Achromatopsia Research Articles

Retinal Development in Infants and Young Children with Achromatopsia

Retinal Development in Infants and Young Children with Achromatopsia

Computer Assisted Quantitative Electroretinography II: Separation of Rod and Cone Components of the Electroretinogram in Congenital Achromatopsia and Congenital Nyctalopia

Computer Assisted Quantitative Electroretinography II: Separation of Rod and Cone Components of the Electroretinogram in Congenital Achromatopsia and Congenital Nyctalopia

Novel CNGA3 mutations in Chinese patients with achromatopsia

ObjectiveTo study the clinical features and to identify the pathogenic mutations in Chinese patients with achromatopsia (ACHM).DesignFifteen patients from 10 unrelated families were included in this study. Detailed ocular examinations...

Genetics and Disease Expression in the CNGA3 Form of Achromatopsia: Steps on the Path to Gene Therapy

Achromatopsia (ACHM) is a congenital, autosomal recessive retinal disease that manifests cone dysfunction, reduced visual acuity and color vision, nystagmus, and photoaversion. Five genes are known causes of ACHM. The present study took steps toward performing a trial of gene therapy in ACHM by characterizing the genetics of ACHM in Israel and the Palestinian Territories and analyzing retinal function and structure in CNGA3 ACHM patients from the Israeli-Palestinian population and US patients with other origins. Case series study. Patients with clinically suspected ACHM, cone dysfunction phenotypes, and unaffected family members were included. The protocol was approved by the local institutional review board and informed consent was obtained from all participants. Genetic analyses included homozygosity mapping and exome sequencing. Phenotype was assessed with electroretinography (ERG), optical coherence tomography, psychophysics, and photoaversion testing. Single nucleotide polymorphism microarray, exome analysis, DNA sequence analysis, visual function testing including ERG, and photoaversion. We identified 148 ACHM patients from 57 Israeli and Palestinian families; there were 16 CNGA3 mutations (5 novel) in 41 families and 5 CNGB3 mutations (1 novel) in 8 families. Two CNGA3 founder mutations underlie >50% of cases. These mutations lead to a high ACHM prevalence of ∼1:5000 among Arab-Muslims residing in Jerusalem. Rod ERG abnormalities (in addition to cone dysfunction) were detected in 59% of patients. Retinal structure in CNGA3 ACHM patients revealed persistent but abnormal foveal cones. Under dark- and light-adapted conditions, patients use rod-mediated pathways. Photoaversion was readily demonstrated with transition from the dark to a dim light background. Among Israeli and Palestinian patients, CNGA3 mutations are the leading cause of ACHM. Retinal structural results support the candidacy of CNGA3 ACHM for clinical trials for therapy of cone photoreceptors. Efficacy outcome measures would include chromatic light-adapted psychophysics, with attention to the photoreceptor basis of the response, and quantitation of photoaversion.

A Case of Joubert Syndrome Accompanied with Retinal Abnormality

Purpose: Joubert syndrome is a rare disorder which affects the cerebellum and the brain stem. Herein, we report a case of Joubert syndrome accompanied with retinal abnormality. Case summary: A 9-year-old female visited our hospital with chief complaints of low vision in both eyes, nystagmus, and lack of gaze movement. The best-corrected visual acuity in her right eye was 20/80 and in the left 20/80 and heterotropia was not observed. She appeared to have incomplete total color blindness on the color vision test. The anterior segment test showed no abnormal findings other than diffuse pigmentation and degeneration of the peripheral retina, vascular attenuation, and pale optic disc in both eyes on fundus examination. The patient showed overall developmental delay and decreased muscle tension, but genetic and congenital metabolic disease tests were normal. The molar tooth sign of the midbrain, defect in the lower part of the cerebellum and dilatation of the fourth ventricle were observed on magnetic resonance imaging. Conclusions: Appropriate evaluation of retinitis pigmentosa and visual function should be performed in Joubert syndrome patients. J Korean Ophthalmol Soc 2015;56(8):1304-1309

Genotype-dependent variability in residual cone structure in achromatopsia: toward developing metrics for assessing cone health.

