Sporadic Retinitis Pigmentosa Research Articles

Whole exome sequencing analysis identifies novel Stargardt disease-related gene mutations in Chinese Stargardt disease and retinitis pigmentosa patients.

To delineate the disease-causing mutations of the Stargardt disease-related genes in Chinese patients diagnosed with Stargardt disease or retinitis pigmentosa (RP) by whole exome sequencing analysis. A total of 123 sporadic RP or Stargardt disease patients and 2 Stargardt disease families were recruited. All sporadic patients and the probands of the families were subjected to whole exome sequencing analysis. The candidate mutations were verified by direct sequencing based on the cosegregation pattern and in 200 control subjects and by the bioinformatics analyses. A total of three reported ABCA4 mutations were identified in the probands of the two Stargardt disease families. The probands and the affected family members with either homozygous or compound heterozygous mutations showed typical Stargardt disease features, which was absent in their unaffected family members. The cosegregation pattern confirmed the mode of recessive inheritance. Moreover, two sporadic Stargardt disease patients were identified to carry two novel ABCA4 and one PROM1 mutations. In addition, 13 novel variants were found in 119 sporadic RP patients in 7 Stargardt disease-related genes, and 8 novel missense variants were conserved across different species and predicted to be damaging to the protein. All 15 novel variants were absent in our 200 control subjects. This study revealed 22.4% study subjects carrying Stargardt disease-related gene mutations with total 15 novel variants in seven Stargardt disease-related genes, assuring that targeted next-generation sequencing analysis is a high throughput strategy to facilitate the clinical diagnosis from suspicious patients and recommended as a routine examination for inherited retinal dystrophies.

Retinitis pigmentosa associated with systemic light chain amyloidosis (AL amyloidosis)

Retinitis pigmentosa (RP) or hereditary retinal dystrophy is a rare disease that can be isolated (non-syndromic RP) or associated with other systemic signs (syndromic RP). Kidney damage is exceptionally reported in patients with RP, particularly in syndromic forms. Association with renal amyloidosis remains unusual with only one reported case of RP and hereditary gelsolin amyloidosis due to a G654A gelsolin mutation defining the new syndrome of Ardalan-Shoja-Kiuru. Apart from this publication, no case associating RP and AL amyloidosis has been found. We report an original case of renal damage revealing kappa-type systemic light chains amyloidosis (AL amyloidosis) in 35-year-old man with sporadic RP. Our observation is, to our knowledge, the first to report this association.

Whole exome sequencing identifies novel USH2A mutations and confirms Usher syndrome 2 diagnosis in Chinese retinitis pigmentosa patients

Retinitis pigmentosa (RP) is a common phenotype in multiple inherited retinal dystrophies (IRD). Disease gene identification can assist the clinical diagnosis of IRD patients for better clinical management, treatment and counseling. In this study, we aimed to delineate and characterize the disease-causing mutations in Chinese familial and sporadic patients with initial diagnosis of RP. Four unrelated Chinese families and 118 sporadic RP patients were recruited for whole exome sequencing analysis. A total of 5 reported and 3 novel USH2A mutations were identified in four Chinese probands. The probands and their family members showed typical RP features and mild to severe hearing impairment, confirming the diagnosis of Usher syndrome 2 (USH). Moreover, 11 sporadic RP patients were identified to carry the compound heterozygous mutations in the USH2A gene, confirming the diagnosis of USH2. The patients carried the truncating mutations had a younger age of first visit than the patients carried only the missense mutations (p = 0.017). In summary, this study revealed 8 novel USH2A variants in Chinese familial and sporadic RP patients, assuring that whole exome sequencing analysis is an adequate strategy to facilitate the clinical diagnosis of USH from the sporadic RP patients.

