Mutant frizzled-4 disrupts retinal angiogenesis in familial exudative vitreoretinopathy.

Familial exudative vitreoretinopathy (FEVR) is a hereditary ocular disorder characterized by a failure of peripheral retinal vascularization. Loci associated with FEVR map to 11q13-q23 (EVR1; OMIM 133780, ref. 1), Xp11.4 (EVR2; OMIM 305390, ref. 2) and 11p13-12 (EVR3; OMIM 605750, ref. 3). Here we have confirmed linkage to the 11q13-23 locus for autosomal dominant FEVR in one large multigenerational family and refined the disease locus to a genomic region spanning 1.55 Mb. Mutations in FZD4, encoding the putative Wnt receptor frizzled-4, segregated completely with affected individuals in the family and were detected in affected individuals from an additional unrelated family, but not in normal controls. FZD genes encode Wnt receptors, which are implicated in development and carcinogenesis. Injection of wildtype and mutated FZD4 into Xenopus laevis embryos revealed that wildtype, but not mutant, frizzled-4 activated calcium/calmodulin-dependent protein kinase II (CAMKII) and protein kinase C (PKC), components of the Wnt/Ca(2+) signaling pathway. In one of the mutants, altered subcellular trafficking led to defective signaling. These findings support a function for frizzled-4 in retinal angiogenesis and establish the first association between a Wnt receptor and human disease.

Retinal ganglion cell (RGC) death in glaucomatous beagles is not associated with mutations in p53 and NTF4 genes.

BACKGROUND: Glaucoma in humans is a second leading cause of irreversible vision loss in the world and can affect all age groups as well as all populations. The precise mechanism of retinal ganglion cell (RGC) death and progressive degeneration of optic nerve in glaucoma is not understood. It has been suggested that apoptosis is the common pathway that leads to the death of RGCs in glaucoma and that neurotrophin 4 (NTF4) protein plays a role in the protection of RGCs by activating tyrosine kinase receptors. Additionally, one previous study suggested that p53 codon 72 polymorphism (R72P) might have a greater susceptibility to apoptosis in some ethnic population. Glaucoma also occurs in dogs, and the primary glaucoma in beagles is inherited as an autosomal recessive trait. Although recently a candidate gene has been isolated, the mechanism underlying RGC death is not understood. METHOD: To understand whether the same p53 and NTF4 pathway mechanism is involved in a beagle model of glaucoma, we have isolated NTF4 gene from dog and analyzed both p53 and NTF4 genes for mutations in glaucomatous animals. RESULTS: Our analyses failed to identify any disease-causing mutations in both genes with the exception of two polymorphisms in NTF4 gene. However, these are not pathogenic changes because they are also present in normal animals and are not segregated with the disease. CONCLUSION: These results suggest that impaired neurotrophin signaling or compromised trophic support to the retina and p53-mediated apoptosis may not be the underlying mechanism of RGCs death in a beagle model of glaucoma.

Genetic susceptibility to advanced retinopathy of prematurity (ROP).

Retinopathy of prematurity (ROP) is a vascular vitreoretinopathy that affects infants with short gestational age and low birth-weight. The condition is a multifactorial disease and is clinically similar to familial exudative vitreoretinopathy (FEVR), which is a bilateral hereditary eye disorder affecting full-term infants. Both of them are characterized by the abnormal vessel growth in the vitreous that can lead to vitreoretinal traction, retinal detachment and other complications resulting in blindness. Despite the recent advances in diagnosis and treatment, ROP remains a major cause of childhood blindness in developed countries. The etiology of pathogenesis of advanced ROP is currently unknown. In the past, many causative factors such as length of time exposed to supplemental oxygen, excessive ambient light exposure and hypoxia have been suggested but evidence for these as independent risk factors in recent years is not compelling. It is not clear why ROP in a subset of infants with low birth-weight progresses to a severe stage (retinal detachment) despite timely intervention whereas in other infants with similar clinical characteristics ROP regresses spontaneously. Recent research with candidate gene approach, higher concordance rate in monozygotic twins and other clinical and experimental animal studies, suggest a strong genetic predisposition to ROP besides environmental factors such as prematurity. Three genes, which are involved in the Wnt signaling pathway, are mutated in both FEVR and in a small percentage of ROP disorder. However, none of the genetic factors identified thus far in ROP, account for a substantial number of patient population. Future studies involving genomics, bioinformatics and proteomics may provide a better understanding of the pathophysiology and management of ROP.

Genetic diversity and medicinal drug response in eye care.