Gene therapy trials for inherited photoreceptor disorders are planned. Anatomical metrics to select the best candidates and outcomes are needed. Adaptive optics (AO) imaging enables visualization of photoreceptor structure, although analytical tools are lacking. Here we present criteria to assess residual photoreceptor integrity in achromatopsia (ACHM). Two AOSLOs, at the Medical College of Wisconsin and Moorfields Eye Hospital, were used to image the photoreceptor mosaic of 11 subjects with ACHM and 7 age-matched controls. Images were obtained, processed, and montaged using previously described methods. Cone density and reflectivity were quantified to assess residual cone photoreceptor structure. All subjects with ACHM had reduced numbers of cone photoreceptors, albeit to a variable degree. In addition, the relative cone reflectivity varied greatly. Interestingly, subjects with GNAT2-associated ACHM had the greatest number of residual cones and the reflectivity of those cones was significantly greater than that of the cones in the subjects with CNGA3/CNGB3-associated ACHM. We present cone reflectivity as a metric that can be used to characterize cone structure in ACHM. This method may be applicable to subjects with other cone disorders. In ACHM, we hypothesize that cone numerosity (and/or density) combined with cone reflectivity could be used to gauge the therapeutic potential. As gene replacement would not be expected to add cones, reflectivity could be a more powerful AO-metric for monitoring the cellular response to treatment and could provide a more immediate indicator of efficacy than behavioral measures, which may take longer to change.

A prospective longitudinal study of retinal structure and function in achromatopsia.

To longitudinally characterize retinal structure and function in achromatopsia (ACHM) in preparation for clinical gene therapy trials. Thirty-eight molecularly confirmed ACHM subjects underwent serial assessments, including spectral domain optical coherence tomography (SD-OCT), microperimetry, and fundus autofluorescence (FAF). Foveal structure on SD-OCT was graded and compared for evidence of progression, along with serial measurements of foveal total retinal thickness (FTRT) and outer nuclear layer (ONL) thickness. Fundus autofluorescence patterns were characterized and compared over time. Mean follow-up was 19.5 months (age range at baseline, 6-52 years). Only 2 (5%) of 37 subjects demonstrated change in serial foveal SD-OCT scans. There was no statistically significant change over time in FTRT (P = 0.83), ONL thickness (P = 0.27), hyporeflective zone diameter (P = 0.42), visual acuity (P = 0.89), contrast sensitivity (P = 0.22), mean retinal sensitivity (P = 0.84), and fixation stability (P = 0.58). Three distinct FAF patterns were observed (n = 30): central increased FAF (n = 4), normal FAF (n = 11), and well-demarcated reduced FAF (n = 15); with the latter group displaying a slow increase in the area of reduced FAF of 0.03 mm(2) over 19.3 months (P = 0.002). Previously published cross-sectional studies have described conflicting findings with respect to the age-dependency of progression. This study, which constitutes the largest and longest prospective longitudinal study of ACHM to date, suggests that although ACHM may be progressive, any such progression is slow and subtle in most patients, and does not correlate with age or genotype. We also describe the first serial assessment of FAF, which is highly variable between individuals, even of similar age and genotype.

Causes and consequences of inherited cone disorders.

Hereditary cone disorders (CDs) are characterized by defects of the cone photoreceptors or retinal pigment epithelium underlying the macula, and include achromatopsia (ACHM), cone dystrophy (COD), cone-rod dystrophy (CRD), color vision impairment, Stargardt disease (STGD) and other maculopathies. Forty-two genes have been implicated in non-syndromic inherited CDs. Mutations in the 5 genes implicated in ACHM explain ∼93% of the cases. On the contrary, only 21% of CRDs (17 genes) and 25% of CODs (8 genes) have been elucidated. The fact that the large majority of COD and CRD-associated genes are yet to be discovered hints towards the existence of unknown cone-specific or cone-sensitive processes. The ACHM-associated genes encode proteins that fulfill crucial roles in the cone phototransduction cascade, which is the most frequently compromised (10 genes) process in CDs. Another 7 CD-associated proteins are required for transport processes towards or through the connecting cilium. The remaining CD-associated proteins are involved in cell membrane morphogenesis and maintenance, synaptic transduction, and the retinoid cycle. Further novel genes are likely to be identified in the near future by combining large-scale DNA sequencing and transcriptomics technologies. For 31 of 42 CD-associated genes, mammalian models are available, 14 of which have successfully been used for gene augmentation studies. However, gene augmentation for CDs should ideally be developed in large mammalian models with cone-rich areas, which are currently available for only 11 CD genes. Future research will aim to elucidate the remaining causative genes, identify the molecular mechanisms of CD, and develop novel therapies aimed at preventing vision loss in individuals with CD in the future.

