**10. The potential role of epigenetics in glaucoma**

**8. Pigmentary dispersion syndrome, pigmentary glaucoma and Axenfeld-**

A number of ocular conditions such as pigment dispersion syndrome (PDS), Axenfeld-Rieg‐ er syndrome (ARS) can lead to secondary open-angle glaucoma. PDS affects the young peo‐ ple and is characterized by the presence of TM pigmentation, iris-transillumination defects, Krukenberg spindle and backward bowing of the iris [50]. It is transmitted in a direct linear manner from parent to sibling [51]. Genetic analysis revealed a homozygous mutation (C677T) in methylenetetrahydrofolate reductase gene (MTHFR) in a patient [52] and the higher level of plasma homocysteine was suggested to be associated with pigmentary glau‐ coma. Additionally, a gene responsible for the PDS has been mapped to chromosome 7q35 q36 [53]. Regarding pigmentary glaucoma, the risk of developing it from PDS is about 10% at 5 years. Young myopic men are most likely to develop the disorder [54]. Interestingly, PDS and pigmentary glaucoma are not associated with mutations in lysyl oxidase like-1 (LOXL1) and tyrosinase related protein-1 (TYRP1) genes [55-56]. Another anterior segment disease with the risk of developing congenital glaucoma is called ARS. It is a rare autosomal dominant disorder with genetic heterogeneity and exhibits a range of congenital malforma‐ tions of the anterior segment of the eye. In addition, patients with ARS may present system‐ ic malformations such as mild tooth abnormalities, craniofacial dysmorphism, sensory hearing loss and congenital heart defect. It is caused by mutations in paired-like homeodo‐ main 2 (PITX2) and forkhead box C1 (FOXC1) genes [57-61]. In the United States, it has been estimated that mutations in PITX2 and FOXC1 genes are associated with 25% - 30% cases of ARS [62]. In severely affected patients, digenic inheritance of mutations in PITX2 and

**9. Epigenetics: Three major types of epigenetic modifications**

A vast spectrum of epigenetic changes has been described. The most common epigenetic variations involve DNA methylation, various modifications of histones, microRNA (miR‐ NA) and small non-coding RNA expression. All these factors can modulate the expression of genes that in turn may affect phenotypes and response to drugs. DNA methylation may be tissue specific [64] and disrupts the transcriptional activity of genes by affecting the ac‐ cessibility of transcription factors. A large number of CpG residues are concentrated in a re‐ gion of DNA sequence (CpG island). Methylation of cytosine may reduce or prevent the binding of sequence specific transcription factors. This results in changes in gene expression. The CpG region methylation also regulates the expression of a large number of miRNA. On the other hand, genomic hypomethylation may lead to genome instability. This kind of epi‐ genetic abnormality can be influenced by environmental factors such as tobacco smoking, dioxin and nutrition [65] and can lead to complex disorders. Studies including monozygotic twins also suggest that non-Mendelian and complex diseases (including neurological and psychiatric disorders) are likely to be caused by the combination of genetic and epigenetic factors [66]. DNA methylation and its maintenance may depend upon chromatin-associated

**Rieger syndrome**

62 Glaucoma - Basic and Clinical Aspects

FOXC1 has also been reported [63].

The eye is a model organ for epigenetic studies because external ocular tissues are exposed to the outside environment and may be sensitive to epigenetic effects. Although the epigenetics is well known in diseases such as cancer [71], and hereditary and environmental determi‐ nants have been long suspected for eye disorders [72], epigenetic studies on eye disorders are slowly progressing [9; 73-74]. For instance, retinal and lens differentiation involves specific changes in DNA methylation, expression of non-coding RNA and nucleolar organization [73]. In addition, cell-specific DNA methylation may play an important role in modulating eye specific genes [64]. Similarly, histone modifications were involved in the pathologic course of retinal ganglion cells [75] and site-specific DNA hypomethylation permits the ex‐ pression of interphotoreceptor retinoid binding protein (IRBP) gene [76]. Overexpression of mutant OPTN (E50K) is also found to induce RGC apoptosis [77-78]. Recently, it was also shown that histone deacetylase 4 (HDAC4) was involved in the survival of retinal neurons by preventing apoptosis of rod photoreceptor and bipolar cells [79-80]. Additionally, histone acetyltransferase p300 was found to promote intrinsic axonal regeneration [81]. Similarly, in an animal model (rat/mice), it has been observed that there was a regional gene expression changes including pro-survival, pro-death and acute stress genes [82-84]. Moreover, miRNAs can act as either oncogenes or tumor suppressor genes and can influence the growth of uveal melanoma [85]. Similarly, smoking and nutritional factors were involved in the etiology of age-related macular degeneration (AMD) in addition to genetic susceptibility [65].

