CEP290 Research Articles

Novel biallelic loss-of-function variants in CEP290 cause Joubert syndrome in two siblings

BackgroundJoubert syndrome (JS) is a rare genetic disorder, which can be defined by brain stem malformation, cerebellar vermis hypoplasia, and consequent “molar tooth sign” (MTS). JS always shares variety of phenotypes in development defects. With the development of next-generation sequencing, dozens of causative genes have been identified to JS so far. Here, we investigated two male siblings with JS and uncovered a novel pathogenesis through combined methods.ResultsThe siblings shared similar features of nystagmus, disorders of intellectual development, typical MTS, and abnormal morphology in fourth ventricle. Whole-exome sequencing (WES) and chromosome comparative genomic hybridization (CGH) were then performed on the proband. Strikingly, a maternal inherited nonsense variant (NM_025114.3: c.5953G>T [p.E1985*]) in CEP290 gene and a paternal inherited deletion in 12q21.32 including exons 1 to 10 of CEP290 gene were identified in the two affected siblings. We further confirmed the two variants by in vitro experiments: quantitative PCR and PCR sequencing.ConclusionsIn this study, we first reported a novel causative mechanism of Joubert syndrome: a copy number variation (CNV) combined with a single-nucleotide variant in CEP290 gene, which can be helpful in the genetic diagnosis of this disease.

Results of targeted next-generation sequencing in children with cystic kidney diseases often change the clinical diagnosis.

Cystic kidney diseases are a very heterogeneous group of chronic kidney diseases. The diagnosis is usually based on clinical and ultrasound characteristics and the final diagnosis is often difficult to be made. Next-generation sequencing (NGS) may help the clinicians to find the correct final diagnosis. The aim of our study was to test the diagnostic yield of NGS and its ability to improve the diagnosis precision in a heterogeneous group of children with cystic kidney diseases. Next-generation sequencing of genes responsible for the formation of cystic kidneys was performed in 31 unrelated patients with various clinically diagnosed cystic kidney diseases gathered at the Department of Pediatrics of Motol University Hospital in Prague between 2013 and 2018. The underlying pathogenic variants were detected in 71% of patients (n = 22), no or only one (in case of autosomal recessive inheritance) pathogenic variant was found in 29% of patients (n = 9). The result of NGS correlated with the clinical diagnosis made before the NGS in 55% of patients (n = 17), in the remaining 14 children (45%) the result of NGS revealed another type of cystic kidney disease that was suspected clinically before or did not find causal mutation in suspected genes. The most common unexpected findings were variants in nephronophthisis (NPHP) genes in children with clinically suspected autosomal recessive polycystic kidney disease (ARPKD, n = 4). Overall, 24 pathogenic or probably pathogenic variants were detected in the PKHD1 gene, 8 variants in the TMEM67 gene, 4 variants in the PKD1 gene, 2 variants in the HNF1B gene and 2 variants in BBS1 and NPHP1 genes, respectively. NGS is a valuable tool in the diagnostics of various forms of cystic kidney diseases. Its results changed the clinically based diagnoses in 16% (n = 5) of the children.

P1799GENOTYPE-PHENOTYPE CORRELATION IN A PEDIATRIC ADPKD COHORT

Abstract Background and Aims The correlation between genotype and phenotype is well described in ADPKD adults. PKD2 is milder than PKD1 disease, with end stage kidney disease (ESKD) occurring on average 20 years later, and patients with PKD1 truncating mutations having a more severe outcome than PKD1 non-truncating mutations. Still, large differences in outcome occur even within families carrying the same gene variation. Only a few cases series reported the genetic profile of severely affected ADPKD children and suggest an additional effect of hypomorphic genes. We therefore aim to analyse the geno-phenotype profile in a well characterized pediatric ADPKD cohort. Method Clinical, familial, biological and imaging data were collected longitudinally in children diagnosed with ADPKD. Genotypic analysis was done using a custom Agilent SureSelect gene panel containing 136 ciliopathy-associated genes, including PKD1 and PKD2. Mutations and/or variants identified were individually evaluated for pathogenicity. Results 57 ADPKD children from 44 families were diagnosed at a mean (± SD) age of 4.1 (±4.8) years. ADPKD diagnosis was made in 32 children (56%) because of asymptomatic screening as requested by the family; 7 (12%) due to presenting symptoms (6 due to urinary tract infection and 1 due to post-traumatic macroscopic hematuria); 9 (16%) due to a coincidental finding of renal cysts on US performed for another reason and in 9 cases (16%) a prenatal diagnosis was performed. Twenty-nine children (51%) met the definition of very-early onset (VEO) disease. We identified pathogenic mutations in 100% of our patients, in which the prevalence of PKD1 truncating, PKD1 non-truncating, PKD2 and GANAB mutations was 75%, 19%, 4%, and 2%, respectively. Four cases (7%) were due to a de novo mutation. Interestingly, in 29 patients (51%) the germline mutation was the only identified mutation. However, in the rest of the subjects additional variants were identified in other ciliopathy-associated genes. In 12 cases (21%) the additional identified variants found in either the PKD, PMM2, HNF1B, DNAJC1, CEP290, NEK1, MKKS, NPHP4 or PKHD1 genes were scored to have a potential phenotypic effect, which will be evaluated by continued follow-up of this cohort. Conclusion We report the first large cohort of genotyped ADPKD children, including an extensive panel of ciliopathy genes next to the PKD genes. Interestingly, we found a high prevalence of additional and potentially modifying variants in this young population.

