Role identified for glaucoma gene and signaling pathway

November 1, 2008

Another piece of the puzzle of the mechanism involved in the elevation of IOP has been identified with the discovery of the role of the gene sFRP-1 in the Wnt signaling pathway.

Key Points

Iowa City, Iowa-In a development that one day could lead to a novel disease intervention strategy for glaucoma, investigators have found that an antagonist of the Wnt signaling pathway is involved in the development of elevated IOP, according to John Fingert, MD, PhD, assistant professor in the Department of Ophthalmology and Visual Sciences, University of Iowa Carver College of Medicine, Iowa City.

The team discovered that the protein gene sFRP-1 (secreted frizzled-related protein-1), a Wnt antagonist, was overexpressed in glaucomatous human trabecular meshwork cells. In a mouse model, however, restoration of Wnt signaling reduced IOP levels that had become elevated following intravitreal injection of an adenoviral vector encoding sFRP-1.

"This set of experiments identified a gene that's part of a signaling pathway that seems to play a role in regulating eye pressure, a key risk factor for glaucoma," Dr. Fingert said. "That's what's exciting; it's a new biologic pathway that can lead to elevated pressure in the eye. It represents some new information about the potential causes of glaucoma, and we hope that this will help us understand who is at risk for glaucoma, and that down the road it might also provide some insights for developing new interventions."

The investigators began their latest experiments with cultured trabecular meshwork cells derived from normal and glaucomatous donors and compared them using RNA differential display. They found increased sFRP-1 expression in the glaucomatous cells when compared with controls. Because sFRP-1 is an antagonist of Wnt signaling, the investigators also wanted to determine if Wnt signaling pathway genes were expressed in the trabecular meshwork. After examining cells and tissues from normal donors, they detected mRNA for several Wnt genes, receptors, antagonists, coreceptors, and other members of the Wnt pathway.

Addition of sFRP-1

The next step was the addition of sFRP-1 to ex vivo perfusion-cultured human eyes. Because the trabecular meshwork regulates aqueous humor outflow and IOP, the objective was to determine whether the addition of sFRP-1 would change those functions. Investigators found that the outflow rate was reduced in eyes that were perfused in medium containing recombinant human sFRP-1 compared with control eyes, starting 1 day after treatment; the reduction reached 55% at day 4. Another result was a decrease in protein levels of beta-catenin, a Wnt signaling mediator.

Those findings were substantiated in a mouse model. The mice received intravitreal injections of an adenoviral vector encoding sFRP-1. A highly significant pressure increase (p < 0.001) was found 5 days after injection in the injected eyes, whereas IOP remained unchanged in eyes given a null injection.

For the final step in the process, the researchers administered glycogen synthase kinase (GSK), a selective inhibitor of a downstream suppressor of Wnt signaling, in the mouse eyes. The increase in IOP that had been induced by injection of sFRP-1 was significantly diminished by the topical ocular administration of GSK.

That observation provides further evidence that the Wnt signaling pathway plays a role in maintaining normal IOP and that alterations in this pathway caused by increased expression of sFRP-1 can produce ocular hypertension, Dr. Fingert said. He added that a natural future direction would be further study of the Wnt pathway.

"We know from our current report that the Wnt signaling pathway is involved in eye pressure regulation, and it's very exciting to think about experiments that might look at this pathway as a target for intervention," Dr. Fingert said. "Other genes also may be important in regulating eye pressure and might be good targets for developing new treatments. We're just learning about the biology of the Wnt signaling pathway in glaucoma, and I think it's very exciting that it may have important clinical implications for better treatment and diagnosis in the future."

According to Allan R. Shepard, PhD, assistant director, Discovery Research, Alcon Research, Ltd., this approach of blocking signaling and reversing elevated pressure also could be used with other genes that have been associated with glaucoma.

"In the future, what we're trying to do is take other genes that we've identified through all of our discovery work on differential forms of gene expression and develop the same paradigm of administering a viral vector into a mouse efficacy model to determine the effect on mouse IOP, with the presumption that it will correlate to human IOP in glaucoma," Dr. Shepard said. He was not one of the authors of the recently published study on the Wnt pathway but is familiar with the work.

Drugs that might emerge from this research approach would reverse elevated eye pressure by blocking the build-up of extracellular matrix that impedes aqueous outflow, thereby reversing the disease process, Dr. Shepard said. In contrast, the glaucoma drugs on the market today do not specifically address the disease mechanism.