Autosomal Dominant Retinal Dystrophies Caused by a Founder Splice Site Mutation, c.828+3A>T, in PRPH2 and Protein Haplotypes in trans as Modifiers.

PurposeWe determined the phenotypic variation, disease progression, and potential modifiers of autosomal dominant retinal dystrophies caused by a splice site founder mutation, c.828+3A>T, in the PRPH2 gene.MethodsA total of 62 individuals (19 families) harboring the PRPH2 c.828+3A>T mutation, had phenotype analysis by fundus appearance, electrophysiology, and visual fields. The PRPH2 haplotypes in trans were sequenced for potential modifying variants and generalized estimating equations (GEE) used for statistical analysis.ResultsSeveral distinct phenotypes caused by the PRPH2 c.828+3A>T mutation were observed and fell into two clinical categories: Group I (N = 44) with mild pattern dystrophies (PD) and Group II (N = 18) with more severe cone-rod dystrophy (CRD), retinitis pigmentosa (RP), and central areolar chorioretinal dystrophy (CACD). The PRPH2 Gln304-Lys310-Asp338 protein haplotype in trans was found in Group I only (29.6% vs. 0%), whereas the Glu304-Lys310-Gly338 haplotype was predominant in Group II (94.4% vs. 70.4%). Generalized estimating equations analysis for PD versus the CRD/CACD/RP phenotypes in individuals over 43 years alone with the PRPH2 haplotypes in trans and age as predictors, adjusted for correlation within families, confirmed a significant effect of haplotype on severity (P = 0.03) with an estimated odds ratio of 7.16 (95% confidence interval [CI] = [2.8, 18.4]).ConclusionsThe PRPH2 c.828+3A>T mutation results in multiple distinct phenotypes likely modified by protein haplotypes in trans; the odds of having the CACD/RP-like phenotype (versus the PD phenotype) are 7.16 times greater with a Glu304-Lys310-Gly338 haplotype in trans. Further functional studies of the modifying haplotypes in trans and PRPH2 splice variants may offer therapeutic targets

North Carolina macular dystrophy (MCDR1) caused by a novel tandem duplication of the PRDM13 gene

PURPOSE: To identify the underlying cause of disease in a large family with North Carolina macular dystrophy (NCMD). METHODS: A large four-generation family (RFS355) with an autosomal dominant form of NCMD was ascertained. Family members underwent comprehensive visual function evaluations. Blood or saliva from six affected family members and three unaffected spouses was collected and DNA tested for linkage to the MCDR1 locus on chromosome 6q12. Three affected family members and two unaffected spouses underwent whole exome sequencing (WES) and subsequently, custom capture of the linkage region followed by next-generation sequencing (NGS). Standard PCR and dideoxy sequencing were used to further characterize the mutation. RESULTS: Of the 12 eyes examined in six affected individuals, all but two had Gass grade 3 macular degeneration features. Large central excavation of the retinal and choroid layers, referred to as a macular caldera, was seen in an age-independent manner in the grade 3 eyes. The calderas are unique to affected individuals with MCDR1. Genome-wide linkage mapping and haplotype analysis of markers from the chromosome 6q region were consistent with linkage to the MCDR1 locus. Whole exome sequencing and custom-capture NGS failed to reveal any rare coding variants segregating with the phenotype. Analysis of the custom-capture NGS sequencing data for copy number variants uncovered a tandem duplication of approximately 60 kb on chromosome 6q. This region contains two genes, CCNC and PRDM13. The duplication creates a partial copy of CCNC and a complete copy of PRDM13. The duplication was found in all affected members of the family and is not present in any unaffected members. The duplication was not seen in 200 ethnically matched normal chromosomes. CONCLUSIONS: The cause of disease in the original family with MCDR1 and several others has been recently reported to be dysregulation of the PRDM13 gene, caused by either single base substitutions in a DNase 1 hypersensitive site upstream of the CCNC and PRDM13 genes or a tandem duplication of the PRDM13 gene. The duplication found in the RFS355 family is distinct from the previously reported duplication and provides additional support that dysregulation of PRDM13, not CCNC, is the cause of NCMD mapped to the MCDR1 locus