Generation of two human induced pluripotent stem cell lines derived from two juvenile nephronophthisis patients with NPHP1 deletion.

Juvenile nephronophthisis is an inherited renal ciliopathy, causing cystic kidney disease, renal fibrosis, and end-stage renal failure. Human induced pluripotent stem cell (hiPSC) lines, derived from two Juvenile nephronophthisis patients, were generated from peripheral blood mononuclear cells by episomal plasmid vectors. Generated hiPSC lines showed self-renewal and pluripotency and carried a large deletion in NPHP1 (Nephrocystin 1) gene. Since the molecular pathogenesis caused by NPHP1 dysfunction remains unclear, these cell resources provide useful tools to establish disease models and to develop new therapies for juvenile nephronophthisis.

A case report of NPHP1 deletion in Chinese twins with nephronophthisis

BackgroundNephronophthisis (NPHP) is a rare autosomal recessive inherited disorder with high heterogeneity. The majority of NPHP patients progress to end-stage renal disease (ESRD) within the first three decades of life. As an inherited disorder with highly genetic heterogeneity and clinical presentations, NPHP still poses a challenging task for nephrologists without special training to make a well-judged decision on its precise diagnosis, let alone its mechanism and optimal therapy.Case presentationA Chinese family with NPHP was recruited in current study. The clinical characteristics (including findings from renal biopsy) of NPHP patients were collected from medical records and the potential responsible genes were explored by the whole exome sequencing (WES). A homozygous deletion of NPHP1 (1–20 exons) was found in both affected patients, which was further confirmed by quantitative PCR.ConclusionsHomozygous full gene deletion of the NPHP1 gene was identified in a Chinese family with NPHP, which was the molecular pathogenic basis of this disorder. Furthermore, identification of the pathogenic genes for those affected patients can help to have a full knowledge on NPHP’s molecular mechanism and precise treatment.

Difference of genomic copy numbers alterations between hairy cell leukemia-variant and classical hairy cell leukemia: a pilot retrospective study in Chinese.

Background: Hairy cell leukemia (HCL) is a rare chronic B-cell lymphoproliferative disorder. It has two pathological subtypes: classical HCL (HCL-C) and HCL-variant (HCL-V). HCL-C and HCL-V are distinct in morphology and immunophenotype. Their differentiation is important for patient management and clinical outcome, with HCL-V responding poorly to conventional HCL treatments. Recently, whole genomic sequencing has been used to identify the difference between HCL-C and HCL-V and mutation of BRAFV600E has been proved to be a molecular hallmark of HCL-C. However, BRAF inhibitors were not effective in all HCL-C cases and HCL-V seems be lack of the high-frequency mutations. Therefore, it is necessary to compare the genomic changes between HCL-C and HCL-V by high-resolution studies, especially in Chinese population, the genomic alterations of HCL have rarely be reported.Methods: In this study, the clinical features of a total of 18 Chinese HCL patients were described. Single nucleotide polymorphism (SNP) array analysis was performed to evaluate the genomic copy number alterations (CNA) and copy neutral loss of heterozygosity (CN-LOH) on six HCL-Vs with CD25-/BRAFV600E- and four HCL-Cs with CD25+/BRAFV600E+.Results: A total of 24 CNAs including seven chromosomal gains and 17 chromosomal losses, and 22 CN-LOHs were revealed. Five of the six cases of HCL-V showed 15 CNAs including four cryptic chromosomal gains and 11 chromosomal losses. Overlapping regions involving micro-deletion of chromosome 2q13 and large chromosomal loss of 14q were showed in HCL-V. In HCL-C, a total of nine CNAs were revealed in three of the four cases including three chromosomal gains and six chromosomal losses. No overlapping area was observed among the CNVs. 15 CN-LOHs were showed in five of the six cases of HCL-V and seven CN-LOHs was demonstrated in all of the four HCL-Cs.Conclusions: Comparing to Westerners, a relatively higher proportion of HCL-V in all HCL is observed in this study. CNAs and CN-LOHs were common in both HCL-V and HCL-C but the CNAs were different in them. HCL-C was characterized with the higher ratio of large chromosomal changes but lacked of recurrent CNAs, while HCL-V was presented with the higher incidence of cryptic CNAs and recurrent CNAs involving tumor-associated genes. It is necessary to further investigate the association of the genes, such as NPHP1 and TRAF3 genes, and HCL-V in the future study.

