Frequency, Genotype, and Clinical Spectrum of Best Vitelliform Macular Dystrophy: Data From a National Center in Denmark

PURPOSE: To estimate the prevalence, genotype, and clinical spectrum of Best vitelliform macular dystrophy (Best disease). DESIGN: Retrospective epidemiologic and clinical and molecular genetic observational study. METHODS: SETTING: National referral center. PARTICIPANTS: Forty-five individuals diagnosed with Best disease. OBSERVATION PROCEDURES: Retrospective review of patients diagnosed according to clinical findings and sequencing of BEST1. Patients with recently established molecular genetic diagnosis were followed up including multifocal electroretinography (mfERG), spectral-domain optical coherence tomography (SD-OCT), and fundus autofluorescence (FAF) imaging. MAIN OUTCOME MEASURES: BEST1 mutations, SD-OCT and FAF findings, mfERG amplitudes, prevalence estimate of Best disease. RESULTS: BEST1 mutations described previously in Danish patients with Best disease are reviewed. In addition, we identified a further 8 families and 1 sporadic case, in whom 6 BEST1 missense mutations were found, 4 of which are novel. The mutation c.904G>T (p.Asp302Asn) was identified in members of 4 unrelated families. Structural alterations ranged from precipitate-like alterations at the level of the photoreceptor outer segments (OS) to choroidal neovascularization. The extent of the former correlated with the reduction of retinal function. A prevalence estimate of Best disease in Denmark based on the number of diagnosed cases was 1.5 per 100 000 individuals. CONCLUSIONS: Our data expand the mutation spectrum of BEST1 in patients with Best disease. Alterations of the OS overlying lesions with subretinal fluid are similar to those seen in central serous retinopathy and may indicate impaired turnover of OS. Our frequency estimate confirms that Best disease is one of the most common causes of early macular degeneration. (Am J Ophthalmol 2012;154:403-412. (c) 2012 by Elsevier Inc. All rights reserved.

Clinical evaluation of two consanguineous families with homozygous mutations in BEST1

Purpose: To describe the clinical and genetic findings in two consanguineous families with Best vitelliform macular dystrophy (BVMD) and homozygous mutations in the bestrophin-1 (BEST1) gene. Methods: Ophthalmologic examination was performed in eight members of two families originating from Spain and Denmark. Mutation screening was performed using the Vitelliform Macular Dystrophy mutation array from Asper Biotech, and by the directed genomic sequencing of BEST1. Results: Two homozygous mutations were detected in these families. Mutation c.936C>A (p.Asp312Glu) has been reported previously in a Danish family; here, we describe four additional individuals in this family demonstrating findings compatible with a severe dominant BVMD, albeit with reduced penetrance in heterozygotes. In the Spanish family, a novel homozygous missense mutation in exon 4, c. 388 C>A (p.Arg130Ser), was identified in the siblings. Homozygous siblings demonstrated evidence of multifocal vitelliform retinopathy, whereas heterozygous family members presented findings ranging from isolated reduction of the electrooculogram Arden ratio to normal values on all clinical parameters. Conclusions: As demonstrated in these consanguineous families, a great clinical variability is associated with homozygous mutations in BEST1, ranging from severe dominant BVMD with reduced penetrance in heterozygotes to autosomal recessive bestrophinopathy

Interactome analysis reveals that FAM161A, deficient in recessive retinitis pigmentosa, is a component of the Golgi-centrosomal network

Defects in FAM161A, a protein of unknown function localized at the cilium of retinal photoreceptor cells, cause retinitis pigmentosa, a form of hereditary blindness. By using different fragments of this protein as baits to screen cDNA libraries of human and bovine retinas, we defined a yeast two-hybrid-based FAM161A interactome, identifying 53 bona fide partners. In addition to statistically significant enrichment in ciliary proteins, as expected, this interactome revealed a substantial bias towards proteins from the Golgi apparatus, the centrosome and the microtubule network. Validation of interaction with key partners by co-immunoprecipitation and proximity ligation assay confirmed that FAM161A is a member of the recently recognized Golgi-centrosomal interactome, a network of proteins interconnecting Golgi maintenance, intracellular transport and centrosome organization. Notable FAM161A interactors included AKAP9, FIP3, GOLGA3, KIFC3, KLC2, PDE4DIP, NIN and TRIP11. Furthermore, analysis of FAM161A localization during the cell cycle revealed that this protein followed the centrosome during all stages of mitosis, likely reflecting a specific compartmentalization related to its role at the ciliary basal body during the G0 phase. Altogether, these findings suggest that FAM161A's activities are probably not limited to ciliary tasks but also extend to more general cellular functions, highlighting possible novel mechanisms for the molecular pathology of retinal diseas

FAM161A, associated with retinitis pigmentosa, is a component of the cilia-basal body complex and interacts with proteins involved in ciliopathies

Retinitis pigmentosa (RP) is a retinal degenerative disease characterized by the progressive loss of photoreceptors. We have previously demonstrated that RP can be caused by recessive mutations in the human FAM161A gene, encoding a protein with unknown function that contains a conserved region shared only with a distant paralog, FAM161B. In this study, we show that FAM161A localizes at the base of the photoreceptor connecting cilium in human, mouse and rat. Furthermore, it is also present at the ciliary basal body in ciliated mammalian cells, both in native conditions and upon the expression of recombinant tagged proteins. Yeast two-hybrid analysis of binary interactions between FAM161A and an array of ciliary and ciliopathy-associated proteins reveals direct interaction with lebercilin, CEP290, OFD1 and SDCCAG8, all involved in hereditary retinal degeneration. These interactions are mediated by the C-terminal moiety of FAM161A, as demonstrated by pull-down experiments in cultured cell lines and in bovine retinal extracts. As other ciliary proteins, FAM161A can also interact with the microtubules and organize itself into microtubule-dependent intracellular networks. Moreover, small interfering RNA-mediated depletion of FAM161A transcripts in cultured cells causes the reduction in assembled primary cilia. Taken together, these data indicate that FAM161A-associated RP can be considered as a novel retinal ciliopathy and that its molecular pathogenesis may be related to other ciliopathie

