aim 1 : Characterize viral transduction of Htt proteins within the retina. We will use AAV2 to express various forms of Htt and determine their effects on retinal neurons over time.
Aim 2 : Use ERG to measure effects on retinal physiology and compare mutants. We will use electroretinography to measure effects of various Htt proteins on retinal physiology.
Aim 3. Use optomotry to measure effects on retinal neuron function and compare mutants. We will use a specialized behavior assay to measure the effects of various Htt proteins on visual function. The work is based on the use of recombinant adeno-associated virus, micro-injection of this virus into the intraocular space of wild-type mice, and the determination of the consequences of virus-mediated Htt gene expression within the eye. We will also use state-of-the-art techniques to monitor retinal pathology, physiology, and function. If successful, this work will set the stage for much more rapid in vivo analyses of chemical and genetic modifiers of neurodegeneration. This could have an enormous impact on human health, since neurodegenerative disease is one of the single biggest health problems faced by the United States.
It is very difficult to translate cellular studies of neurodegenerative disease into animal models. Cell models are very useful for identifying early drug leads that could be translated into patient use, as well as for the identification of genes that modify the neurodegenerative disease process. However, it is very difficult, time consuming, and expensive to evaluate experimental compounds in whole animals, and to test genetic modifiers. Evaluation of small molecules is difficult because it requires large amounts of compound, and any early compound must be safe, and easily administered, with good bioavailability. Evaluation of genetic modifiers is difficult because this requires creating hybrid mice that express multiple genes of interest. This is a proposal to develop a mouse retinal model of neurodegenerative disease that could be useful for evaluating experimental compounds and modifier genes. It is based on the idea that experimental compounds can be safely administered locally to each individual eye via injection, and the genetic modifications can be delivered by a single injection of a specialized virus. If we are successful, we will show that it is possible to direct expression of a disease-associated gene to the eye, and then use highly quantitative measures of visual function to determine the relative toxicity of various mutants. We will be studying huntingtin, the protein that causes Huntington disease, a devastating neurodegenerative condition.
In Aim 1 we will create specialized viruses that allow us to efficiently express various forms of the huntingtin protein within the retina. We will determine that we are getting good expression, and will monitor animals over long periods to determine any effects of gene expression on the integrity of retinal neurons.
In Aim 2 we will use a technique called electroretinography to measure the effects of the various forms of the huntingtin protein on the physiology of neurons in the mouse retina. We will correlate any changes with pathology observed in Aim 1. We will independently measure the physiologic changes in the left vs. right eyes of the animals. This will enable much more effective studies of relative effects of the various huntingtin mutations.
In Aim 3 we will use a state-of-the-art behavior technique to measure the effects of huntingtin protein expression on visual acuity in the injected mice. This allows us to independently measure the function of the left vs. right retina, which enables much more effective studies of the various huntingtin mutations. Relevance to public health Neurodegenerative diseases represent an enormous cost to society, and afflict millions of people. There are no effective therapies, and the methods to develop new treatments and to study these diseases are relatively slow and cumbersome. If successful, this work will develop a new method to study neurodegenerative disease that could vastly improve our ability to develop new drugs and to study the basic mechanisms of disease. This will help speed the development of more effective treatments.
|Zheng, Zhiqiang; Li, Aimin; Holmes, Brandon B et al. (2013) An N-terminal nuclear export signal regulates trafficking and aggregation of Huntingtin (Htt) protein exon 1. J Biol Chem 288:6063-71|
|Li, Mei; Yasumura, Douglas; Ma, Aye Aye K et al. (2013) Intravitreal administration of HA-1077, a ROCK inhibitor, improves retinal function in a mouse model of huntington disease. PLoS One 8:e56026|