Safety Assessment of Docosahexaenoic Acid in X-Linked Retinitis Pigmentosa: The 4-Year DHAX Trial

Citation: Hughbanks-Wheaton DK, Birch DG, Fish GE, et al. Safety assessment of docosahexaenoic acid in X-linked retinitis pigmentosa: the 4-year DHAX trial. Invest Ophthalmol Vis Sci. 2014;55:4958-4966. DOI: 10.1167/iovs.14-14437 PURPOSE. Docosahexaenoic acid (DHA) continues to be evaluated and recommended as treatment and prophylaxis for various diseases. We recently assessed efficacy of high-dose DHA supplementation to slow vision loss in patients with X-linked retinitis pigmentosa (XLRP) in a randomized clinical trial. Because DHA is a highly unsaturated fatty acid, it could serve as a target for free-radical induced oxidation, resulting in increased oxidative stress. Biosafety was monitored during the 4-year trial to determine whether DHA supplementation was associated with identifiable risks. METHODS. Males (n ¼ 78; 7-31 years) meeting entry criteria were enrolled. The modified intent-to-treat cohort (DHA ¼ 33; placebo ¼ 27) adhered to the protocol ‡ 1 year. Participants were randomized to an oral dose of 30 mg/kg/d DHA or placebo plus a daily multivitamin. Comprehensive metabolic analyses were assessed for group differences. Treatment-emergent adverse events including blood chemistry metabolites were recorded. RESULTS. By year 4, supplementation elevated plasma and red blood cell-DHA 4.4-and 3.6-fold, respectively, compared with the placebo group (P < 0.00001). Over the trial duration, no significant differences between DHA and placebo groups were found for vitamin A, vitamin E, platelet aggregation, antioxidant activity, lipoprotein cholesterol, or oxidized LDL levels (all P > 0.14). Adverse events were transient and not considered severe (e.g., gastrointestinal [GI] irritability, blood chemistry alterations). One participant was unable to tolerate persistent GI discomfort. CONCLUSIONS. Long-term, high-dose DHA supplementation to patients with XLRP was associated with limited safety risks in this 4-year trial. Nevertheless, GI symptoms should be monitored in all patients taking high dose DHA especially those with personal or family history of GI disturbances. (ClinicalTrials.gov number, NCT00100230.) Keywords: biosafety, fatty acids, adverse events R etinitis pigmentosa (RP) is a retinal degenerative disease characterized by night blindness and visual field constriction 1 with four underlying inheritance patterns. The X-linked form of RP (XLRP) is among the most severe with night blindness often detectable by age 5 years 2-4 and legal blindness by the second or third decade. Gene defects are known to cause retinal degeneration, yet factors such as environment, diet, stress, and/or metabolism may modify disease severity. Many patients with RP have lower plasma and red blood cell (RBC) levels of the n3 polyunsaturated fatty acid docosahexaenoic acid (DHA; 22:6n3) than normally-sighted controls. 5 Blood DHA was significantly correlated with age-adjusted ERG responses in XLRP such that patients with lower RBC-DHA tended to have lower ERG amplitudes. 6 These findings were similarly documented in approximately 70% of female XLRP carriers (Hoffman DR, et al. IOVS 1998;39:ARVO Abstract 725). A reduction in DHA biosynthesis was demonstrated in XLRP using stable isotopes to assess in vivo metabolism 7 suggesting that downregulation of hepatic D 5 desaturase may contribute to subnormal blood DHA levels. Thus, daily supplementation with DHA may bypass any decrease in DHA biosynthesis. Docosahexaenoic acid comprises 1% to 5% of membrane fatty acids in most human tissues; however, it is the most abundant fatty acid in the retina. 8 This n3 fatty acid can increase membrane fluidity and modify the mobility of vital proteins and activities of retinal enzymes, 9,10 promote photoreceptor differentiation, 11 and antiapoptotic activity. 12 The highly unsaturated nature of DHA makes it a potential target for free radical oxidative damage. Increased polyunsaturated fatty acid (PUFA) intake, particularly long-chain PUFAs (LCPUFAs; >18 carbons), may lead to elevated oxidative stress and subsequent membrane damage. 15 Numerous n3-supplementation studies report elevations in low-and highdensity lipoprotein (LDL and HDL)-cholesterol