Another example to illustrate the epigenetic effect is the pseudoexfoliation syndrome (XFS), which is one of the most common subtypes of POAG. It is the major risk factor for secon‐ dary POAG. The condition is characterized by a pathological accumulation of the whitish material in the anterior segment of the eye, predisposing to glaucomatous optic neuropathy [86]. The disorder is frequent among Icelanders, increases with age and rarely identified in people below the age of 50. Mutations in the LOXL1 gene were found to be associated with XFS in the Caucasian Australian population. [87]. However, this does not account for the large difference in disease prevalence between different populations. This raises the possi‐ bility of unidentified genetic, racial and environmental modulators [88]. In support of this is the finding that XFS may be associated with geographic and climatic factors such as sun ex‐ posure and ambient temperature [89]. The mechanisms involved are not known at present. Retinal cell death, the most common pathophysiology of all forms of glaucoma involves many factors such as oxidative stress, mitochondrial dysfunction, excitotoxic damage, axo‐ nal transport failure, deprivation of neurotrophic factors and activation of intrinsic and ex‐ trinsic apoptotic signals [90-91]. Some of these could be modulated by epigenetic changes. In support of this is the finding that heavy smoking, exposure to pesticides and nutrient intake was significantly associated with POAG [92-94]. This suggests that the interaction between gene and environmental factors may play a role in the pathogenesis of glaucoma. Intrauter‐ ine exposure (obesity and diabetes), variable DNA methylation and environmental factors may also have profound influence on adult epigenetic status. Thus in general, epigenetic may provide an additional layer of important information on inherited as well as age-relat‐ ed eye disorders including glaucoma.

genotype alone. This missing link could be due to several factors including environmental factors such as chemicals, alcohol, tobacco, diet and other drugs. In addition, age and gender may contribute to the physiological and biochemical status of the targeted cells (with respect to gene expression). Therefore, it is not simply genetics or environment but it is the interplay

Emerging Concept of Genetic and Epigenetic Contribution to the Manifestation of Glaucoma

http://dx.doi.org/10.5772/52279

65

**Figure 2.** A schematic illustration of the relationship between genotype and drug response of an individual. Two hori‐ zontal lines 1 and 2 (panels A to C) denote a pair of homologous genes encoding a drug-metabolizing enzyme. In panel A, genes are normal and hence the individual is a fast responder and metabolizes the drug more efficiently. Therefore high doses are needed to treat. In panel B, the individual is heterozygous for the mutation and metabolizes the drug slowly. Therefore, lower doses are needed to avoid side effect or toxicity. In panel C, the individual is homozy‐ gous for the mutation and metabolizes the drug very poorly. Therefore, it may have fatal effect. The X mark denotes