Contribution du SD-OCT de la macula dans le diagnostic de l’achromatopsie congénitale : à propos de 6 patients

Achromatopsia (ACH) is a congenital autosomal recessive cone disorder. The puspose is to describe particular SD-OCT macular images in ACH. The study included 6 patients from 3 consanguineous Tunisian families with congenital nystagmus and amblyopia with ACH. All patients had clinical examination with fundus photography, autofluorescence, 100-Hue Color vision and the appearance and thickness of all retinal layers were evaluated by spectral-domain optical coherence tomography (SD-OCT). All patients had ACH. The feature was loss of inner- and outer-segments (IS/OS) with disruption of the ciliary layer on OCT and an appearance of partial-thickness hole in the outer macular retina. This feature seems to be characteristic of ACH. SD-OCT correlated to clinic signs help the diagnosis.

Retinal Structure and Function in Achromatopsia: Implications for Gene Therapy

To characterize retinal structure and function in achromatopsia (ACHM) in preparation for clinical trials of gene therapy. Cross-sectional study. Forty subjects with ACHM. All subjects underwent spectral domain optical coherence tomography (SD-OCT), microperimetry, and molecular genetic testing. Foveal structure on SD-OCT was graded into 5 distinct categories: (1) continuous inner segment ellipsoid (ISe), (2) ISe disruption, (3) ISe absence, (4) presence of a hyporeflective zone (HRZ), and (5) outer retinal atrophy including retinal pigment epithelial loss. Foveal and outer nuclear layer (ONL) thickness was measured and presence of hypoplasia determined. Photoreceptor appearance on SD-OCT imaging, foveal and ONL thickness, presence of foveal hypoplasia, retinal sensitivity and fixation stability, and association of these parameters with age and genotype. Forty subjects with a mean age of 24.9 years (range, 6-52 years) were included. Disease-causing variants were found in CNGA3 (n = 18), CNGB3 (n= 15), GNAT2 (n= 4), and PDE6C (n= 1). No variants were found in 2 individuals. In all, 22.5% of subjects had a continuous ISe layer at the fovea, 27.5% had ISe disruption, 20% hadan absent ISe layer, 22.5% had an HRZ, and 7.5% had outer retinal atrophy. No significant differences in age (P= 0.77), mean retinal sensitivity (P= 0.21), or fixation stability (P= 0.34) across the 5 SD-OCT categories were evident. No correlation was found between age and foveal thickness (P= 0.84) or between age and foveal ONL thickness (P= 0.12). The lack of a clear association of disruption of retinal structure or function in ACHM with age suggests that the window of opportunity for intervention by gene therapy is wider in some individuals than previously indicated. Therefore, the potential benefit for a given subject is likely to be better predicted by specific measurement of photoreceptor structure rather than simply by age. The ability to directly assess cone photoreceptor preservation with SD-OCT and/or adaptive optics imaging is likely to prove invaluable in selecting subjects for future trials and measuring the trials' impact.

Genomic deletion of CNGB3 is identical by descent in multiple canine breeds and causes achromatopsia