Genetics Mutations in RPGR and RP2 Account for 15% of Males with Simplex Retinal Degenerative Disease

PURPOSE. To determine the proportion of male patients presenting simplex retinal degenerative disease (RD: retinitis pigmentosa [RP] or cone/cone-rod dystrophy [COD/CORD]) with mutations in the X-linked retinal degeneration genes RPGR and RP2. METHODS. Simplex males were defined as patients with no known affected family members. Patients were excluded if they had a family history of parental consanguinity. Blood samples from a total of 214 simplex males with a diagnosis of retinal degeneration were collected for genetic analysis. The patients were screened for mutations in RPGR and RP2 by direct sequencing of PCR-amplified genomic DNA. RESULTS. We identified pathogenic mutations in 32 of the 214 patients screened (15%). Of the 29 patients with a diagnosis of COD/CORD, four mutations were identified in the ORF15 mutational hotspot of the RPGR gene. Of the 185 RP patients, three patients had mutations in RP2 and 25 had RPGR mutations (including 12 in the ORF15 region). CONCLUSIONS. This study represents mutation screening of RPGR and RP2 in the largest cohort, to date, of simplex males affected with RP or COD/CORD. Our results demonstrate a substantial contribution of RPGR mutations to retinal degenerations, and in particular, to simplex RP. Based on our findings, we suggest that RPGR should be considered as a first tier gene for screening isolated males with retinal degeneration. (Invest Ophthalmol Vis Sci. 2012;53:8232-8237

Allelic heterogeneity and genetic modifier loci contribute to clinical variation in males with X-linked retinitis pigmentosa due to RPGR mutations.

Mutations in RPGR account for over 70% of X-linked retinitis pigmentosa (XlRP), characterized by retinal degeneration and eventual blindness. The clinical consequences of RPGR mutations are highly varied, even among individuals with the same mutation: males demonstrate a wide range of clinical severity, and female carriers may or may not be affected. This study describes the phenotypic diversity in a cohort of 98 affected males from 56 families with RPGR mutations, and demonstrates the contribution of genetic factors (i.e., allelic heterogeneity and genetic modifiers) to this diversity. Patients were categorized as grade 1 (mild), 2 (moderate) or 3 (severe) according to specific clinical criteria. Patient DNAs were genotyped for coding SNPs in 4 candidate modifier genes with products known to interact with RPGR protein: RPGRIP1, RPGRIP1L, CEP290, and IQCB1. Family-based association testing was performed using PLINK. A wide range of clinical severity was observed both between and within families. Patients with mutations in exons 1-14 were more severely affected than those with ORF15 mutations, and patients with predicted null alleles were more severely affected than those predicted to make RPGR protein. Two SNPs showed association with severe disease: the minor allele (N) of I393N in IQCB1 (p = 0.044) and the common allele (R) of R744Q in RPGRIP1L (p = 0.049). These data demonstrate that allelic heterogeneity contributes to phenotypic diversity in XlRP and suggest that this may depend on the presence or absence of RPGR protein. In addition, common variants in 2 proteins known to interact with RPGR are associated with severe disease in this cohort

Founder Effect of a c.828+3A>T Splice Site Mutation in Peripherin 2 (PRPH2) Causing Autosomal Dominant Retinal Dystrophies