A phenotypic and genetic study on β-propeller protein-associated neurodegeneration

Objective: To summarize the clinical and genetic features of β-propeller protein-associated neurodegeneration (BPAN). Methods: The clinical data of 17 patients with BPAN with WDR45 gene variants were retrospectively collected at Children's Hospital of Fudan University, Peking University First Hospital, Capital Institute of Pediatrics, Shengjing Hospital of China Medical University and Shanghai Children's Hospital from June 2016 to December 2018, and their clinical manifestations, electroencephalogram, neuroimaging and genetics were analyzed. Results: Seventeen cases (13 females, 4 males), aged 1.1-8.8 years, were included. The median age of seizure onset was 14.5 months, from 3 months to 24 months of age, manifested with epileptic spasm in 6 cases and focal seizures in 5 cases. Eight patients had only one seizure type and 8 patients had two or more seizure types. Nine patients had complete remission of seizures. All 16 patients with seizures had developmental delay before the seizure onset, of whom 13 patients had moderate to severe seizures. The brain magnetic resonance imaging (MRI) was abnormal in 13 patients, including cerebral atrophy (10 cases) and thinning of the corpus callosum (9 cases). The brain magnetic susceptibility weighted imaging (SWI) in preschool stage showed prominent T2 hypointense signals in bilateral globus pallidus and brainstem ventral in two cases. Five seizure types (spasm, focal, absence, myodonic and generalized tonic clonic seizures)were found on ictal electroencephalogram(EEG) recordings. Compared to female patients(17(6-24) months of ege), male cases had earlier seizure onset (3, 4, 5, 18 months of age) . All patients had de novo variations in WDR45(6 nonsense, 4 frameshift, 3 missense and 4 splicing variations), with hemizygous variants in 3 males, mosaic variants in a male and heterozygous variants in 13 females, within which 5 variations had not been reported (c.977-1C>T,c.976+1G>C,c.10C>T,c.806del and c.110T>C). Conclusions: The patients with BPAN have profound developmental delay and are vulnerable to seizures. The male patients with BPAN tend to have more severer clinical phenotype than females. Early brain SWI could facilitate the timely diagnosis of this disease.

Copy-number variation of the NPHP1 gene in patients with juvenile Nephronophthisis

ABSTRACT Objective: Juvenile nephronophthisis (NPHP) is an autosomal recessive cystic disease of the kidney. It represents the most frequent genetic cause of chronic renal failure in children. Methods: we investigated clinical and molecular features in two children with Juvenile nephronophthisis using firstly Multiplex ligation-dependent probe amplification (MLPA) and secondly multiplex PCR. Results: we report a homozygous NPHP1 deletion in two children. Conclusion: NPHP1 deletion analysis using diagnostic methods (e.g. MLPA, Multiplex PCR) should always be considered in patients with nephronophthisis, especially from consanguineous families. Our results provide insights into genotype-phenotype correlations in juvenile nephronophthisis that can be utilized in genetic counseling.

Gene mutation and clinical analysis of nephronophthisis diagnosed using whole exome sequencing: Experience from China .