BackgroundAchromatopsia is an autosomal recessive disease characterized by the loss of cone photoreceptor function that results in day-blindness, total colorblindness, and decreased central visual acuity. The most common causes for the disease are mutations in the CNGB3 gene, coding for the beta subunit of the cyclic nucleotide-gated channels in cones. CNGB3-achromatopsia, or cone degeneration (cd), is also known to occur in two canine breeds, the Alaskan malamute (AM) and the German shorthaired pointer.ResultsHere we report an in-depth characterization of the achromatopsia phenotype in a new canine breed, the miniature Australian shepherd (MAS). Genotyping revealed that the dog was homozygous for a complete genomic deletion of the CNGB3 gene, as has been previously observed in the AM. Identical breakpoints on chromosome 29 were identified in both the affected AM and MAS with a resulting deletion of 404,820 bp. Pooled DNA samples of unrelated purebred Australian shepherd, MAS, Siberian husky, Samoyed and Alaskan sled dogs were screened for the presence of the affected allele; one Siberian husky and three Alaskan sled dogs were identified as carriers. The affected chromosomes from the AM, MAS, and Siberian husky were genotyped for 147 SNPs in a 3.93 Mb interval within the cd locus. An identical shared affected haplotype, 0.5 Mb long, was observed in all three breeds and defined the minimal linkage disequilibrium (LD) across breeds. This supports the idea that the mutated allele was identical by descent (IBD).ConclusionWe report the occurrence of CNGB3-achromatopsia in a new canine breed, the MAS. The CNGB3-deletion allele previously described in the AM was also observed in a homozygous state in the affected MAS, as well as in a heterozygous carrier state in a Siberian husky and Alaskan sled dogs. All affected alleles were shown to be IBD, strongly suggesting an affected founder effect. Since the MAS is not known to be genetically related to the AM, other breeds may potentially carry the same cd-allele and be affected by achromatopsia.

Development of a pharmacorphore model for pharmacological chaperones targeting mutant trafficking-deficient CNG channels

Complete colorblindness (achromatopsia) is caused by autosomal recessively inherited mutations in the retinal phototransduction pathway, predominantly in the CNGA3- and CNGB3-subunit of the cyclic nucleotide-gated (CNG) channels in cone photoreceptors. CNGA3, which is mutated in about 25% of the achromatopsia patients, mainly harbors missense mutations which frequently impair the folding and/or trafficking of the mutant CNGA3-channels [1]. Pharmacological chaperones stabilizing the folding of the mutant protein may be used to overcome this folding-/trafficking-deficiency. More than 50 compounds were evaluated in their ability to restore signal transduction using a calcium imaging-based bioassay utilizing the CNGA3-mutant E228K [2]. With this data we created several pharmacophore models using Schrodinger Phase [3], which describe the chemical features of potential pharmacological chaperones targeting achromatopsia. We used several approaches leading to different pharmacophore hypotheses: a) Training with the complete set of experimental data (see Figure ​Figure11) Figure 1 Pharmacophore depicting potential features of CNG channel-chaperones. b) Training with only dihydropyridines since this group showed the highest experimental activity, and c) Training with a data set excluding dihydropyridines. Our in-house database TueScreen, which includes ZINC12 [4], was screened to identify potentially active compounds. As a result, several potential molecule classes could be found that may be useful as pharmacological chaperones to improve folding/trafficking of mutant CNG-channels. We will experimentally validate these predictions in a calcium imaging-based bioassay.

RhodopsinF45L Allele Does Not Cause Autosomal Dominant Retinitis Pigmentosa in a Large Caucasian Family

To ascertain the potential pathogenicity of a retinitis pigmentosa (RP)-causing RHO F45L allele in a family affected by congenital achromatopsia (ACHM). Case series/observational study that included two patients with ACHM and 24 extended family members. Molecular genetic analysis was performed to identify RHO F45L carrier status in the family and a control population. An adaptive optics scanning light ophthalmoscope (AOSLO) was used to image the photoreceptor mosaic and assess rod and cone structure. Spectral domain optical coherence tomography (SD-OCT) was used to examine retinal lamination. Comprehensive clinical testing included acuity, color vision, and dilated fundus examination. Electroretinography was used to assess rod and cone function. Five carriers of the RHO F45L allele alone (24-80 years) and three carriers in combination with a heterozygous CNGA3 mutant allele (10-64 years) were all free of the classic symptoms and signs of RP. In heterozygous carriers of both mutations, SD-OCT showed normal retinal thickness and intact outer retinal layers; rod and cone densities were within normal limits on AOSLO. The phenotype in two individuals affected with ACHM and harboring the RHO F45L allele was indistinguishable from that previously reported for ACHM. The RHO F45L allele is not pathogenic in this large family; hence, the two ACHM patients would unlikely develop RP in the future. The combined approach of comprehensive molecular analysis of individual genomes and noninvasive cellular resolution retinal imaging enhances the current repertoire of clinical diagnostic tools, giving a substantial impetus to personalized medicine.