ImportanceScreening for splice site mutation c.828+3A>T in the peripherin 2 (PRPH2) gene should be a high priority in families with highly variable retinal dystrophies. The correction of missplicing is a potential therapeutic target.ObjectiveTo determine the prevalence, genetic origin, and molecular mechanism of a donor c.828+3A>T mutation in the PRPH2 (peripherin 2, retinal degeneration slow) gene in individuals with retinal dystrophies.Design, setting, and participantsCase-control study that took place at the University of Texas Health Science Center, the University of Iowa, and the Retina Foundation of the Southwest, from January 1, 1987, to August 1, 2014, including affected individuals from 200 families with a diagnosis of autosomal dominant retinitis pigmentosa, 35 families with unspecified macular dystrophies, and 116 families with pattern dystrophy. Participants were screened for the c.828+3A>T mutation by restriction-enzyme digest, single-strand conformational polymorphism screening, or bidirectional sequencing. Haplotypes of polymorphic markers flanking the PRPH2 locus and sequence variants within the gene were determined by denaturing gel electrophoresis or automated capillary-based cycle sequencing. The effect of the splice site mutation on the PRPH2 transcript was analyzed using NetGene2, a splice prediction program and by the reverse transcription polymerase chain reaction of illegitimate transcripts from peripheral white blood cells.Main outcomes and measuresResults of testing for splice site mutation, haplotypes, and alternate transcripts.ResultsThe PRPH2 mutation was found in 97 individuals of 19 independently ascertained families with a clinical diagnosis of retinitis pigmentosa, macular dystrophy, and/or pattern dystrophy. All affected individuals also shared a rare haplotype of approximately 644 kilobase pairs containing the c.828+3A>T mutation, which extends from the short tandem repeat polymorphism D6S282 to c.1013G>A (rs434102, a single-nucleotide polymorphism) in exon 3 of PRPH2, suggesting this mutation is from a common ancestor and is a founder mutation. It has a prevalence of 2% in families diagnosed as having autosomal dominant retinitis pigmentosa and 10% in families with variable clinical diagnosis of pattern, macular, and retinal dystrophies. Individuals with the c.828+3A>T mutation expressed a PRPH2 transcript not found in control participants and that was consistent with abnormal splicing.Conclusions and relevanceThe PRPH2 c.828+3A>T splice site mutation is a frequent cause of inherited retinal dystrophies and is owing to the founder effect. The likely cause of disease is the missplicing of the PRPH2 message that results in a truncated protein product. Identifying the genetic etiology assists in more accurate management and possible future therapeutic options

Founder Effect of a c.828+3A>T Splice Site Mutation in Peripherin 2 (PRPH2) Causing Autosomal Dominant Retinal Dystrophies

O acesso à informação através da internet ganha a cada dia mais espaço em nossa sociedade, inclusive entre crianças e adolescentes. O estudo teve como objetivo identificar tendências de pesquisas sobre o uso seguro da informação on line por crianças e adolescentes relacionadas à competência informacional. Foi realizado um levantamento na base de dados SCOPUS e após refinamento dos resultados obteve-se 71 referências. Como os resultados observou-se que os Estados Unidos e o Reino Unido são os países que mais publicam sobre a temática e que 52.9 % das produções se tratam de artigos científicos. Verificou-se que os documentos analisados ressaltam a importância de que orientações sobre o tema ocorram na escola, com destaque para o papel da biblioteca escolar, visto que na escola pública este é o local em que os estudantes mais acessam a internet. Verificou-se também que o tema uso seguro da informação abrange muitos outros subtemas: exposição de dados, abuso sexual, cyberbullying. Concluiu-se que a pesquisa sobre uso seguro da informação em ambientes online é complexa e que a área carece de pesquisas sobre a temática, em particular no Brasil, visando obter mais subsídios que permitam conhecer melhor esta questão e tendo a escola como espaço de aplicação, discussão e orientação que possibilite o desenvolvimento de habilidades para uma navegação mais segura por adolescentes.Access the information using the internet is gaining more and more space in our society, including among children and adolescents. The study aimed to identify research trends related to the safe use of online information and the development of information literacy. A survey was performed in the SCOPUS database and after refining the results, 71 references were obtained. As a result it is observed that the United States and the United Kingdom are the largest producers of content on the subject, 52.9% of the productions are through scientific articles. It was found that the documents underscore the importance of guidance on the subject in the school, especially the school library in this process, being in public school, the place that students most access the internet. It has also been found that the safe use of information theme covers many other subthemes: data exposure, sexual abuse, cyberbullying. It was concluded that the research on secure use of information in online environments is complex and that the area lacks research on the subject aiming to obtain more subsidies to better understand this issue and having the school as a space for application, discussion and guidance that enable the development of skills for safer browsing by teenagers.Facultad de Humanidades y Ciencias de la Educació