Nephronophthisis (NPH) is an autosomal recessive cystic kidney disease. Its onset is obscure, and its early clinical manifestations and pathological changes lack specificity, which makes clinical diagnosis difficult. At present, as many as 90 genetic alterations can result in NPH, which exhibits significant genetic heterogeneity. Therefore, high-throughput sequencing technology provides an effective method to identify and characterize novel NPH pathogenic genes when compared to Sanger sequencing. This study summarizes the gene mutations and clinical data of whole exome sequencing, which was used to diagnose 5 NPH patients to improve the understanding of the causative genes and clinical phenotypes of NPH. The clinical manifestations, laboratory examination indexes, and imaging data of 5 patients of NPH were reported. Whole exome sequencing was performed in 5 children, and the causative genes and mutation sites were analyzed by bioinformatics and genetics. The mutation sites were verified in children and their parents using Sanger direct sequencing. Among the 5 patients (3 male and 2 female), 2 patients had infantile NPH, and 3 patients had juvenile NPH. The 2 infantile NPH patients were characterized by the onset of liver dysfunction accompanied by hypertension and left ventricular change, and the renal function progressed to end-stage renal disease (ESRD) after 7months and 9months, respectively. The 2 cases of infantile NPH had NPHP3 mutations, with one carrying compound heterozygous mutations (c.1358A>G, c.2369A>G) and the other simultaneously carrying a c.1174C>T IVS26-3A>G cleavage site mutation from the father and a nonsense mutation (p.392R>X, 939) from the mother. The 2 juvenile NPH children had entered ESRD at the onset of the disease, including 1 patient with Joubert syndrome. The 2 patients with juvenile NPH had frameshift mutations (c.1583 to 1596: deletion) and homozygous point mutations (7 c.640G>T) of the NPHP1 gene. In addition, another patient with frequent urination and nocturia resulting in stage CKD3 renal function had a complex heterozygous mutation of the NPHP2 gene (c.2686G>A, c.1943A>G). The urine A1MU/creatinine and urinary transferrin increased in all 5 patients without hematuria. Whole exome sequencing identified the causative genes of NPH in 5 children. In NPH children with NPHP3 gene mutations, renal functional damage was characterized by early onset and rapid progression to ESRD, often accompanied by liver dysfunction and hypertension.

The N-Terminal Domain of NPHP1 Folds into a Monomeric Left-Handed Antiparallel Three-Stranded Coiled Coil with Anti-apoptotic Function.

Mutations in the NPHP1 gene, coding for human nephrocystin-1 (NPHP1), cause the autosomal recessive disease nephronophthisis, the most common cause of end-stage renal disease in children and adolescents. The function and structure of NPHP1 are still poorly characterized. NPHP1 presents a modular structure well in keeping with its role as an adaptor protein: it harbors an SH3 domain flanked by two glutamic acid-rich regions and a conserved C-terminal nephrocystin homology domain (NHD). Similar to other NPHP protein family members, its N-terminus contains a putative coiled-coil domain (NPHP1CC) that is supposed to play an important role in NPHP1 self-association and/or protein-protein interactions. Structural studies proving its structure and its oligomerization state are still lacking. Here we demonstrate that NPHP1CC is monomeric in solution and unexpectedly folds into an autonomous domain forming a three-stranded antiparallel coiled coil suitable for protein-protein interactions. Notably, we found that the NPHP1CC shares remarkable structural similarities with the three-stranded coiled coil of the BAG domain protein family, which is known to mediate the anti-apoptotic function of these proteins, suggesting a possible similar role for NPHP1CC. In agreement with this hypothesis, we show that in the context of the full-length protein the NPHP1CC is fundamental to regulate resistance to apoptotic stimuli.

Orphan designation: Recombinant adeno-associated viral vector serotype 5 encoding Staphylococcus aureus Cas9 endonuclease and two guide RNAs complementary to two regions of intron 26 of the CEP290 gene, Treatment of leber's congenital amaurosis

Orphan designation: Recombinant adeno-associated viral vector serotype 5 encoding Staphylococcus aureus Cas9 endonuclease and two guide RNAs complementary to two regions of intron 26 of the CEP290 gene, Treatment of leber's congenital amaurosis

Two novel homozygous mutations in NPHP1 lead to late onset end-stage renal disease: a case report of an adult nephronophthisis in a Chinese intermarriage family

BackgroundNephronophthisis (NPHP) is an autosomal recessive hereditary disease with highly variable clinical characteristics for which 20 genes (NPHP1–20) have been identified. NPHP1 is the major subtype leading to pediatric end-stage renal disease (ESRD). Reports of adult NPHP1 are rare.Case presentationHere, we report a 27-year-old male from a Chinese intermarriage family who was diagnosed as NPHP from clinical presentations and molecular genetic analysis by whole-exome sequencing. The genetic investigation revealed a novel homozygous nonsense mutation, p. E697X,37 and a novel homozygous missense mutation, p. F691 L, in the NPHP1 gene. His parents and fraternal twin harbored heterozygous mutations of the two loci and had no renal symptoms. His elder sister developed ESRD and died at 23 years of age.ConclusionsThe report indicated that adult NPHP should be taken into consideration for adults with ESRD of uncertain cause. The genotype-phenotype correlation requires further investigation.