Epigenetic is an emerging field in ophthalmology. One benefit of understanding epigenetic changes is at the level of treatment. Epigenetic modifications are reversible. For instance, disease associated DNA methylation can be reversed by inhibitors such as adenosine or deoxycytidine. However, these reagents might become cytotoxic and may lead to a wide spread DNA hypome‐ thylation that may be resulting in and causing destabilization of genome. We need to develop less toxic inhibitors of DNA mythyltransferases. Similarly, inhibitors of histone deacetylase (HDAC) may have some therapeutic applications. For instance, HDAC inhibitors have been found to have protective effects in animal model of ischemia and optic nerve damage in the retina [71, 102-103]. At present IOP is the only modifiable risk factor for the prevention or progression of glaucoma and low IOP is associated with reduced progression of visual field defect [104-105]. Recent devel‐ opment on stem-cell therapy may be interesting. The initial results of clinical trials in patients us‐ ing stem-cell therapy showed some visual benefits with no sign of tumorigenicity [106-111]. Therefore, stem-cell therapy may be a promising approach to treat patients with retinal disease in the future. However, further research will be needed and an understanding of the role of epige‐ netics is also important to the success of the stem cell-based therapies [8]. In the future, studies will uncover the epigenetic mechanism contributing to glaucoma. A strong emphasis must be placed on epigenetics in the analysis of complex phenotypic variation. It may be necessary to de‐ velop a human methylation map to understand the difference in transcript expression. Epigenet‐

between them that is important in pharmacology and medicine.

ic mechanisms in ophthalmology are truly exciting areas of research.

mutation.

**12. Concluding remarks**

### **11. Pharmacogenetics and pharmacoepigenetics in glaucoma**

Adverse drug reactions (ADRs) and individual variations in drug response were well known in medicine. There are many systemic and other drugs that produce adverse effects in eye care [95-96]. For instance, many steroid drugs induce glaucoma in some patients [97]. Therefore, efficacy and safety are important aspects of initiation of any medication. Present‐ ly, there are no biochemical markers (proteins or genes) to predict which group of patients develops ADR and which group does not. Physicians in all medical branches have to make a guessing game to find out, which medication will work best for a given patient. This trialand error method is often inefficient. Now because of the advancement in genetics, physi‐ cians will have better opportunities to treat individual patients based on their genotype (Fig. 2). In order to understand the relationship between genes and inter-individual variations in drug response, two related fields namely pharmacogenetics and pharmacogenomics have been developed. They have taken massive studies on genetic personalization of drug re‐ sponse [98]. Some of the pharmacogenetic studies that are related to eye disorders including glaucoma have been discussed previously [99-100]. For instance, heterozygosity in N363S mutation in glucocorticoid receptor gene has been found to be associated with steroid in‐ duced ocular hypertension in Hungarian population although it may not be the major risk factor in the pathogenesis of elevated IOP. Similarly, a beta-adrenergic antagonist timolol has been used for the treatment of glaucoma. However, a topically administered eye drop may cause adverse cardiovascular and respiratory effects. Recent investigation of a single nucleotide polymorphism (SNP) in beta-adrenergic receptor suggests that this polymor‐ phism may be associated with positive clinical response to topical beta-blockers. In addition, R296C polymorphism in CYP 2D6 (cytochrome P450) gene may confer susceptibility to tim‐ olol induced bradycardia. Patients with CC genotype were unlikely to suffer from timolol induced bradycardia and those with TT genotype were found to suffer. Many studies ad‐ dress the pharmacology of several glaucoma medications but it is still not possible to ex‐ plain the variable IOP response to glaucoma drugs between patients [101] using their genotype alone. This missing link could be due to several factors including environmental factors such as chemicals, alcohol, tobacco, diet and other drugs. In addition, age and gender may contribute to the physiological and biochemical status of the targeted cells (with respect to gene expression). Therefore, it is not simply genetics or environment but it is the interplay between them that is important in pharmacology and medicine.

**Figure 2.** A schematic illustration of the relationship between genotype and drug response of an individual. Two hori‐ zontal lines 1 and 2 (panels A to C) denote a pair of homologous genes encoding a drug-metabolizing enzyme. In panel A, genes are normal and hence the individual is a fast responder and metabolizes the drug more efficiently. Therefore high doses are needed to treat. In panel B, the individual is heterozygous for the mutation and metabolizes the drug slowly. Therefore, lower doses are needed to avoid side effect or toxicity. In panel C, the individual is homozy‐ gous for the mutation and metabolizes the drug very poorly. Therefore, it may have fatal effect. The X mark denotes mutation.