BACKGROUND: Individual variation in drug response and adverse drug reactions are a serious problem in medicine. This inter-individual variation in drug response could be due to multiple factors such as disease determinants, environmental and genetic factors. Much has been published in the literature in recent years about the potential of pharmacogenetic testing and individualized medicine. The development of personalized medicine is truly an exciting area of research. METHODS: This pharmacogenetic concept in ophthalmology has existed for more than a century. Although substantial studies that link genetic variants to inter-individual difference in drug response have been reported in several diseases such as cancer and heart diseases, such studies are progressing slowly in the eye field. In this short article, an attempt has been made to summarize these results. RESULTS: Recently, there have been some small-scale studies that seem to associate the drug response to the genotype of patients in two major eye disorders, namely age-related macular degeneration (ARMD) and glaucoma. CONCLUSION: These studies are still in their infancy, and do not suggest that a pharmacogenetic basis of drug development is a credible concept and can become reality in the future. This is because most drug responses involve a large number of genes that have several polymorphisms and it is unlikely that any one single gene dictates the drug response. Therefore, a polygenic approach, whole genome single nucleotide polymorphism (SNP) analysis and a molecular understanding of disease itself may provide a better insight in the future about genetic predisposing factors for adverse drug reactions.

Persistent hyperplastic primary vitreous: congenital malformation of the eye.

Persistent hyperplastic primary vitreous (PHPV), also known as persistent fetal vasculature, is a rare congenital developmental malformation of the eye, caused by the failure of regression of the primary vitreous. It is divided into anterior and posterior types and is characterized by the presence of a vascular membrane located behind the lens. The condition can be of an isolated type or can occur with other ocular disorders. Most cases of PHPV are sporadic, but it can be inherited as an autosomal dominant or recessive trait. Inherited PHPV also occurs in several breeds of dogs and cats. In a limited number of cases, Norrie disease and FZD4 genes are found to be mutated in unilateral and bilateral PHPV. These genes when mutated also cause Norrie disease pseudoglioma and familial exudative vitreoretinopathy that share some of the clinical features with PHPV. Mice lacking arf and p53 tumour suppressor genes as well as Norrie disease pseudoglioma and LRP5 genes suggest that these genes are needed for hyaloid vascular regression. These experiments also indicate that abnormalities in normal apoptosis and defects in Wnt signalling pathway may be responsible for the pathogenesis of PHPV. Identification of other candidate genes in the future may provide a better understanding of the pathogenesis of the condition that may lead to a better therapeutic approach and better management.

Evaluation of the common variants of the ABCA4 gene in families with Stargardt disease and autosomal recessive retinitis pigmentosa.

Stargardt disease (STGD) is one of the most common autosomal recessive retinal dystrophies with an estimated incidence of one in 10,000. It affects the central retina (macula). Retinitis pigmentosa (RP) comprises a large and exceptionally heterogeneous group of hereditary disorders of the retina. It is caused by the loss of photoreceptors. The condition is a degenerative disorder characterized by retinal pigment deposits and has an estimated incidence of one in 4,000. Although, to date, 45 known loci have been identified, none of them independently account for a substantial portion of RP. Recently, the photoreceptor cell-specific ATP-binding cassette transporter (ABCA4) gene was found to be mutated in patients with STGD as well as autosomal recessive RP. In order to further understand the contribution of this gene to the susceptibility to STGD and RP, we analyzed three unrelated STGD families and one autosomal recessive RP family specifically for the more common variants (A1038V, G1961E, 2588G-->C, R943Q or 2828G-->A) in the ABCA4 gene. Our analyses employing standard techniques such as polymerase chain reaction, restriction fragment length polymorphism, and direct DNA sequencing of amplified products were able to identify one common variant (R943Q) in all three STGD families but not in the RP family. All three affected STGD individuals, however, were heterozygous for this variation, and this alteration did not segregate with the disease and was also present in the normal controls. Similar analysis of other common variants revealed no pathogenic mutations in the STGD and RP families. It is likely that the variant identified in this study represents a rare polymorphism (non-pathogenic). Although, at present we cannot eliminate the possibility of this gene as a candidate gene, future extensive studies on this as well as other candidate genes may uncover the susceptibility gene for these recessive forms of the disorders in these families.

Bipolar disorder: an update.

