Primary open-angle glaucoma genes

A substantial fraction of glaucoma has a genetic basis. About 5% of primary open angle glaucoma (POAG) is currently attributed to single-gene or Mendelian forms of glaucoma (ie glaucoma

caused by mutations in myocilinor optineurin). Mutations in these genes have a high likelihood of leading to glaucoma and are rarely seen in normal subjects. Other cases of POAG have a more

complex genetic basis and are caused by the combined effects of many genetic and environmental risk factors, each of which do not act alone to cause glaucoma. These factors are more

frequently detected in patients with POAG, but are also commonly observed in normal subjects. Additional genes that may be important in glaucoma pathogenesis have been investigated using

quantitative traits approaches. Such studies have begun to identify genes that control the magnitude of important quantitative features of glaucoma that may also be important risk factors

for POAG, such as central corneal thickness. Each of these different approaches to study glaucoma genetics is providing new insights into the pathogenesis of POAG.

Research breakthroughs have shown that genes have key roles in the pathogenesis of common eye diseases, including age-related macular degeneration,1, 2, 3, 4 Fuchs corneal endothelial

dystrophy,5 exfoliation syndrome,6 and primary open-angle glaucoma (POAG).7, 8, 9 The recent discovery of important risk factors for these common eye diseases underscores the utility of

studying ophthalmic genetics.

The identification of disease-causing genes provides information about the pathogenesis of heritable eye diseases at the most basic level. For example, disease-causing genes may be part of

important biological pathways that once identified may help clarify the mechanisms that lead to disease. The discovery of disease genes will also continue to provide insights into the normal

function of the eye.

Discovery of the genes that cause eye disease may also provide useful information for patients and their physicians. Identifying these genes will enable the design of DNA-based tests that

may help physicians assess their patient's risk for disease and may also differentiate between clinically similar disorders. Many such tests are already available on both a fee-for-service

and research basis (http://www.genetests.org). Identification of the specific mutation or mutations that are responsible for a patient's disease not only solidifies the diagnosis, but may

also help predict its likely clinical course. Several mutation-specific phenotypes of hereditary eye diseases have already been reported, including glaucoma,10 retinitis pigmentosa,11 and

Von Hippel Lindau syndrome.12 Genetic variations may also influence a patient's response to therapeutic interventions and will help guide selection of their clinical and surgical care.

Discovery of the genes that cause disease is a vital step in the development of new treatments for heritable eye conditions. The biological function of a disease-causing gene may in some

cases suggest the application of currently available medical and surgical therapies. In other cases, new interventions may be developed to compensate a genetic defect after it is identified.

Such gene-directed therapies might include currently available or newly designed medications, gene therapy (replacing a mutant gene with a normal copy), and/or other molecular genetic

approaches such as blocking mutant gene expression.13, 14

Many eye diseases, including glaucoma, are both genetically and mechanistically heterogeneous, meaning that it is unlikely that one therapy will be effective for all forms of a disease.

Genetic studies of complex diseases may also provide crucial information for future animal and clinical treatment trials. The most relevant animal models of eye disease will be those that

are designed to have the same genetic defects that are found in human disease. Such animal models would have great value for testing both the safety and efficacy of new therapies.

Additionally, researchers may use genetic tests to identify relatively homogeneous populations of study patients for treatment trials or to study patients with the same molecular cause of

disease.

The idea that heredity has an important role in glaucoma pathogenesis is not new. Some of the earliest evidence came from reports of large pedigrees in which glaucoma was passed down from

generation to generation in a Mendelian pattern that demonstrated that at least some cases of glaucoma have a genetic basis. Twin studies and familial clustering studies have also indicated

that some portion of glaucoma is caused by heredity.15, 16, 17, 18, 19, 20 Finally, several domesticated animal breeds, including the DBA/2J mouse (pigmentary glaucoma), cats, and dogs, have

been documented to be afflicted by inherited glaucoma.21, 22, 23, 24, 25 Together these data provide strong evidence that genes have an important role in the pathogenesis of glaucoma and

provide support for efforts to find these genes.