Autosomal Dominant Retinal Dystrophies Caused by a Founder Splice Site Mutation, c.828+3A>T, in PRPH2

PURPOSE: We determined the phenotypic variation, disease progression, and potential modifiers of autosomal dominant retinal dystrophies caused by a splice site founder mutation, c.828+3A>T, in the PRPH2 gene. METHODS: A total of 62 individuals (19 families) harboring the PRPH2 c.828+3A>T mutation, had phenotype analysis by fundus appearance, electrophysiology, and visual fields. The PRPH2 haplotypes in trans were sequenced for potential modifying variants and generalized estimating equations (GEE) used for statistical analysis. RESULTS: Several distinct phenotypes caused by the PRPH2 c.828+3A>T mutation were observed and fell into two clinical categories: Group I (N = 44) with mild pattern dystrophies (PD) and Group II (N = 18) with more severe cone-rod dystrophy (CRD), retinitis pigmentosa (RP), and central areolar chorioretinal dystrophy (CACD). The PRPH2 Gln304-Lys310-Asp338 protein haplotype in trans was found in Group I only (29.6% vs. 0%), whereas the Glu304-Lys310-Gly338 haplotype was predominant in Group II (94.4% vs. 70.4%). Generalized estimating equations analysis for PD versus the CRD/CACD/RP phenotypes in individuals over 43 years alone with the PRPH2 haplotypes in trans and age as predictors, adjusted for correlation within families, confirmed a significant effect of haplotype on severity (P = 0.03) with an estimated odds ratio of 7.16 (95% confidence interval [CI] = [2.8, 18.4]). CONCLUSIONS: The PRPH2 c.828+3A>T mutation results in multiple distinct phenotypes likely modified by protein haplotypes in trans; the odds of having the CACD/RP-like phenotype (versus the PD phenotype) are 7.16 times greater with a Glu304-Lys310-Gly338 haplotype in trans. Further functional studies of the modifying haplotypes in trans and PRPH2 splice variants may offer therapeutic targets

