Researchers track the complex roots of glaucoma in DNA

Researchers are deconstructing the genetic map of glaucoma to aid future efforts aimed at screening, prevention and treatment.

As the second leading cause of blindness in the United States after age-related macular degeneration, glaucoma has naturally drawn the interest of genetic researchers. Much is now known about the genetic components of the disease, but much remains to be uncovered.

Often the payoff of genetic research is said to be 5 to 10 years from practical clinical application. But according to researchers, what is known today about glaucoma’s genetics can already benefit some patients in the clinic.

What makes glaucoma so daunting to study, experts say, is that it is a heterogeneous disease, involving probably as many as 30 different genes that sit in different chromosomal regions, or loci. Where the mutations for the disease are located may have a bearing on how it is treated, researchers say.

“Glaucoma is not really one disease. We believe it is no less than six diseases and probably even more than that,” said John R. Samples, MD.

“Once you know the mutations for each gene at each locus you can start to predict how the glaucoma can be managed, in terms of which medicines to use and who needs surgery, and so forth,” said Dr. Samples, a professor of ophthalmology at Oregon Health Sciences University and the director of the glaucoma service at Casey Eye Institute.

“The complexity of glaucoma stems from the predicted number of genes involved,” said Mansoor Sarfarazi, PhD, who directs the Molecular Ophthalmic Genetics Lab at the University of Connecticut. “Its complex nature does not mean that glaucoma is induced by environmental factors. It can be secondary to many other biological triggers, but so far there is no direct evidence that environmental factors play a significant role in the etiology of glaucoma,” Dr. Sarfarazi said.

At least half of the types of glaucoma are thought to be inherited. This presents a double-edged sword to researchers.

On one hand, they are faced with an inherited disease that goes undetected in most cases and can have devastating effects on vision in over generations.

On the other hand, the genetic nature of the disease can be viewed as an advantage, a tool that researchers can seize upon to help track its lineage and find genetic-based methods of screening, prevention and treatment.

The genetic nature of the disease also means there is a potential for money to be made in the burgeoning business of genetic research — an element that is not lost on anyone.

Not one disease

There are currently six loci that have been labeled for open-angle glaucoma. They are known as GLC1-A to GLC1-F. With these loci labeled, the task at hand involves identifying the specific genes that encode the mutations that lead to glaucoma.

This task has been long and costly thus far, and the competition is intense.

At present, three glaucoma-related genes have been conclusively identified. The most recent one was identified by Dr. Sarfarazi and his team of researchers, who published their findings in the February issue of Science.

The optineurin gene, located on chromosome 10p14, is known as OPTN, for “optic neuropathy-inducing protein.” OPTN is expressed in the trabecular meshwork, nonpigmented ciliary epithelium, retina and brain, and it is anticipated to play a neuroprotective role.

The other two genes that have been identified are CYP1B1, which encodes cytochrome P4501B1 enzyme and is mutated in primary congenital glaucoma, and MYOC, which encodes myocilin and is mutated in juvenile-onset primary open-angle glaucoma (POAG).

The CYP1B1 gene was identified by Dr. Sarfarazi’s group about 6 years ago. It is “now proven to be a major gene for primary congenital glaucoma,” according to Dr. Sarfarazi.

“Our estimate is that CYP1B1 is involved in about 85% of familial and about 33% of sporadic cases with congenital glaucoma. We have been able to identify mutations in families which have two or more affected subjects, as well as other families with no prior history of glaucoma,” he said.

The myocilin protein — associated with the MYOC gene — is also well-documented in the literature. Dr. Samples described it as interesting that the protein has a “myosin-like domain” — the kind of myosin found in the motor proteins of skeletal and muscle tissue.

He said the other side of the myocilin protein has a mucin layer that surrounds olfactory neurons. The protein is able to interact with itself because of a “zipper-like” structure between the two sides.

The myocilin protein is believed to account for up to 14% of juvenile-onset glaucoma and 1% to 5% of adult-onset glaucoma, according to Dr. Samples.

Big genetic news

Dr. Samples believes the biggest genetic news so far of 2003 was the announcement by researchers at Wills Eye Hospital that they had linked the progression of primary open-angle glaucoma to the presence or absence of the myocilin mutation, or the mt.1 variant.

“This was the first study to do this convincingly,” Dr. Samples said, noting that his research group had tried to make the same correlation but found it to be “too messy to publish.”

Over 6 to 10 years, patients with the mt.1 variant showed a 20% progression in glaucomatous symptoms, as opposed to a 10% progression in those without the variant. The difference was more apparent when the results were evaluated over 16 to 20 years.

The study found no statistical evidence for an effect of the mt.1 variant on intraocular pressure (IOP) measurements. The authors based their conclusions on strong associations for both optic disc and visual field measurements, which they claim support the role of clinical genetic testing for the variant.

This study was presented at the annual meeting of the Association for Research in Vision and Ophthalmology.

“This means we might be able to say that a person who is mt.1 positive needs to be managed more aggressively. That’s where this all leads — with a lot of precautions and with a lot of asterisks and footnotes,” Dr. Samples said.

Still, others are skeptical about assigning such biological significance to promoter mutations such as mt.1.

Dr. Sarfarazi is among those who believe that there is thus far no compelling evidence for or against assigning any biological significance to these polymorphisms, generally known as SNPs (for single nucleotide polymorphisms).