In the last two decades significant progress has been made in unraveling the genetic basis of POAG. Mutations that lead to POAG can be divided into two groups with very distinct

characteristics (Table 1). One class of mutations are capable of ‘causing’ POAG on their own with little influence from other genes or the environment. These single-gene forms of glaucoma

are responsible for disease that is transmitted as a Mendelian trait, often with an autosomal dominant inheritance pattern. Individuals that carry these types of mutations almost always

develop POAG and these mutations are rarely observed in subjects with normal eyes. Mutations in the myocilin (MYOC) and optineurin (OPTN) genes are examples of mutations that ‘cause’ POAG

and are discussed in more detail below. The other class of mutations, ‘risk alleles’, may promote the development of POAG when combined with other glaucoma risk alleles and environmental

factors but do not cause disease on their own (Table 1). These alleles or genetic risk factors are statistically more common in POAG patients, although they are very frequently detected in

both patients and controls. In this report, the current state of our knowledge of the genetic basis for glaucoma is reviewed, including glaucoma causing genes, glaucoma genetic risk factors,

and quantitative traits related to the development of glaucoma.

Myocilin was the first gene to be associated with POAG, and mutations in this gene are the most common cause of glaucoma with a known molecularly defined basis. Glaucoma-causing mutations in

myocilin were first detected in linkage-based studies of large pedigrees with juvenile open angle glaucoma (JOAG).7, 27 Myocilin mutations have since been found in 3–4% of POAG patients7,

26 and a range of glaucoma-associated myocilin mutations have now been cataloged in review articles28, 29 and online at http://www.myocilin.com. In most cases, myocilin-associated glaucoma

appears to be transmitted as an autosomal dominant trait and is associated with markedly elevated IOP.

Myocilin and JOAG. Glaucoma-causing mutations have been identified in many JOAG pedigrees with glaucoma that is inherited as an autosomal dominant trait. Myocilin mutations are relatively

common in patients with JOAG, and have been detected in 8–36% of patients in published case series.30, 31, 32 The likelihood of detecting myocilin mutations in subjects with JOAG appears to

be greater in patients with stronger family histories of glaucoma than in patients with apparently sporadic glaucoma.

Myocilin and POAG. Myocilin mutations are also co-inherited with POAG in an autosomal dominant pattern and have been detected in cohorts of POAG patients from around the world. However, due

to the later onset of disease, the POAG pedigrees are smaller and the mode of inheritance is not as obvious as is observed with JOAG families.26 Many mutations are only observed in specific

geographic or ethnic populations, but one myocilin mutation (GLN368STOP) has been detected in nearly all of the examined POAG populations, including African-Americans and Caucasians from the

United States, Canada, Australia, Europe, and South America.28 This most common myocilin mutation has not, however, been observed in Asian glaucoma patients. Some studies have suggested

that a founder effect may be responsible for the high relative frequency of the GLN368STOP mutation in some populations.26

Myocilin genotype–phenotype correlations. Individual myocilin mutations have been associated with specific clinical features of myocilin-related glaucoma such as age of diagnosis, maximum

intraocular pressure (IOP), and response to medical therapy. The best correlations between genotype and phenotype come from the two classes of mutations that have been detected in most

glaucoma patients: myocilin mutations that cause JOAG and the GLN368STOP mutation. In addition to having an earlier age of onset, the myocilin mutations that cause JOAG are also associated

with higher IOP and greater resistance to medical therapies than mutations that cause POAG. Although a number of reports have shown that subjects with JOAG respond poorly to medical

treatment and generally require surgical interventions,33, 34 a study by Graul et al35 has shown that POAG patients with a GLN368STOP myocilin mutation have similar rates of laser

trabeculoplasty and surgery as POAG patients with no myocilin mutations. However, another study by Craig et al36 reported that glaucoma patients with GLN368STOP mutations had increased rates

of filtration surgery compared with patients with no myocilin mutations.

Myocilin and glaucoma pathogenesis. Myocilin was first identified as a glaucoma gene in 1997, but we still know little about the normal function of the protein it encodes. In health,

myocilin protein is produced by many cell types of the eye and is secreted into the aqueous humor for an unknown purpose. Studies of both cell culture and human tissue have shown that the

abnormal protein that is produced by myocilin mutations is poorly secreted and is retained within trabecular meshwork cells.37, 38, 39, 40 Accumulation of abnormal myocilin protein may be

toxic to trabecular meshwork cells and may lead to their dysfunction or death, which may ultimately produce decreased aqueous outflow, elevated IOP, and glaucoma.41, 42 Animal models

harboring myocilin mutations have been developed and are beginning to reveal the specific molecular steps that lead from mutations in myocilin to the elevated IOP that is characteristic of

myocilin-related glaucoma.43, 44, 45

Optineurin was identified as a glaucoma-causing gene through investigations of a large normal tension glaucoma (NTG) pedigree. The dominantly inherited glaucoma in this family was shown to

be caused by a GLU50LYS mutation in the optineurin gene.9, 46 Subsequently, most studies of optineurin in large populations of glaucoma patients have suggested that mutations in this gene

may be responsible for up to 1.5% of NTG cases.47, 48, 49 The strongest data linking mutations in optineurin with glaucoma are focused on the GLU50LYS mutation. The links between other

optineurin mutations and glaucoma are more complex, as these mutations are associated with glaucoma in some but not all populations. For example, the MET98LYS variant in optineurin is

statistically more common in NTG patients than control subjects in some Caucasian and Asian populations,47, 50, 51 but not in others.47, 52, 53 Overall, mutations in optineurin do not appear

to be associated with cases of POAG that have elevated IOP in most47, 48, 52, 54, 55, 56, 57 but not all populations.51, 55 The significance of some optineurin variations appears to depend

upon the population in which they are observed.