Genomic Landscape of Sporadic Retinitis Pigmentosa: Findings from 877 Spanish Cases

We aimed to unravel the molecular basis of sporadic retinitis pigmentosa (sRP) in the largest cohort reported to date. Case series. A cohort of 877 unrelated Spanish sporadic cases with a clinical diagnosis of retinitis pigmentosa (RP) and negative family history. The cohort was studied by classic genotyping or targeted next-generation sequencing (NGS). Multiplex ligation-dependent probe amplification (MLPA) and array-based comparative genomic hybridization were performed to confirm copy number variations detected by NGS. Quantitative fluorescent polymerase chain reaction was assessed in sRP cases carrying de novo variants to confirm paternity. The study of the sRP cohort showed a high proportion of causal autosomal dominant (AD) and X-linked (XL) variants, most of them being de novo. Causative variants were identified in 38% of the patients studied, segregating recessively in 84.5% of the solved cases. Biallelic variants detected in only 6 different autosomal recessive genes explained 50% of the cases characterized. Causal AD and XL variants were found in 7.6% and 7.9% of cases, respectively. Remarkably, 20 de novo variants were confirmed after trio analysis, explaining 6% of the cases. In addition, 17% of the solved sRP cases were reclassified to a different retinopathy phenotype. This study highlights the clinical utility of NGS testing for sRP cases, expands the mutational spectrum, and provides accurate prevalence of mutated genes. Our findings evidence the underestimated role of de novo variants in the etiology of RP, emphasizing the importance of segregation analysis as well as comprehensive screening of genes carrying XL and AD variants in sporadic cases. Such in-depth study is essential for accurate family counseling and future enrollment in gene therapy-based treatments.

Next-generation sequencing identifies unexpected genotype-phenotype correlations in patients with retinitis pigmentosa.

Retinitis pigmentosa (RP) is an inherited degenerative disease causing severe retinal dystrophy and visual impairment mainly with onset in infancy or adolescence. Targeted next-generation sequencing (NGS) has become an efficient tool to encounter the enormous genetic heterogeneity of diverse retinal dystrophies, including RP. To identify disease-causing mutations in unselected, consecutive RP patients, we conducted Sanger sequencing of genes commonly involved in the suspected genetic RP subtype, followed by targeted large-panel NGS if no mutation was identified, or NGS as primary analysis. A high (70%) detection rate of disease-causing mutations was achieved in a large cohort of 116 unrelated patients. About half (48%) of the solved RP cases were explained by mutations in four genes: RPGR, EYS, PRPF31 and USH2A. Overall, 110 different mutations distributed across 30 different genes were detected, and 46 of these mutations were novel. A molecular diagnosis was achieved in the majority (82–100%) of patients if the family history was suggestive for a particular mode of inheritance, but only in 60% in cases of sporadic RP. The diagnostic potential of extensive molecular analysis in a routine setting is also illustrated by the identification of unexpected genotype-phenotype correlations for RP patients with mutations in CRX, CEP290, RPGRIP1, MFSD8. Furthermore, we identified numerous mutations in autosomal dominant (PRPF31, PRPH2, CRX) and X-linked (RPGR) RP genes in patients with sporadic RP. Variants in RP2 and RPGR were also found in female RP patients with apparently sporadic or dominant disease. In summary, this study demonstrates that massively parallel sequencing of all known retinal dystrophy genes is a valuable diagnostic approach for RP patients.

Mutation screening of the USH2A gene in retinitis pigmentosa and USHER patients in a Han Chinese population

ObjectivesUSH2A encodes for usherin, a basement membrane protein in the inner ear and retina. USH2A can cause retinitis pigmentosa (RP) with or without hearing loss. The aim of this study was to detect USH2A mutations in a Chinese cohort of 75 small RP families and 10 Usher syndrome families.MethodsWe performed a direct Sanger sequencing analysis of the USH2A gene to identify mutations for this cohort.ResultsWe identified a total of eight mutations in four of the 75 small RP families (5.3%) and two mutations in one of the 10 Usher families (10%); all families were detected to have compound heterozygous mutations. In families with nonsyndromic RP, we identified the compound heterozygous mutations p.Pro4818Leuand p.Leu2395Hisfs*19 in family No. 19114, p.Arg4493His and p.His1677Glnfs*15 in family No.19162, c.8559-2A > G and p.Arg1549* in family No.19123 and p.Ser5060Pro and p.Arg34Leufs*41 in family No.19178. We also identified the heterozygous mutations p.Arg3719His and p.Cys934Trp in family No.19124, which was the Usher syndrome family. These mutations were predicted to be harmful by SIFT, PROVEAN, Mutation Taster or PolyPhen-2.ConclusionsOur results revealed six novel mutations in the USH2A gene in a Chinese population, which is beneficial for the clinical use of genetic testing of USH2A in patients with autosomal-recessive or sporadic RP and Usher syndrome.