“Everyone is trying to correlate the presence of a given SNPs to a clinical parameter, but the issue is more complicated than just saying it is present or absent,” Dr. Sarfarazi said. “What this SNP is doing biologically has never been established. Nobody is looking at the biological and function significance of these SNPs in glaucoma research. If you look at the entire genome, some 25 million SNPs have been identified, but what all these 25 million do, nobody is clear on that yet.”

Dr. Sarfarazi said he believes there is a significant lack of understanding in general regarding what an SNP is and what it does. Until these knowledge gaps are addressed, he said, no definitive conclusions about their role in glaucoma can be drawn.

“We have to understand how it works before we can correlate it to glaucoma,” he said of the mt.1 variant.

A test

Yet even if researchers do not understand the biological mechanisms of the mt.1 promoter, its presence has already demonstrated a potential commercial value, according to Dr. Samples.

A test called OcuGene, marketed by InSite Vision, provides a test for the mt.1 promoter, as well as other less well-known mutations.

The test is designed to detect the genetic marker mt.1 in the promoter region of the glaucoma-related TIGR gene. It can be used to screen for the presence of a number of mutations in the coding region of the gene in high-risk people such as relatives of glaucoma patients, ocular hypertensives and suspected glaucoma patients.

There is debate in the ophthalmic genetics community over the test’s usefulness and cost-effectiveness. At this point, the OcuGene test kit is rarely used and expensive. It costs about $300 and is not reimbursed by Medicare.

Yet Dr. Samples said the recent study linking the mt.1 variant to glaucoma progression provides a strong argument for increased use of the test in the future.

“If we now know that, for mt.1 positives, you’ve got a 20% vs. a 10% progression in 6 to 10 years, there’s a test that theoretically will help you predict if your glaucoma is going to be progressive. So if you get a patient who demands testing, it is out there,” he said.

Irene Maumenee, MD, a specialist in genetic eye disease and a professor at Johns Hopkins University’s Wilmer Eye Institute, agreed that the test may prove to be useful in screening for people who need to be examined more carefully for the disease.

“If I came from a family where this was a problem, I would want (this test) done. It may turn out that it only helps a few people, but if you can get to the right members of the right families it can make a big difference,” Dr. Maumenee said. “If you can afford it, I think you should do it. Anything is going to be cheaper than continually following someone (with the disease) over their lifetime.”

Funding enough?

Regarding the commercialization of glaucoma genetic testing, Dr. Samples makes this argument; research and development money will become more available as these types of tests are developed and deployed commercially.

The National Eye Institute devoted nearly $160 million to funding gene-based research in 2002, representing some 29% of its total grant allocations.

That number has more than doubled from the roughly $70 million the NEI dedicated in 1998, but some in the research community think it is not enough.

Dr. Samples has argued that the government is not devoting enough money to product development for glaucoma gene therapies. (See the June 1 issue, page 1.)

Yet Dr. Maumenee said she predicts it will not be more than 5 to 10 years until patients will routinely reap the benefits from the genetic knowledge that is already out there today.

For certain eye diseases, such as retinoblastoma, the gene identification happened so many years ago that “it has already made a tremendous difference in management,” according to Dr. Maumenee.

“It will happen for other diseases as well,” she said.

Dr. Sarfarazi said a full understanding of the genetics of glaucoma is still years away, and therefore the full incorporation of technology such as genetic testing and targeted drug therapy into most practices will not be a reality for some time.

“There is so much to be done, but this can only happen once we firmly establish how a (glaucoma-related) gene functions both normally and abnormally,” he said.

However, he noted, “the genetics of glaucoma has not been unrewarding.”

“Especially for families with identified mutations, at least we can identify the siblings and children who are at risk decades before they are ever going to develop glaucoma,” he said

If a healthy person is curious about his or her likelihood of developing the glaucoma that runs in the family, researchers may be able to find the gene sequence for the particular mutation they know exists in the family. With some testing they may be able to tell the healthy relative whether he or she will develop glaucoma in the future.

“We refer patients to an ophthalmologist and say they have an extremely high risk for developing glaucoma. Previously that risk would have been at 50%, but now we can change that to 99% or to 1%,” Dr. Sarfarazi said. “We can put the patients on one side of the scale or the other. This has been a major reward of our research so far.”

For Your Information:

• Mansoor Sarfarazi, PhD, can be reached at the Molecular Ophthalmic Genetics Laboratory, Surgical Research Center, Department of Surgery, University of Connecticut Health Center, 263 Farmington Ave., Farmington, CT 06030; (860) 679-3629; fax: (860) 679-7524; e-mail: mansoor@neuron.uchc.edu; Web site: www.uchc.edu.
• John R. Samples, MD, can be reached at Casey Eye Institute, 3375 SW Terwilliger Blvd., Portland, OR 97239; (503) 494-7667; fax: (503) 494-3017; e-mail: samplesj@ohsu.edu.
• Irene Maumenee, MD, can be reached at Maumenee Bldg. Ste. 517, 600 North Wolfe St., Baltimore, MD 21287-9237; (410) 955-5214; fax: (410) 614-4363; e-mail: maumenee@welchlink.welch.jhu.edu.

• InSite Vision, manufacturer of the OcuGene glaucoma genetic test, can be reached at 965 Atlantic Ave., Alameda, CA 94501; (510) 865-8800; fax: (510) 865-5700; Web site: www.insitevision.com.