Animal model connects gene mutation and normal-tension glaucoma

Take home message: Transgenic mice engineered to have extra copies of the TBK1 gene showed effects similar to those in human patients with normal-tension glaucoma, providing clues to pathophysiology and potential treatments.

By Vanessa Caceres; Reviewed by John H. Fingert, MD, PhD

Iowa City, IA-Mice that have a gene defect associated with normal-tension glaucoma (NTG) can provide clues to the causes of human disease and to future treatments of NTG, according to John H. Fingert, MD, PhD, associate professor of ophthalmology and visual sciences, University of Iowa Carver College of Medicine, Iowa City.

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Dr. Fingert initially addressed the genetics of NTG, which sometimes is caused by a single gene and other times by the combined actions of many genes.

The first gene associated with NTG, optineurin (OPTN), was discovered in 2002. Multiple studies have confirmed this finding, and it is associated with 1% to 2% of NTG cases, he said.

More recently, Dr. Fingert’s lab has discovered that gene duplications and triplications of the TANK binding kinase 1 gene (TBK1) are also associated with NTG about 1% to 2% of the time. Studies in the United States, Japan, and Australia have confirmed that duplication of TBK1 is associated with the disease.

“TBK1 is expressed in just the right place in the eye to cause normal-pressure glaucoma,” Dr. Fingert said.

The gene is known to have roles in important biological pathways, including autophagy.

“In autophagy, cells engulf and clear abnormal proteins, intracellular bacteria, and defective organelles,” Dr. Fingert said.

Autophagy is a known response to stress, such as a nutritional deficit. Because autophagy includes three genes known to lead to glaucoma, his lab became interested in studying the pathway.

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The hypothesis was that increased TBK1 activity from a gene duplication may activate optineurin, which, in turn, stimulates autophagy and ultimately may lead to retinal ganglion death and glaucoma, he said.

To test this hypothesis, Dr. Fingert's lab used an engineered line of transgenic mice to mimic what is seen in human patients with glaucoma. The mice that they studied had the normal two copies of mouse TBK1 plus an additional copy of the human TBK1 gene.

“We basically bred these mice to see if extra copies of the TBK1 gene would cause low-pressure glaucoma in them as in our human patients,” he said.

The first analyses of the mice showed that those animals with an extra copy of the TBK1 gene did not have higher IOPs than mice with a normal number of copies, which indicated these mice are a good model of NTG. When analyzing the retina, the first finding was increased TBK1 protein in retinal ganglion cells of the mice.

“This supported our model that an extra dose of the gene has an effect in TBK1 gene activity,” he said.

Lab researchers evaluated the mice for glaucoma with histological methods. They counted retinal ganglion cells in the transgenic mice and compared them with what was seen in a control group of normal littermates. Similar to human glaucoma, researchers observed a significant reduction in retinal ganglion cells in transgenic mice with an extra copy of the TBK1 gene. These losses were consistent with NTG in humans.

Researchers in Dr. Fingert’s lab then bred mice to have two extra doses of the TBK1 gene, and they noticed that the mice with two extra doses had even more retinal ganglion cell loss.

Finally, the researchers also analyzed optic nerve tissue by counting individual axons in these animals. They similarly found significantly fewer axons in mice aged to 12 months that had an extra TBK1 gene.

“This is more evidence that the mice developed features of NTG,” Dr. Fingert said.

The research shows that animals engineered to have the same TBK1 gene defect as human patients develop key features of human glaucoma, Dr. Fingert said.

This continuing research provides strong evidence that mutation of the TBK1 gene can lead to glaucoma and may provide insights into disease mechanisms and future treatments, Dr. Fingert concluded.

“Hopefully, this will open up a new field for low-pressure glaucoma research and treatment,” he said.

John H. Fingert, MD, PhD

E: john-fingert@uiowa.edu

This article was adapted from Dr. Fingert’s presentation at the 2014 meeting of American Academy of Ophthalmology. He did not indicate any proprietary interest in the subject matter.