Bipolar disorder (BPD) is one of the most severe forms of mental illness and is characterized by swinging moods. It affects both sexes equally in all age groups and its worldwide prevalence is approximately 3-5%. The clinical course of illness can vary from a mild depression to a severe form of mania. The condition has a high rate of recurrence and if untreated, it has an approximately 15% risk of death by suicide. It is the third leading cause of death among people aged 15-24 years and is a burden on society and families. The pathophysiology of the disorder is poorly understood. However, a variety of imaging studies suggests the involvement of structural abnormalities in the amygdala, basal ganglia and prefrontal cortex. There are two main biological models that have been proposed for depression. These are called the serotonin and norepinephrine hypotheses. Multiple lines of evidence support both of them. It is a life-long disease and runs in families but has a complex mode of inheritance. Family, twin and adoption studies suggest genetic factors but the candidate susceptibility genes, which when mutated can account for a substantial portion of BPD patients, have not yet been conclusively identified. There have been an increasing number of new generation antidepressant drugs developed to treat BPD. However, lithium salt is only the drug that is most efficient in long-term preventive treatment and it also has an anti-suicidal effect. The condition can be well managed by physicians and psychiatrists along with family and patient education. Identification of risk genes in the future may provide a better understanding of the nature of pathogenesis that may lead to a better therapeutic target.

Neurodegenerative disorders of protein aggregation.

In recent years, it has become increasingly clear that many neurodegenerative diseases involve aggregation and deposition of misfolded proteins such as amyloid beta, tau, alpha-synuclein and polyglutamine containing proteins. This abnormal deposition of misfolded proteins produce malfunctioning of a distinctive set of neurons. It may also induce oxidative and endoplasmic reticulum stress and proteosomal and mitochondrial dysfunction that ultimately leads to neuronal death. While hereditary forms of disorders are caused by genetic mutations, many sporadic cases are likely to be due to genetic and environmental factors. These disorders are progressive in nature. Therefore, treatment is difficult. However, for some diseases, a growing number of treatment options such as drugs, antioxidants, cell transplantation, surgery, rehabilitation procedures and preimplantation diagnosis is available. It should be noted that many of these treatments produce unacceptable risks or adverse effects and they are of only minimal benefit for patients. In future, an understanding of the causes of protein aggregation and genetic and environmental susceptibility factors of a specific individual (or specific individual determinants) may provide a better opportunity for an effective therapeutic intervention.

Molecular genetics of attention-deficit hyperactivity disorder (ADHD): an update.

Attention-deficit hyperactivity disorder (ADHD) is a heritable and behavioral condition of childhood, affecting 5-10% of school-age children worldwide. Affected patients exhibit various behavioral problems such as carelessness, restlessness, disobedience and failure to stay quiet in class. The etiology of ADHD is not known. However, family, twin and adoption studies have provided strong evidence for a genetic etiology of the disorder. A genome-wide scan has identified six chromosomal loci with LOD scores suggestive of linkage. Animal studies suggest the involvement of the brain dopamine pathway and its alteration in ADHD but there is no direct evidence to support this hypothesis. In addition, there are at least 20 candidate genes of small effect that have been studied but none of them appear to be the major gene causing ADHD. Medical intervention along with psychosocial therapy proved to be beneficial for controlling ADHD, although some undesirable side effects have been encountered during medical treatment. In the future, identification of environmental factors, study of additive gene effects and the interaction of genes and environmental factors may provide better insight into the pathophysiology of ADHD. This may lead to an effective new treatment strategy.

Evaluation of the ABCR and glutathione peroxidase-3 genes in familial and sporadic cases of exudative age-related macular degeneration.

Age-related macular degeneration (ARMD) is the most common cause of blindness in older patients and is a major health care epidemic in developed countries. The exact cause of ARMD is not known. It has been recently reported that heterozygous missense ABCR mutations are associated with age-related macular degeneration. In addition, one of the susceptible loci for maculopathy is on chromosome 5 that is very close to the plasma glutathione peroxidase (GPX) gene. Since the retina is highly sensitive to peroxidation and the GPX gene product protects cells from oxidative damage, and the fact that the ABCR gene is considered as a major disease gene in macular degeneration we reasoned that they might serve as candidate genes in a subset of ARMD cases. To test our hypothesis, we have carried out a pilot study by analyzing 8 exudative ARMD patients for allelic variations in the GPX gene and three statistically significant mutations in the ABCR gene (R943Q, G1961E and D2177N). Our analysis failed to identify the above three major alterations in the ABCR gene as well as mutations in the coding sequence of the GPX gene. However, we have been able to identify two polymorphic heterozygous mutations in the promoter region of the GPX gene in one sporadic patient. These mutations have not been seen in any other patients. On the other hand, when 28 individuals from 6 different ethnic backgrounds with no evidence of ARMD were analyzed, four of them showed the same alterations in the GPX promoter region. Although we cannot completely exclude the possibility of alterations in the coding regions of the GPX gene, the promoter mutation identified in the present study statistically may not be associated with this disease. However, it may be associated with other additive factors as might be expected for a complex disorder.