THE COLOUR VISION DEFICIENCIES AMONG STUDENTS OF HUE SECONDARY SCHOOL OF TRANSPORTATION

Background: Colorblind disability causes difficulties for people in daily life. Derived from the critical requirement to detect the patients in order to help prevent inappropriate careers, especially careers related to transportation, this research aim to determine the situations and the distributions of different types of visual disabilities. Materials: 1174 students (787 boys and 387 girls) including 2 groups: occupation group and transportation group were tested with ISHIHARA chromatic plates for colour vision deficiencies (CVD) (CVD iclude the total colour blindness, red- green blindness, red-blindness, green- blindness). The results are showed as follow: (i) Frequency of CVD boys among boy group is 4.70%; (ii) Frequency of CVD girls among girls group is 2.58%; (iii) In boy group, among 3 types of red- green blindness, the green-blindness has the higher frequency (3.18%) comparision with these ones of the red- green blindness and red-blindness. The total colour defiency was hardly; (iv) Frequency of CVD students among occupation group is 4.15%; (v) Frequency of CVD students among transportation group is 3.83%.

A Nonsense Mutation in PDE6H Causes Autosomal-Recessive Incomplete Achromatopsia

Achromatopsia (ACHM) is an autosomal-recessive retinal dystrophy characterized by color blindness, photophobia, nystagmus, and severely reduced visual acuity. Its prevalence has been estimated to about 1 in 30,000 individuals. Four genes, GNAT2, PDE6C, CNGA3, and CNGB3, have been implicated in ACHM, and all encode functional components of the phototransduction cascade in cone photoreceptors. Applying a functional-candidate-gene approach that focused on screening additional genes involved in this process in a cohort of 611 index cases with ACHM or other cone photoreceptor disorders, we detected a homozygous single base change (c.35C>G) resulting in a nonsense mutation (p.Ser12(∗)) in PDE6H, encoding the inhibitory γ subunit of the cone photoreceptor cyclic guanosine monophosphate phosphodiesterase. The c.35C>G mutation was present in three individuals from two independent families with a clinical diagnosis of incomplete ACHM and preserved short-wavelength-sensitive cone function. Moreover, we show through immunohistochemical colocalization studies in mouse retina that Pde6h is evenly present in all retinal cone photoreceptors, a fact that had been under debate in the past. These findings add PDE6H to the set of genes involved in autosomal-recessive cone disorders and demonstrate the importance of the inhibitory γ subunit in cone phototransduction.

Progress on study of achromatopsia and targeted gene therapy

Achromatopsia is an early onset retinal dystrophy that causes severe visual impairment. To date, four genes have been found to be implicated in achromatopsia-associated mutations: guanine nucleotide-binding protein (GNAT2), cyclic nucleotide-gated channel alpha-3 (CNGA3), cyclic nucleotide-gated channel beta-3 (CNGB3) and phosphodiesterase 6C (PDE6C). Even with early onset, the slow progress and the good responses to gene therapy in animal models render achromatopsia a very attractive candidate for human gene therapy after the successful of the Phase I clinical trials of Leber's congenital amaurosis. With the development of molecular genetics and the therapeutic gene replacement technology, the adeno-associated viral (AAV) vector-mediated gene therapy for achromatopsia in the preclinical animal experiments achieved encouraging progress in the past years. This article briefly reviews the recent research achievements of achromatopsia with gene therapy.