Significance of CEP78 and WDR62 gene expressions in differentiated thyroid carcinoma: Possible predictors of lateral lymph node metastasis.

This study aimed at investigating the clinical significance of CEP78 and WDR62 in differentiated thyroid carcinoma (DTC). This study also aimed at finding predictors that help in detecting patients with DTC who have high risk for lateral lymph node metastasis (LNM). Quantitative real-time polymerase chain reaction (RT-qPCR) was performed to examine CEP78, and WDR62 mRNA expressions in 40 tissue specimens of DTC, and 40 goiter tissue specimens. Additionally, we reviewed clinical, ultrasound, laboratory, pathological data of patients to analyze the associations between these characteristics and lateral LNM. Our results demonstrated that relative CEP78 mRNA levels were significantly decreased in thyroid cancer tissues than goiter tissues (P=0.002). ROC curve analysis confirmed the diagnostic value of CEP78 mRNA expression, providing an AUC equals to 0.698 (95% confidence intervals (CI), 0.583-0.813; P=0.002). The relative WDR62 mRNA expression was not statistically different in DTC tissues and goiter tissues (P=0.686). Furthermore, the DTC patients had been included to examine risk factors for lateral LNM. In multivariate analysis, the significant factors for predicting lateral LNM were low CEP78 mRNA expression (cut off value ≤0.54; P=0.03; OR=19.62; 95% CI, 1.3-296.23), central LNM (P=0.011; OR=33.6; 95% CI, 2.24-503.6) and calcifications (P=0.023; OR=27.187; 95% CI, 1.57-469.5). CEP78 can be used as a promising molecular biomarker for differentiation between DTC and goiter tissues, in addition it might serve as a predictor of lateral LNM in DTC along with central LNM and calcifications. Unlike CEP78, WDR62 mRNA expression was not statistically different in DTC and goiter.

Diagnosing neurodegeneration with brain iron accumulation before iron starts to accumulate

Introduction
 Neurodegeneration with brain iron accumulation (NBIA) consists of a heterogeneous group of disorders with brain iron accumulation as common radiological endpoint. Mutations in multiple genes have been associated with NBIA. We present 2 cases with a different type of NBIA, in whom the diagnosis was confirmed before brain iron accumulation became evident on MRI.
 Case description
 The first patient was referred because of frequent falls at the age of 4 years. She had an ataxic gait and weak Achilles tendon reflexes. Two years later, pyramidal and more prominent cerebellar signs became evident. A skin and muscle biopsy revealed intra-axonal spheroids in the peri-and endomysial myelinated nerve bundles as well as in the motor endplates, which led to the diagnosis of PLA2G6-associated neurodegeneration (PLAN). Brain iron accumulation occurred at follow-up MRI at 9 years of age.
 The second patient was referred because of developmental stagnation and detection of elevated liver enzymes at 3 months of age. Seizures started at 15 months of age, and were refractory to treatment with multiple anti-epileptic drugs. Molecular genetic testing using an epilepsy gene panel revealed a mutation in the WDR45 gene, a known cause of beta-propeller protein-associated neurodegeneration (BPAN). Brain MRI at 14 months of age showed diffuse hypomyelination in the absence of BIA.
 Discussion
 This report highlights that NBIA can be suspected on a clinical basis and confirmed by genetic testing before iron accumulation becomes present on brain MRI. Early diagnosis will provide a longer timeframe for potential disease modulating treatments in the future.

Partial sequencing of ESR1 and CDK5RAP2 genes in dogs with mammary tumours

Canine mammary tumours (CMT) are among the most common canine cancer types in female dogs. Dogs provide an adaptable model system for human breast cancer studies. It is important to identify the underlying genetic basis to improve knowledge of pathways related to cancer pathogenesis in both species. In this study, we investigated CMT associated single nucleotide polymorphisms (SNP) in target regions of ESR1 and CDK5RAP2 genes. Partial sequencing of two genes in 25 cases with mammary tumours and 10 dogs with healthy mammary glands was performed. Two previously reported SNPs in ESR1 gene and one previously reported SNP and two novel SNPs were genotyped downstream CDK5RAP2 gene. According to the association analysis performed in cases and controls, no statistically significant association was found between these SNPs and CMTs. Comparison of the results from other studies revealed the genetic heterogeneity of ESR1 and CDK5RAP2 between different dog breeds. Larger datasets of different breeds should be analyzed in further studies to identify the possible effects of the two genes in mammary tumour development.

