Maternal uniparental isodisomy of chromosome 6 reveals a TULP1 mutation as a novel cause of cone dysfunction.

PURPOSE: The majority of the genetic causes of autosomal recessive (ar) cone dystrophy (CD) and cone-rod dystrophy (CRD) are currently unknown. We used a high-resolution homozygosity mapping approach in a cohort of patients with CD or CRD to identify new genes for ar cone disorders. DESIGN: Case series. PARTICIPANTS: A cohort of 159 patients with ar CD and 91 patients with CRD. METHODS: The genomes of 83 patients with ar CD and 73 patients with CRD were analyzed for homozygous regions using single nucleotide polymorphism (SNP) microarrays. One patient showed homozygosity of SNPs across chromosome 6, and segregation analysis was performed using microsatellite markers. Direct sequencing of all retinal disease genes on chromosome 6 revealed a novel pathogenic TULP1 mutation in this patient. A cohort of 159 individuals with CD and 91 individuals with CRD was screened for this particular mutation using the restriction enzyme HhaI. The medical history of patients carrying the TULP1 mutation was reviewed and additional ophthalmic examinations were performed, including electroretinography (ERG), perimetry, optical coherence tomography (OCT), fundus autofluorescence (FAF), and fundus photography. MAIN OUTCOME MEASURES: TULP1 mutations, age at diagnosis, visual acuity, fundus appearance, color vision defects, visual field, ERG, FAF, and OCT findings. RESULTS: In 1 patient, homozygosity mapping and subsequent segregation analysis revealed maternal uniparental disomy (UPD) of chromosome 6. A novel homozygous missense mutation (p.Arg420Ser) was identified in TULP1, whereas no mutations were detected in other retinal disease genes on chromosome 6. The mutation affects a highly conserved amino acid residue in the Tubby domain and is predicted to be pathogenic. The same homozygous mutation was also identified in an additional, unrelated patient with CRD. Both patients carrying the p.Arg420Ser mutation presented with a bull's eye maculopathy. The first patient had progressive loss of visual acuity with a relatively preserved ERG, whereas the second patient developed loss of visual acuity, peripheral degeneration, and severely reduced ERG responses in a cone-rod pattern. CONCLUSIONS: Maternal UPD of chromosome 6 unmasked a mutation in the TULP1 gene as a novel cause of cone dysfunction. This expands the disease spectrum of TULP1 mutations from Leber congenital amaurosis and early-onset retinitis pigmentosa to cone-dominated disease. FINANCIAL DISCLOSURE(S): The author(s) have no proprietary or commercial interest in any materials discussed in this article.

Clinical course, genetic etiology, and visual outcome in cone and cone-rod dystrophy.

OBJECTIVE: To evaluate the clinical course, genetic etiology, and visual prognosis in patients with cone dystrophy (CD) and cone-rod dystrophy (CRD). DESIGN: Clinic-based, longitudinal, multicenter study. PARTICIPANTS: Consecutive probands with CD (N = 98), CRD (N = 83), and affected relatives (N = 41 and N = 17, respectively) from various ophthalmogenetic clinics in The Netherlands, Belgium, and the United Kingdom. METHODS: Data on best-corrected Snellen visual acuity, color vision, ophthalmoscopy, fundus photography, Goldmann perimetry, and full-field standard electroretinogram (ERG) from all patients were registered from medical charts over a mean follow-up of 19 years. The ABCA4, CNGB3, KCNV2, PDE6C, and RPGR genes were analyzed by direct sequencing in autosomal recessive (AR) and X-linked (XL), respectively. Genotyping was not undertaken for autosomal-dominant cases. MAIN OUTCOME MEASURES: The 10-year progression of all clinical parameters and cumulative lifetime risk of low vision and legal blindness were assessed. RESULTS: The mean age onset for CD was 16 years (standard deviation, 11), and of CRD 12 years (standard deviation, 11; P = 0.02). The pattern of inheritance was AR in 92% of CD and 90% of CRD. Ten years after diagnosis, 35% of CD and 51% of CRD had a bull's eye maculopathy; 70% of CRD showed absolute peripheral visual field defects and 37% of CD developed rod involvement on ERG. The mean age of legal blindness was 48 (standard error [SE], 3.1) years in CD, and 35 (SE, 1.1; P<0.001) years in CRD. ABCA4 mutations were found in 8 of 90 (9%) of AR-CD, and in 17 of 65 (26%) of AR-CRD. Other mutations were detected in CNGB3 (3/90; 3%), KCNV2 (4/90; 4%), and in PDE6C (1/90; 1%). The RPGR gene was mutated in the 2 XL-CD and in 4 of 5 (80%) of XL-CRD. ABCA4 mutations as well as age of onset <20 years were significantly associated with a faster progression to legal blindness (P<0.001). CONCLUSIONS: Although CD had a slightly more favorable clinical course than CRD, both disorders progressed to legal blindness in the majority of patients. Mutations in the ABCA4 gene and early onset of disease were independent prognostic parameters for visual loss. Our data may serve as an aid in counseling patients with progressive cone disorders.