Novel Mutations in PRPF31 Causing Retinitis Pigmentosa Identified Using Whole-Exome Sequencing.

The purpose of this study was to investigate the disease-causing mutations for retinitis pigmentosa (RP) patients and function of mutations. We recruited RP families and sporadic RP patients, and performed whole-exome sequencing (WES) to screen for sequence variations. Subsequently, we investigated the expression of green fluorescent protein (GFP) merged expression vectors containing PRPF31 wild type (WT) and its variants. We determined protein stability by cycloheximide (CHX) treatment. Two frameshift variants, c.547delG (p.E183fs) and c.804delG (p.L268fs), and one stopgain variant, c.1060C>T (p.R354X), in the pre-mRNA processing factor 31 gene (PRPF31) were identified in three RP families. In comparison with WT, the expressions of GFP-fused PRPF31 (GFP-PRPF31) protein with the mutation c.547delG or c.804delG in HEK293 cells were significantly reduced. However, the expression of GFP-PRPF31 containing the stopgain mutation (GFP-PRPF31sg) was increased. CHX treatment of HEK293 showed the GFP-PRPF31sg protein was more stable than GFP-PRPF31 WT. The WT protein expression was localized in the nuclei, and the mutants in both nuclei and cytoplasm. We screened for PRPF31 mutations in 131 sporadic RP patients by WES and successfully identified three novel mutations: c.G781C (p.G261R), c.A1373T (p.Q458L), and c.C1222T (p.R408W). Our study revealed novel mutations of PRPF31 in RP. Our results also showed that the two mutations (c.547delG or c.804delG) affect gene expression and GFP-PRPF31sg has increased protein stability.

Analysis of mutations of disease-causing gene and the inheritance in sporadic retinitis pigmentosa

Objective To observe the disease-causing genes and the inheritance in sporadic retinitis pigmentosa (sRP) in Ningxia region. Methods 49 sRP patients and 128 family members were recruited for this study. All the patients and family members received complete ophthalmic examinations including best corrected visual acuity, slit-lamp microscope, indirect ophthalmoscopy, fundus color photography, visual field, optic coherence tomography, full view electroretinogram. DNA was extracted from patients and family members. Using exon combined target region capture sequencing chip to screen the 230 candidate disease-causing gene mutations, polymerase chain reaction and direct sequencing were used to confirm the disease-causing mutations. Results 24/49 patients (49.0%) had identified disease-causing genes, totally 16 genes were involved. There were 41 mutation sites were found, including 32 new mutations (78.0%). The disease-causing genes include USH2A, C2orf71, GNGA1, RPGR1, IFT140, TULP1, CLRN1, RPE65, ABCA4, GUCA1, EYS, CYP4V2, GPR98 and ATXN7. Based on pedigree analysis, 20 patients were autosomal recessive retinitis pigmentosa, 3 patients were autosomal dominant retinitis pigmentosa and 1 patient was X linked retinitis pigmentosa. 3/7 patients with USH2A mutations were identified as Usher syndrome. Conclusions USHZA is the main disease-causing of sRP patients in Ningxia region. 83.3% of sRP in this cohort are autosomal recessive retinitis pigmentosa. Key words: Retinitis pigmentosa/genetics; Inheritance patterns; Genes; Mutation; High-throughput nucleotide sequencing

Novel mutations in PDE6B causing human retinitis pigmentosa.