SNPs and haplotypes: genetic markers for disease and drug response (review).

The genetic make-up of an individual not only determines disease susceptibility but also response to drug treatment. It is because of this reason tremendous progress has recently been made in cataloging human sequence variations with an intention such that a high-density map will provide necessary tools to develop genetic-based diagnostic and therapeutic options. The availability of the human DNA sequence, its variation between individuals and the functional understanding of genetic determinants between individuals may enable pharmaceutical companies to discover safer and effective drugs. This will provide physicians with better tools to select the most appropriate treatment for each patient and avoid drugs that cause adverse effects. When a personalized medication becomes a more realistic and beneficial treatment, every newborn child in the neonatal unit will probably be genotyped as a routine procedure for improved treatment. This newly developed toxicogenomic or pharmacogenomic field is rapidly advancing and may change the practice of medicine in the future, providing personalized medicine for each individual.

Schizophrenia: a genetic perspective (review).

Schizophrenia is a complex neurodevelopmental disorder characterized by mental dysfunction across multiple domains of the brain. It affects 1% of world's general population and the nature of neurobiological lesions in the schizophrenic brain are not known. Although the exact etiology of the disorder is not understood, twin, family and adoption studies have provided consistent evidence that genetic factors play a major role in the pathogenesis. A genome-wide genetic linkage screen identified loci on chromosomes 1, 2, 4, 5, 6, 7, 8, 9, 10, 11, 13, 15, 18, 22 and the X with positive lod scores, thus excluding a single major locus for schizophrenia. Association studies have generated disappointing results in identifying the susceptible DNA sequence variants and the anticipation hypothesis on trinucleotide repeat expansion provided equivocal results or lack of enthusiasm. Although there are no biological markers at present, the recent finding that human endogenous retrovirus is activated in cerebrospinal fluid as well as in the postmortem schizophrenic brain may change our understanding of the etiopathogenesis of this disease. Meanwhile, treatment with newly developed anti-psychotic drugs combined with educational and cognitive rehabilitation procedure may help the patients to cope with the illness.

Pharmacogenomics and pharmacoepigenomics in pediatric medicine.

In the past several years, human genetics studies have progressed from monogenic to complex and common diseases because of the advancement in technologies. There is increased knowledge of the pharmacokinetics and pharmacogenomics of the drugs in adults as well as in children. These technological developments provided new diagnostic, prognostic, and therapeutic opportunities. We are now in a position to address many additional ambitious questions. For instance, in clinical medicine, interindividual variation in drug response is a major problem. Some of the heterogeneity of drug safety and efficacy among individuals can be explained by pharmacogenomics. It has also the potential to improve the treatment in both adults and children. In pediatrics however, there is ontogeny and metabolic capacity in children is different compared to adults. Several specific developmental changes may underlie some of the variability in drug response seen in children. They may also be responsible for adverse drug reactions (ADRs). Therefore, much of the diversity in drug effects cannot be explained by studying the genomic diversity alone. It is necessary to include the effect of growth (involves variations in gene expression) along with genetic differences when explaining the variability in treatment response. In this respect epigenomics may expand the scope of pharmacogenomics towards optimization of drug therapy. Future studies must focus on periods of maturation of the drug-metabolizing enzymes and polymorphisms in their genes by using candidate gene approach, gene expression analysis, genome-wide haplotype mapping, and proteomics. The integration of genetic data and clinical phenotypes along with the role of other factors is necessary to evaluate both efficacy and ADRs of any drug. It may require extensive genetic epidemiological studies spanning over many years.

Genetic susceptibility to primary angle closure glaucoma (PACG).

Glaucoma is a group of heterogeneous optic neuropathy and is the second leading cause of irreversible blindness worldwide. The two most common clinical types of glaucoma include primary open-angle (POAG) and primary angle-closure glaucoma (PACG). PACG is characterized by the closure of angles between iris and trabecular meshwork (iridocorneal angles) mainly because of anatomic abnormalities. The condition is more prevalent in Chinese, Asian Indians, and Eskimos. Because of an unusually high incidence of PACG among siblings of affected patients, it was suggested that genetic factors were involved in its pathology and the action of a large number of grouped or independently inherited genes along with environmental factors result in anatomical abnormalities of PACG. In PACG, the genetic basis is not well understood. Genome-wide association studies have identified several candidate genes in relation to PACG in several different populations. However, they are not reproduced from population to population or the results are controversial. This may indicate that the involvement of genetic abnormality in the pathogenesis of PACG is complex. The availability of spontaneously occurring large animal models such as dogs may provide an opportunity to identify genes responsible for the pathophysiology of PACG in the future. This article summarizes the current status of genetic investigations on PACG which is the most common cause of blindness worldwide.