Optineurin and glaucoma pathogenesis. The mechanism by which optineurin causes glaucoma has been investigated using in vitro and in vivo studies. There is some evidence that optineurin may

have neuro-protective effects that are reduced or eliminated by disease-causing mutations. Overexpression of wild-type optineurin appears to provide some protection from apoptosis induced by

oxidative stress in a cell culture system. This protective effect is not observed with overexpression of mutant optineurin protein.58 More recently, experiments with transgenic mice have

demonstrated that the GLU50LYS mutation in optineurin leads to apoptosis of retinal ganglion cells. Further studies of these mice have suggested that optineurin-mediated glaucoma may result

from a disruption of an interaction between optineurin and a GTP-binding protein, Rab8, and its effects on protein trafficking.59

TBK1 encodes a kinase that regulates the expression of genes in the NF-κB signaling pathway. Copy number variations (duplications) that encompass the TBK1 gene were recently shown to be

associated with glaucoma through family-based studies.60 First, linkage analysis of a large African-American NTG pedigree mapped a new glaucoma gene to chromosome 12q14. Subsequent

investigations of this part of the genome demonstrated that all family members with glaucoma possessed a duplication of the TBK1 gene and some neighboring genes. This particular duplication

was never seen in control subjects. When a cohort of additional NTG patients were similarly tested for copy number variations, 2(1.3%) of 152 NTG patients were found to have unique but

overlapping duplications of chromosome 12q14 that also spanned the TBK1 gene.60 These data suggest that an extra copy of TBK1 leads to NTG and may be responsible for some fraction of

sporadic-appearing NTG cases.

The role of TBK1 in glaucoma pathogenesis is also supported by previous studies of optineurin, the only other known NTG gene. In 2008, Morton et al61 showed that the protein encoded by

optineurin interacts with TBK1 and that this interaction is influenced by an optineurin mutation (GLU50LYS) that was previously shown to be associated with glaucoma. This interaction with

optineurin, a known NTG gene, further supports a role for TBK1 in glaucoma pathogenesis.

Little is known about how copy number variations in TBK1 might cause glaucoma. The interaction between TBK1 and genes in the NF-κB signaling pathway may influence important processes that

are involved in the pathogenesis of glaucoma, including apoptosis and modulation of the immune system. Initial studies have shown that TBK1 is expressed in human retinal ganglion cells and

that duplication of this gene significantly alters its expression in cultured fibroblasts.60 Therefore, it is a plausible hypothesis that copy number variations of TBK1 cause a dysregulation

of NF-κB signaling that ultimately leads to apoptosis of retinal ganglion cells and the development of NTG.

Linkage analysis of two POAG pedigrees mapped a glaucoma gene to chromosome 5q22, and subsequent studies of genes in this locus suggested that mutations in WDR36 might cause some cases of

POAG.62 However, many subsequent studies failed to confirm this link.63, 64, 65 Furthermore, two additional POAG pedigrees have been identified with glaucoma that is linked to the same

chromosome 5q22 locus, but these pedigrees were found to harbor no WDR36 mutations. These results suggest the presence of a different glaucoma gene in the region.66, 67 Nonetheless, a few

studies have identified rare WDR36 variants that may be associated with POAG in some populations. As a result, there is continued controversy over the role of WDR36 in glaucoma pathogenesis.

Although WDR36 was initially identified in studies focused on genes that ‘cause’ Mendelian forms of POAG, some researchers have investigated the possibility that variants in this gene may

either contribute to the risk for developing complex, polygenic forms of glaucoma68, 69 or influence the severity of disease.63 The mechanism by which such common WDR36 variants might

contribute to the risk for POAG is unknown. However, some common variants of WDR36 have been shown to alter cell viability in yeast70 and axon growth in mouse retinal ganglion cells.71

Family-based studies have been successful in discovering a number of genes that are capable of causing POAG with minimal influence from other genes or the environment. However, the known

glaucoma-causing genes are together responsible for