To identify the genetic defects of a Chinese patient with sporadic retinitis pigmentosa (RP). Ophthalmologic examinations were performed on the sporadic RP patient, 144 genes associated with retinal diseases were scanned with capture next generation sequencing (CNGS) approach. Two heterozygous mutations in PDE6B were confirmed in the pedigree by Sanger sequencing subsequently. The carrier frequency of PDE6B mutations of reported PDE6B mutations based on the available two public exome databases (1000 Genomes Project and ESP6500 Genomes Project) and one in-house exome database was investigated. We identified compound heterozygosity of two novel nonsense mutations c.1133G>A (p.W378X) and c.2395C>T (p.R799X) in PDE6B, one reported causative gene for RP. Neither of the two mutations in our study was presented in three exome databases. Two mutations (p.R74C and p.T604I) in PDE6B have relatively high frequencies in the ESP6500 and in-house databases, respectively, while no common dominant mutation in each of the database or across all databases. We demonstrates that compound heterozygosity of two novel nonsense mutations in PDE6B could lead to RP. These results collectively point to enormous potential of next-generation sequencing in determining the genetic etiology of RP and how various mutations in PDE6B contribute to the genetic heterogeneity of RP.

Whole-exome sequencing reveals a novel frameshift mutation in the FAM161A gene causing autosomal recessive retinitis pigmentosa in the Indian population

Retinitis pigmentosa (RP) is a heterogenous group of inherited retinal degenerations caused by mutations in at least 50 genes. To identify genetic mutations underlying autosomal recessive RP (arRP), we performed whole-exome sequencing study on two consanguineous marriage Indian families (RP-252 and RP-182) and 100 sporadic RP patients. Here we reported novel mutation in FAM161A in RP-252 and RP-182 with two patients affected with RP in each family. The FAM161A gene was identified as the causative gene for RP28, an autosomal recessive form of RP. By whole-exome sequencing we identified several homozygous genomic regions, one of which included the recently identified FAM161A gene mutated in RP28-linked arRP. Sequencing analysis revealed the presence of a novel homozygous frameshift mutation p.R592FsX2 in both patients of family RP-252 and family RP-182. In 100 sporadic Indian RP patients, this novel homozygous frameshift mutation p.R592FsX2 was identified in one sporadic patient ARRP-S-I-46 by whole-exome sequencing and validated by Sanger sequencing. Meanwhile, this homozygous frameshift mutation was absent in 1000 ethnicity-matched control samples screened by direct Sanger sequencing. In conclusion, we identified a novel homozygous frameshift mutations of RP28-linked RP gene FAM161A in Indian population.

Epidemiological characteristics and clinical phenotype of sporadic retinitis pigmentosa

Background Retinitis pigmentosa (RP) is a group of progressive monogenic inheritance disease.Seldom epidemiology is performed to summarize the varied clinical phenotypes,especially some sporadic cases with untypical genetic history. Objective The aim of this survey was to investigate the clinical epidemiological characteristics and phenotype of sporadic RP. Methods A prospective cohort study was designed.A survey of a series of clinically diagnosed sporadic primary RP patients was conducted at the Southwest Eye Hospital from July 2010 to November 2011.A total of 130 patients that matched the inclusion criteria were enrolled in this survey.Clinical ocular examinations and questionnaire surveys were given,including ophthalmoscopic examination,best corrective visual acuity( BCVA ),perimetry and Ganzfield electroretinogram (ERG)and color fundus photo.RP with different phenotypes were classified. Results A total of 130 sporadic RP patients were collected in this survey.Of them,66 were male and 64 were female with a mean age of (36.9±14.4) years.The average onset age of these subjects was (21.2±18.4) years.Seven (5.38％) patients had consanguineous marriage history,and 13 ( 10.00％ )patients had systemic disease.Forty-four (33.85％) patients had outdoor jobs,and 86 (66.15％ ) worked indoor.Eighty-nine patients had typical RP ( 68.5％ ),and the number of patients that developed central RP and sine pigmento RP were 16 ( 12.3％ ) and 16( 12.3％ ),respectively.An absence of a- and b-waves in full-field ERG was detected in 99 (76.15％ ) cases.The longest duration of night blindness was identified in typical RP patients and the lowest BCVA in central RP patients. Conclusions The age at first onset is early in sporadic RP.There are wide variations in different types of RP,but the ERG outcome is specific for all RP types. Key words: Retinitis pigmentosa/sporadic; Epidemiological characteristics; Phenotype; Full-field electroretinogram; Questionnaire