Pharmacogenomics and its importance in pediatric medicine.

Individual variation in drug response and adverse drug reactions (ADRs) are a serious problem in medicine. This individual variation in drug response could be due to multiple factors but there is strong evidence that genetic factors play a significant role in drug response variability and toxicity. Although substantial studies that link genetic variants to inter-individual difference in drug response in adults have been reported, such studies are comparatively rare in pediatric medicine. The ultimate goal of medical research is to improve human health in every disease and every patient. Many diseases such as asthma, autism, epilepsy, juvenile rheumatoid arthritis and attention-deficit hyperactivity disorder develop during childhood. Human development is a rapidly changing process. In children, there are differences in absorption, distribution, excretion and metabolizing capabilities of a drug compared with adults. Therefore, many pharmacological and toxicological actions of drugs in children are not predictable from adult experience. It is also possible that children may experience a different range of ADRs that may have long-term implications for their development. Therefore, an improved understanding of the drug transformation pathways for all age groups is necessary. Such studies could provide insight into the susceptibility of a child to ADRs. The availability of the complete sequence of human genome and the biochip technology may help in identifying the polymorphic variations in drug related genes. In this regard, pharmacogenetic and pharmacogenomic studies may play an important role in providing markers of increased risk or susceptibility. Based on this genetic information, children at risk can be identified before therapy is initiated and pediatric ADRs may be minimized. In this short article, an attempt has been made to emphasize the importance of pharmacogenomics in pediatrics.

Pharmacogenomics in ophthalmology.

Inter-individual variation in drug response and adverse drug reactions (ADRs) are well known in medicine. This individual variation in drug response could be at least, in part, due to genetic diversity among individuals. Although substantial studies that connect genetic variants to inter-individual variation in drug response have been documented in several diseases such as cancer and heart diseases, such studies are slowly progressing in ophthalmology. In recent years, advancement in technologies has led to the identification of genes associated with several eye disorders. At the same time, some small-scale studies have demonstrated the association of various genotypes or haplotypes with response to drug therapies. However, its integration into clinical practice in ophthalmology is not possible at present. This is because there are many challenging questions that remain to be addressed. For instance, in the case of complex disorders a single gene study is not enough. Multiple genes, environmental factors, multiple single nucleotide polymorphisms (SNPs), and rare or low frequency variants may contribute to the disease and they must be considered. The functional aspects of many genetic variants are not known. This raises questions of their biological importance and their clinical usefulness. In addition, there are legal, ethical, and social issues that need to be regulated. Moreover, physicians and patients must be educated about the limitation and sensitivity of genetic testing. At present pharmacogenetic studies in ophthalmology are still in their infancy and do not suggest that a pharmacogenetic basis of drug development in ophthalmology is a concept that can yield immediate results, but can become a reality in the future. In this article an attempt has been made to summarize some of the recent small-scale pharmacogenetic studies on two major eye disorders, age-related macular degeneration (AMD) and glaucoma.

Common polymorphisms of the CFH, LOC 387715/ARMS2 and HTRA1 genes may not influence the intra-familial variability of X-linked juvenile retinoschisis.

X-linked juvenile retinoschisis (XLRS) is the leading cause of juvenile macular degeneration in males and is rare in females. Previous studies have shown that there is a marked intra- and inter-familial variation in disease severity and progression. This suggests that additional factors, such as genetic modifiers and environmental elements, influence disease severity. In order to understand the contribution of genetic modifiers, we aimed to ascertain whether common variants of the CFH, LOC 387715/ARMS2 and HTRA1 genes, which are major risk factors in age-related macular degeneration, contribute to the phenotypic variability of the XLRS disorder. Two unrelated XLRS families were selected, one harboring the missense mutation and the second a nonsense mutation in the RS gene. Both families exhibited variations in clinical phenotype. Genomic DNA from family members were analyzed for the above three genes using the polymerase chain reaction-based restriction fragment length polymorphism method. Our analyses revealed that both families were wild-type with respect to the LOC 387715/ARMS2 and HTRA1 genes. In one family (but not the other), the most severely affected and unaffected individuals were heterozygous for the CFH polymorphisms, while the less severely affected individual was wild-type. However, this alteration did not necessarily influence disease severity. Although we cannot completely rule out the role of the above genes in determining the phenotypic variability of the disorder, and though the statistical significance of the results could not be assessed due to the small scale of the study, it is unlikely that common polymorphisms of the CFH, LOC 387715/ARMS2 and HTRA1 genes serve as disease modifiers of the XLRS disorder.