Coherent form, motion and biological motion perception in congenital achromatopsia

Coherent form, motion and biological motion perception in congenital achromatopsia

Genetic analysis of four Pakistani families with achromatopsia and a novel S4 motif mutation of CNGA3

To identify the causative variants of achromatopsia (ACHM) in four Pakistani families presenting autosomal recessive ACHM. Four families (50, 55, 70 and 74) exhibiting features of achromatopsia were subjected to homozygosity mapping with STS markers flanking known ACHM loci. Mutation screening was done for two of the families linked to CNGA3 and CNGB3 by direct sequencing of the coding regions and exon-intron boundaries of genes to find the pathogenic variant. Homozygosity mapping showed co-segregation of CNGA3 in family 50 and CNGB3 in family 74. Sequencing of coding regions of CNGA3 in family 50 revealed a novel missense mutation, c.827A>G, in exon 7, which results in p.N276S substitution. N276S is located in the S4 motif of the CNGA3 protein and is conserved in all channel proteins. Bioinformatics analysis showed that the N276S substitution altered the channel conformation by shifting the helix. No pathogenic variation was identified in any affected members of family 74 in the coding sequence of CNGB3. The other two families, 55 and 70, were not linked to any known ACHM loci, indicating further heterogeneity of the ACHM phenotype. We describe a novel S4 motif mutation of CNGA3 in a Pakistani family.

A D262n Mutation in Canine CNGB3 in a Highly Conserved Acidic S2 Region Causes Complete Colorblindness in a Canine Model

Cyclic nucleotide-gated (CNG) channels in cone photoreceptors are composed of CNGA3 (A3) and CNGB3 (B3) subunits. In a canine achromatopsia model, loss of cone ERG function and color vision are associated with the mutation of Asp (D) 262 to Asn (N) in the B3 subunit. This Asp residue is conserved in all CNG channels. We examined the D/N mutation in the human (h) B3 channel co-expressed with a wild-type canine (c) A3 and in a mutant cA3 homomeric channel in which the conserved Asp residue was substituted with Asn. In patch clamp experiments, both heteromeric wild type cA3 co-expressed with D/N hB3 and mutant homomeric D/N cA3 CNG channels had no cGMP-activated currents providing molecular insights into the cause of canine achromatopsia. D262 of the cB3 is located in the S2 transmembrane domain and, along with D252 and D256, these residues comprise an acidic motif conserved in the CNG channel family and extending to some members of the Shaker K superfamily. Hints about the role of this Asp motif have been gained from studies of related channels reporting interactions between the negatively-charged residues in S2 and the conserved positively-charged residues in other transmembrane domains. We generated mutant A3 subunits by substituting each of the three conserved S2 Asp residues with Asn, Cys, or Glu. Patch clamp analysis showed all of the mutant homomeric A3 channels lost function. These YFP-tagged A3 mutant channels exhibited changes in cellular localization compared to wild type channels suggesting that any change in the Asp motif alters subunit trafficking. Future studies will focus on understanding the role of this Asp motif in intra-subunit helical contacts and proper trafficking and assembly of cone CNG channels.

Decreased catalytic activity and altered activation properties of PDE6C mutants associated with autosomal recessive achromatopsia

Mutations in the gene encoding the catalytic subunit of the cone photoreceptor phosphodiesterase (PDE6C) have been recently reported in patients with autosomal recessive inherited achromatopsia (ACHM) and early-onset cone photoreceptor dysfunction. Here we present the results of a comprehensive study on PDE6C mutations including the mutation spectrum, its prevalence in a large cohort of ACHM/cone dysfunction patients, the clinical phenotype and the functional characterization of mutant PDE6C proteins. Twelve affected patients from seven independent families segregating PDE6C mutations were identified in our total patient cohort of 492 independent families. Eleven different PDE6C mutations were found including two nonsense mutations, three mutations affecting transcript splicing as shown by minigene assays, one 1 bp-insertion and five missense mutations. We also performed a detailed functional characterization of six missense mutations applying the baculovirus system to express recombinant mutant and wildtype chimeric PDE6C/PDE5 proteins in Sf9 insect cells. Purified proteins were analyzed using Western blotting, phosphodiesterase (PDE) activity measurements as well as inhibition assays by zaprinast and Pγ. Four of the six PDE6C missense mutations led to baseline PDE activities and most likely represent functional null alleles. For two mutations, p.E790K and p.Y323N, we observed reduction in PDE activity of approximately 60% and 80%, respectively. We also observed differences for Pγ inhibition. The p.E790K mutant, with an IC₅₀ value of 2.7 nm is 20.7-fold more sensitive for Pγ inhibition, whereas the p.Y323N mutant with an IC₅₀ of 158 nm is 3-fold less sensitive when compared with the wildtype control.