Screening genes of the visual cycle RGR, RBP1 and RBP3 identifies rare sequence variations

The visual cycle is essential for vision and several genes encoding proteins of the cycle have been found mutated in various forms of inherited retinal dystrophy. We screened 3 genes of the visual cycle. RGR, encoding the retinal pigment epithelium (RPE) G protein-coupled receptor acting in vitro as a photoisomerase; RBP1, encoding the ubiquitous cellular retinol binding protein carrying intracellular all-trans retinoids; RBP3, encoding the interphotoreceptor retinoid binding protein, a retinal-specific protein which shuttles all-trans retinol from photoreceptors to RPE and 11-cis retinal from RPE to photoreceptors. We used denaturing high performance liquid chromatography (D-HPLC) and direct sequencing to screen 216 patients (134 with autosomal recessive or sporadic retinitis pigmentosa (RP) and 82 with other retinal dystrophies) for RBP1 and RBP3, and 331 patients for RGR (79 cases with autosomal dominant RP and 36 RP cases with undetermined inheritance were added to the 216 previous patients). Several variants were found in the 3 genes, including unique amino acid changes, but none of them showed evidence of pathogenicity. It is likely that mutations in RGR, RBP3, and possibly RBP1 occur rarely in inherited retinal dystrophies.

Screening for the Carbonic Anhydrase IV Gene Mutations in Chinese Retinitis Pigmentosa Patients

Purpose: To investigate the significance of carbonic anhydrase IV gene mutations in sporadic retinitis pigmentosa (RP) among the Chinese population.Methods: A total of 116 patients with simplex RP and adRP of Chinese ethnicity were screened for mutations in the eight coding exons of the CA4 gene by sequencing. Immunostaining and Real-Time PCR were used to measure the cellular localization and expression level of CA4 variations.Results: Ten sequence alterations were identified, including a novel variant within exon 1 of CA4 (A12T) in a patient with RP. No notable difference was found between wild-type protein and A12T mutant protein in the distribution pattern. However, there was a decrease in level of mRNA expression of 3′ UTR mutant to wild-type.Conclusions: The results suggest that the expression level of CA IV may be important to maintain retina function in RP.

Silencing gene therapy for mutant membrane, secretory, and lipid proteins in retinitis pigmentosa (RP)

A mutation in retinal photoreceptor-specific proteins causes nearly 50% of retinitis pigmentosa (RP) cases; the other 50% is called sporadic RP, the etiology of which is unknown. To alleviate RP development, gene therapies, including insertion of a wild type gene or replacement of mutant genes, have been conducted in animal models. The result was not always satisfactory. We found that a mutant misfolded rhodopsin induces endoplasmic reticulum (ER) stress and that the resultant unfolded protein response (UPR) induces apoptosis in the retinal photoreceptor cells in transgenic S334ter rats. The levels of mutant misfolded rhodopsin, not those of normal rhodopsin appear to control disease development. Since mutant misfolded rhodopsin induces ER stress, it is essential to eliminate even minute levels of the mutant misfolded rhodopsin from retinal photoreceptor cells. We speculate that a complete elimination of mutant rhodopsin is more important than an insertion of a normal gene. Here, we propose a silencing gene therapy for complete elimination of mutant unfolded rhodopsin from photoreceptor cells. To achieve this therapy, custom synthesized double-stranded small interfering (si)RNAs will be used to silence the mutant rhodopsin mRNA. The silencing gene therapy will be applicable to many inherited diseases caused by ER stress induced by mutant membrane, secretory, and lipid proteins.

Mutations P51U and G122E in retinal transcription factor NRL associated with autosomal dominant and sporadic retinitis pigmentosa.