Progressive loss of cones in achromatopsia: an imaging study using spectral-domain optical coherence tomography.

PURPOSE: Achromatopsia (ACHM) is a congenital autosomal recessive cone disorder with a presumed stationary nature and only a few causative genes. Animal studies suggest that ACHM may be a good candidate for corrective gene therapy. Future implementation of this therapy in humans requires the presence of viable cone cells in the retina. In this study the presence of cone cells in ACHM was determined, as a function of age. METHODS: The appearance and thickness of all retinal layers were evaluated by spectral-domain optical coherence tomography (SD-OCT) in 40 ACHM patients (age range, 4-70 years) with known mutations in the CNGB3, CNGA3, and PDE6C genes. A comparison was made with 55 healthy age-matched control subjects. RESULTS: The initial feature of cone cell decay was loss of inner and outer segments with disruption of the ciliary layer on OCT, which was observed as early as 8 years of age. Cone cell loss further progressed with age and occurred in 8 (42%) of 19 patients below 30 years and in 20 (95%) of 21 of those aged 30+ years. Retinal thickness was significantly thinner in the fovea of all patients (126 µm in ACHM vs. 225 µm in the control; P < 0.001) and correlated with age (ß = 0.065; P = 0.011). Foveal hypoplasia was present in 24 (80%) of 30 patients and in 1 of 55 control subjects. CONCLUSIONS: ACHM is not a stationary disease. The first signs of cone cell loss occur in early childhood. If intervention becomes available in the future, the present results imply that it should be applied in the first decade.

Comprehensive analysis of the achromatopsia genes CNGA3 and CNGB3 in progressive cone dystrophy.

OBJECTIVE: To investigate whether the major achromatopsia genes (CNGA3 and CNGB3) play a role in the cause of progressive cone dystrophy (CD). DESIGN: Prospective multicenter study. PARTICIPANTS: Probands (N = 60) with autosomal recessive (ar) CD from various ophthalmogenetic clinics in The Netherlands. METHODS: All available ophthalmologic data from the arCD probands were registered from medical charts and updated by an additional ophthalmologic examination. Mutations in the CNGA3 and CNGB3 genes were analyzed by direct sequencing. MAIN OUTCOME MEASURES: CNGA3 and CNGB3 mutations and clinical course in arCD probands. RESULTS: In 3 arCD probands (3/60; 5%) we found 2 mutations in the CNGB3 gene. Two of these probands had compound heterozygous mutations (p.R296YfsX9/p.R274VfsX12 and p.R296YfsX9/c.991-3T>g). The third proband revealed homozygous missense mutations (p.R403Q) with 2 additional variants in the CNGA3 gene (p.E228K and p.V266M). These probands did not have a congenital nystagmus, but had a progressive deterioration of visual acuity, color vision, and photopic electroretinogram, with onset in the second decade. In 6 other unrelated probands, we found 6 different heterozygous amino acid changes in the CNGA3 (N = 4) and CNGB3 (N = 2) gene. CONCLUSIONS: The CNGB3 gene accounts for a small fraction of the later onset progressive form of cone photoreceptor disorders, and CNGA3 may have an additive causative effect. Our data indicate that these genes are involved in a broader spectrum of cone dysfunction, and it remains intriguing why initial cone function can be spared despite similar gene defects. FINANCIAL DISCLOSURE(S): The author(s) have no proprietary or commercial interest in any materials discussed in this article.

Homozygosity mapping reveals PDE6C mutations in patients with early-onset cone photoreceptor disorders.