Founder Effect of a c.828+3A>T Splice Site Mutation in Peripherin 2 ( PRPH2

IMPORTANCE: Screening for splice site mutation c.828+3A>T in the peripherin 2 (PRPH2) gene should be a high priority in families with highly variable retinal dystrophies. The correction of missplicing is a potential therapeutic target. OBJECTIVE: To determine the prevalence, genetic origin, and molecular mechanism of a donor c.828+3A>T mutation in the PRPH2 (peripherin 2, retinal degeneration slow) gene in individuals with retinal dystrophies. DESIGN, SETTING, AND PARTICIPANTS: Case-control study that took place at the University of Texas Health Science Center, the University of Iowa, and the Retina Foundation of the Southwest, from January 1, 1987, to August 1, 2014, including affected individuals from 200 families with a diagnosis of autosomal dominant retinitis pigmentosa, 35 families with unspecified macular dystrophies, and 116 families with pattern dystrophy. Participants were screened for the c.828+3A>T mutation by restriction-enzyme digest, single-strand conformational polymorphism screening, or bidirectional sequencing. Haplotypes of polymorphic markers flanking the PRPH2 locus and sequence variants within the gene were determined by denaturing gel electrophoresis or automated capillary-based cycle sequencing. The effect of the splice site mutation on the PRPH2 transcript was analyzed using NetGene2, a splice prediction program and by the reverse transcription polymerase chain reaction of illegitimate transcripts from peripheral white blood cells. MAIN OUTCOMES AND MEASURES: Results of testing for splice site mutation, haplotypes, and alternate transcripts. RESULTS: The PRPH2 mutation was found in 97 individuals of 19 independently ascertained families with a clinical diagnosis of retinitis pigmentosa, macular dystrophy, and/or pattern dystrophy. All affected individuals also shared a rare haplotype of approximately 644 kilobase pairs containing the c.828+3A>T mutation, which extends from the short tandem repeat polymorphism D6S282 to c.1013G>A (rs434102, a single-nucleotide polymorphism) in exon 3 of PRPH2, suggesting this mutation is from a common ancestor and is a founder mutation. It has a prevalence of 2% in families diagnosed as having autosomal dominant retinitis pigmentosa and 10% in families with variable clinical diagnosis of pattern, macular, and retinal dystrophies. Individuals with the c.828+3A>T mutation expressed a PRPH2 transcript not found in control participants and that was consistent with abnormal splicing. CONCLUSIONS AND RELEVANCE: The PRPH2 c.828+3A>T splice site mutation is a frequent cause of inherited retinal dystrophies and is owing to the founder effect. The likely cause of disease is the missplicing of the PRPH2 message that results in a truncated protein product. Identifying the genetic etiology assists in more accurate management and possible future therapeutic options

Autosomal Dominant Retinal Dystrophies Caused by a Founder Splice Site Mutation, c.828+3A>T, in PRPH2 and Protein Haplotypes in trans as Modifiers.

PurposeWe determined the phenotypic variation, disease progression, and potential modifiers of autosomal dominant retinal dystrophies caused by a splice site founder mutation, c.828+3A>T, in the PRPH2 gene.MethodsA total of 62 individuals (19 families) harboring the PRPH2 c.828+3A>T mutation, had phenotype analysis by fundus appearance, electrophysiology, and visual fields. The PRPH2 haplotypes in trans were sequenced for potential modifying variants and generalized estimating equations (GEE) used for statistical analysis.ResultsSeveral distinct phenotypes caused by the PRPH2 c.828+3A>T mutation were observed and fell into two clinical categories: Group I (N = 44) with mild pattern dystrophies (PD) and Group II (N = 18) with more severe cone-rod dystrophy (CRD), retinitis pigmentosa (RP), and central areolar chorioretinal dystrophy (CACD). The PRPH2 Gln304-Lys310-Asp338 protein haplotype in trans was found in Group I only (29.6% vs. 0%), whereas the Glu304-Lys310-Gly338 haplotype was predominant in Group II (94.4% vs. 70.4%). Generalized estimating equations analysis for PD versus the CRD/CACD/RP phenotypes in individuals over 43 years alone with the PRPH2 haplotypes in trans and age as predictors, adjusted for correlation within families, confirmed a significant effect of haplotype on severity (P = 0.03) with an estimated odds ratio of 7.16 (95% confidence interval [CI] = [2.8, 18.4]).ConclusionsThe PRPH2 c.828+3A>T mutation results in multiple distinct phenotypes likely modified by protein haplotypes in trans; the odds of having the CACD/RP-like phenotype (versus the PD phenotype) are 7.16 times greater with a Glu304-Lys310-Gly338 haplotype in trans. Further functional studies of the modifying haplotypes in trans and PRPH2 splice variants may offer therapeutic targets