Retinitis pigmentosa (RP) is the most frequent form of inherited retinopathy. RP is genetically heterogeneous with autosomal dominant, autosomal recessive and X-linked forms. Autosomal dominant retinitis pigmentosa (adRP) accounts for about 20-25% of all RP cases. At least ten adRP loci have so far been mapped. However, mutations causing adRP have been identified only in four retina-specific genes: RHO (encoding rhodopsin) in approximately 20% of adRP families, peripherin/RDS (3-5% of adRP) and recently RP1 (Pierce et al., 1999, Sulivan et al., 1999) and NRL gene. Only one mutation in the NRL gene causing adRP has so far been reported (Bessant et al., 1999). Here we report a novel mutation Pro51Leu in an adRP Spanish family supporting that mutation in NRL is the cause of adRP. A second missense mutation Gly122Glu has been observed in a simplex RP patient that may represent a sporadic case of retinitis pigmentosa. Hum Mutat 17:520, 2001.

160Thr Mutation in the Rhodopsin Gene Associated withRet initis pigmentosa

Mutations in the rhodopsin gene were studied in 23 unrelated Spanish patients with sporadic retinitis pigmentosa (RP). A codon 160 Thr C→A transition was found in 4 of the 23 patients vs. none of the 159 controls (p < 0.001) suggesting that this mutation may be an informative marker in RP.

Clinical and genetic heterogeneity in retinitis pigmentosa

The clinical course of defective vision and blindness has been investigated in relation to different modes of genetic transmission in a large series of 93 families with retinitis pigmentosa (RP). For autosomal dominant RP, two clinical subtypes could be distinguished according to the delay in macular involvement. In the severe form, macular involvement occurred within 10 years, while in the mild form, macular involvement occurred after 20 years. Interestingly, a significant increase of mean paternal age (38.8 years, mean controls in France = 29.1 years, P less than 0.001) was found in this form of RP, a feature which is suggestive of new mutations. For autosomal recessive RP, four significantly different clinical subtypes could be recognized, according to both age of onset and the pattern of development (P less than 0.001), namely cone-rod dystrophy and early-onset severe forms on the one hand (mean age of onset = 7.6 years), late-onset mild forms and senile forms on the other. Similarly, two significantly different clinical subtypes could be recognized in X-linked RP, according to both mode and age of onset, which were either myopia (mean age = 3.5 +/- 0.5 years) or night blindness (mean age = 10.6 +/- 4.1 years. P less than 0.001). By contrast, no difference was noted regarding the clinical course of the disease, which was remarkably severe whatever the clinical subtype (blindness before 25 years). In addition, all obligate carriers in our series were found to have either severe myopia or pigment deposits in their peripheral retina. Finally, sporadic RP represented the majority of cases in our series (42%). There was a considerable heterogeneity in this group, and at least three clinical forms could be recognized, namely cone-rod dystrophy, early onset-severe forms and late onset moderate forms. At the beginning of the disease, the hereditary nature of the sporadic forms was very difficult to ascertain (especially between 7-10 years) and only the clinical course could possibly provide information regarding the mode of inheritance. However, the high level of consanguinity, and the high sex ratio in early onset and severe sporadic forms (including cone-rod dystrophy), was suggestive of an autosomal or X-linked recessive inheritance, while increased paternal age in late onset forms was suggestive of autosomal dominant mutations.

The Frequency of Posterior Subcapsular Cataract in the Hereditary Retinal Degenerations

I analyzed the frequency and severity of posterior subcapsular cataracts in 291 patients with various forms of hereditary retinal degeneration, including typical retinitis pigmentosa (rod-cone degeneration), cone-rod degeneration, Usher's syndrome, and choroideremia. The overall frequency of posterior subcapsular cataract was 41%, less than generally thought. Posterior subcapsular cataracts were least common in cone-rod degeneration, and most common in the group with autosomal dominant retinitis pigmentosa. Only in the group with sporadic retinitis pigmentosa was a significant sex preference noted; females were affected more often (P less than or equal to .002). Severity was related to age of the patient and duration of disease in those affected. Posterior subcapsular cataract formation is unlikely to be an intrinsic expression of the various genes of hereditary retinal degeneration.