Cone photoreceptor disorders form a clinical spectrum of diseases that include progressive cone dystrophy (CD) and complete and incomplete achromatopsia (ACHM). The underlying disease mechanisms of autosomal recessive (ar)CD are largely unknown. Our aim was to identify causative genes for these disorders by genome-wide homozygosity mapping. We investigated 75 ACHM, 97 arCD, and 20 early-onset arCD probands and excluded the involvement of known genes for ACHM and arCD. Subsequently, we performed high-resolution SNP analysis and identified large homozygous regions spanning the PDE6C gene in one sibling pair with early-onset arCD and one sibling pair with incomplete ACHM. The PDE6C gene encodes the cone alpha subunit of cyclic guanosine monophosphate (cGMP) phosphodiesterase, which converts cGMP to 5'-GMP, and thereby plays an essential role in cone phototransduction. Sequence analysis of the coding region of PDE6C revealed homozygous missense mutations (p.R29W, p.Y323N) in both sibling pairs. Sequence analysis of 104 probands with arCD and 10 probands with ACHM revealed compound heterozygous PDE6C mutations in three complete ACHM patients from two families. One patient had a frameshift mutation and a splice defect; the other two had a splice defect and a missense variant (p.M455V). Cross-sectional retinal imaging via optical coherence tomography revealed a more pronounced absence of cone photoreceptors in patients with ACHM compared to patients with early-onset arCD. Our findings identify PDE6C as a gene for cone photoreceptor disorders and show that arCD and ACHM constitute genetically and clinically overlapping phenotypes.

Genetic etiology and clinical consequences of complete and incomplete achromatopsia.

OBJECTIVE: To investigate the genetic causes of complete and incomplete achromatopsia (ACHM) and assess the association between disease-causing mutations, phenotype at diagnosis, and visual prognosis. DESIGN: Clinic-based, longitudinal, multicenter study. PARTICIPANTS: Probands with complete ACHM (n = 35), incomplete ACHM (n = 26), or nonspecific ACHM (n = 2) and their affected relatives (n = 18) from various ophthalmogenetic clinics in The Netherlands. METHODS: Ophthalmologic clinical data were assessed over a life time and were registered from medical charts and updated by ophthalmologic examination. Mutations in the CNGB3, CNGA3, and GNAT2 genes were analyzed by direct sequencing. MAIN OUTCOME MEASURES: Genetic mutations and clinical course of ACHM. RESULTS: CNGB3 mutations were identified in 55 of 63 (87%) of probands and all caused premature truncation of the protein. The most common mutation was p.T383IfsX13 (80%); among the 4 other mutations was the novel frameshift mutation p.G548VfsX35. CNGA3 mutations were detected in 3 of 63 (5%) probands; all caused an amino acid change of the protein. No mutations were found in the GNAT2 gene. The ACHM subtype, visual acuity, color vision, and macular appearance were equally distributed among the CNGB3 genotypes, but were more severely affected among CNGA3 genotypes. Visual acuity deteriorated from infancy to adulthood in 12% of patients, leading to 0.10 in 61%, and even lower than 0.10 in 20% of patients. CONCLUSIONS: In this well-defined cohort of ACHM patients, the disease seemed much more genetically homogeneous than previously described. The CNGB3 gene was by far the most important causal gene, and T383IfsX13 the most frequent mutation. The ACHM subtype did not associate with a distinct genetic etiology, nor were any other genotype-phenotype correlations apparent. The distinction between complete and incomplete subtypes of ACHM has no clinical value, and the assumption of a stationary nature is misleading.

Microarray-based mutation detection and phenotypic characterization of patients with Leber congenital amaurosis.

PURPOSE: To test the efficiency of a microarray chip as a diagnostic tool in a cohort of northwestern European patients with Leber congenital amaurosis (LCA) and to perform a genotype-phenotype analysis in patients in whom pathologic mutations were identified. METHODS: DNAs from 58 patients with LCA were analyzed using a microarray chip containing previously identified disease-associated sequence variants in six LCA genes. Mutations identified by chip analysis were confirmed by sequence analysis. On identification of one mutation, all protein coding exons of the relevant genes were sequenced. In addition, sequence analysis of the RDH12 gene was performed in 22 patients. Patients with mutations were phenotyped. RESULTS: Pathogenic mutations were identified in 19 of the 58 patients with LCA (32.8%). Four novel sequence variants were identified. Mutations were most frequently found in CRB1 (15.5%), followed by GUCY2D (10.3%). The p.R768W mutation was found in 8 of 10 GUCY2D alleles, suggesting that it is a founder mutation in the northwest of Europe. In early childhood, patients with AIPL1 or GUCY2D mutations show normal fundi. Those with AIPL1-associated LCA progress to an RP-like fundus before the age of 8, whereas patients with GUCY2D-associated LCA still have relatively normal fundi in their mid-20s. Patients with CRB1 mutations present with distinct fundus abnormalities at birth and consistently show characteristics of RP12. Pathogenic GUCY2D mutations result in the most severe form of LCA. CONCLUSIONS: Microarray-based mutation detection allowed the identification of 32% of LCA sequence variants and represents an efficient first-pass screening tool. Mutations in CRB1, and to a lesser extent, in GUCY2D, underlie most LCA cases in this cohort. The present study establishes a genotype-phenotype correlation for AIPL1, CRB1, and GUCY2D.