The Effect of Cone Opsin Mutations on Retinal Structure and the Integrity of the Photoreceptor Mosaic

Purpose. To evaluate retinal structure and photoreceptor mosaic integrity in subjects with OPN1LW and OPN1MW mutations. Methods. Eleven subjects were recruited, eight of whom have been previously described. Cone and rod density was measured using images of the photoreceptor mosaic obtained from an adaptive optics scanning light ophthalmoscope (AOSLO). Total retinal thickness, inner retinal thickness, and outer nuclear layer plus Henle fiber layer (ONL+HFL) thickness were measured using cross-sectional spectral-domain optical coherence tomography (SD-OCT) images. Molecular genetic analyses were performed to characterize the OPN1LW/OPN1MW gene array. Results. While disruptions in retinal lamination and cone mosaic structure were observed in all subjects, genotype-specific differences were also observed. For example, subjects with “L/M interchange” mutations resulting from intermixing of ancestral OPN1LW and OPN1MW genes had significant residual cone structure in the parafovea (∼25% of normal), despite widespread retinal disruption that included a large foveal lesion and thinning of the parafoveal inner retina. These subjects also reported a later-onset, progressive loss of visual function. In contrast, subjects with the C203R missense mutation presented with congenital blue cone monochromacy, with retinal lamination defects being restricted to the ONL+HFL and the degree of residual cone structure (8% of normal) being consistent with that expected for the S-cone submosaic. Conclusions. The photoreceptor phenotype associated with OPN1LW and OPN1MW mutations is highly variable. These findings have implications for the potential restoration of visual function in subjects with opsin mutations. Our study highlights the importance of high-resolution phenotyping to characterize cellular structure in inherited retinal disease; such information will be critical for selecting patients most likely to respond to therapeutic intervention and for establishing a baseline for evaluating treatment efficacy

Homozygosity for a novel ABCA4 founder splicing mutation is associated with progressive and severe Stargardt-like disease

PURPOSE. To clinically characterize and genetically analyze members of six families who reside in the same village and manifest a rare form of retinal degeneration. METHODS. Ophthalmic evaluation included a full clinical examination, perimetry, color vision testing, and electroretinography. Genomic DNA was screened for ABCA4 mutations with the use of microarray analysis and direct sequencing. RNA analysis was performed with RT-PCR and sequencing. RESULTS. The authors recruited 15 patients with a unique retinal disease who are members of six highly consanguineous ArabMuslim families from a single village. During early stages of disease, funduscopic and angiographic findings as well as retinal function resemble those of Stargardt disease. However, later in life, severe, widespread cone-rod degeneration ensues. Marked progressive involvement of the retinal periphery distinguishes this phenotype from classic Stargardt disease. Genetic analysis of ABCA4 revealed two novel deletions, p.Cys1150del and c.4254-15del23. One patient, who was a compound heterozygote, manifested typical Stargardt disease. The remaining 14 patients were homozygote for the c.4254-15del23 intronic deletion and had the progressive form of disease. We identified an identical ABCA4 haplotype in all alleles carrying this mutation, indicating a founder mutation. Detailed RT-PCR analysis in normal retina and lymphoblastoid cells revealed expression of the full-length ABCA4 transcript and three novel transcripts produced by alternative splicing. The full-length ABCA4 transcript, however, could not be detected in lymphoblastoid cells of affected homozygote patients. CONCLUSIONS. These results expand the genotype-phenotype correlation of ABCA4, showing that homozygosity for the novel c.4254-15del23 splicing mutation is associated with a severe progressive form of disease. (Invest Ophthalmol Vis Sci

Molecular Anthropology Meets Genetic Medicine to Treat Blindness in the North African Jewish Population: Human Gene Therapy Initiated in Israel

Abstract The history of the North African Jewish community is ancient and complicated with a number of immigration waves and persecutions dramatically affecting its population size. A decade-long process in Israel of clinicalmolecular screening of North African Jews with incurable autosomal recessive blindness led to the identification of a homozygous splicing mutation (c.95-2A > T; IVS2-2A > T) in RPE65, the gene encoding the isomerase that catalyzes a key step in the retinoid-visual cycle, in patients from 10 unrelated families. A total of 33 patients (four now deceased) had the severe childhood blindness known as Leber congenital amaurosis (LCA), making it the most common cause of retinal degeneration in this population. Haplotype analysis in seven of the patients revealed a shared homozygous region, indicating a population-specific founder mutation. The age of the RPE65 founder mutation was estimated to have emerged 100-230 (mean, 153) generations ago, suggesting it originated before the establishment of the Jewish community in North Africa. Individuals with this RPE65 mutation were characterized with retinal studies to determine if they were candidates for gene replacement, the recent and only therapy to date for this otherwise incurable blindness. The step from molecular anthropological studies to application of genetic medicine was then taken, and a representative of this patient subgroup was treated with subretinal rAAV2-RPE65 gene therapy. An increase in vision was present in the treated area as early as 15 days after the intervention. This process of genetically analyzing affected isolated populations as a screen for genebased therapy suggests a new paradigm for disease diagnosis and treatment