A novel WDR62 missense mutation in microcephaly with abnormal cortical architecture and review of the literature.

Autosomal recessive primary microcephaly (MCPH) is a group of rare neurodevelopmental diseases with severe microcephaly at birth. One type of the disorder, MCPH2, is caused by biallelic mutations in the WDR62 gene, which encodes the WD repeat-containing protein 62. Patients with WDR62 mutation may have a wide range of malformations of cortical development in addition to congenital microcephaly. We describe two patients, a boy and a girl, with severe congenital microcephaly, global developmental delay, epilepsy, and failure to thrive. MRI showed hemispherical asymmetry, diffuse pachygyria, thick gray matter, indistinct gray-white matter junction, and corpus callosum and white matter hypoplasia. Whole exome sequencing revealed the same novel homozygous missense mutation, c.668T>C, p.Phe223Ser in exon 6 of the WDR62 gene. The healthy parents were heterozygous for this mutation. The mutation affects a highly conserved region in one of the WD repeats of the WDR62 protein. Haplotype analysis showed genetic relatedness between the families of the patients. Our findings expand the spectrum of mutations randomly distributed in the WDR62 gene. A review is also provided of the brain malformations described in WDR62 mutations in association with congenital microcephaly.

Gene therapy for retinal dystrophy.

Counteracting splice defects in the CEP290 gene using RNA antisense oligonucleotides or Cas9-mediated gene editing is a therapeutic strategy for Leber congenital amaurosis type 10—a severe untreatable retinal dystrophy leading to childhood blindness.

Development of a gene-editing approach to restore vision loss in Leber congenital amaurosis type 10.

Leber congenital amaurosis type 10 is a severe retinal dystrophy caused by mutations in the CEP290 gene1,2. We developed EDIT-101, a candidate genome-editing therapeutic, to remove the aberrant splice donor created by the IVS26 mutation in the CEP290 gene and restore normal CEP290 expression. Key to this therapeutic, we identified a pair of Staphylococcus aureus Cas9 guide RNAs that were highly active and specific to the human CEP290 target sequence. In vitro experiments in human cells and retinal explants demonstrated the molecular mechanism of action and nuclease specificity. Subretinal delivery of EDIT-101 in humanized CEP290 mice showed rapid and sustained CEP290 gene editing. A comparable surrogate non-human primate (NHP) vector also achieved productive editing of the NHP CEP290 gene at levels that met the target therapeutic threshold, and demonstrated the ability of CRISPR/Cas9 to edit somatic primate cells in vivo. These results support further development of EDIT-101 for LCA10 and additional CRISPR-based medicines for other inherited retinal disorders.

Thumb duplication: molecular analysis of different clinical types.

Molecular analysis of different types of thumb duplication and identification of new suspected gene mutations. In a series of patients operated for polydactyly, DNA was extracted from blood samples collected preoperatively. Among these, the samples of two patients with thumb duplication (Wassel types III and IV) were initially selected for molecular analysis. The method of Clinical Exome Solution was used for the study of the phenotype-involved genes. Next-generation sequencing (NGS) was performed on a NextSeq-500 Platform (Illumina), and Sophia DDM® SaaS algorithms were used for the bioinformatics analysis of the data. In total, 8-including 4 new-mutations were detected in CEP290 (1 mutation), RPGRIP1 (2 mutations), TMEM216 (2 mutations), FBN1 (1 mutation), CEP164 (1 mutation), and MEGF8 (1 mutation) genes. NGS revealed 3 mutated genes in the patient with Wassel III thumb duplication and 5 mutated genes in the patient with Wassel IV duplication. The molecular analysis revealed that the patients had 2 mutated genes in common, but they only shared one common mutation. The new detected mutations are most probably associated with thumb duplication, as they belong to genes with already described mutations causing ciliopathies, often including polydactyly in their phenotype. Recognition of these mutations will be helpful to prenatal diagnosis, operative treatment strategy prediction, and possible future experimental applications in gene therapy.

Relative frequency of inherited retinal dystrophies in Brazil

Among the Brazilian population, the frequency rates of inherited retinal dystrophies and their causative genes are underreported. To increase the knowledge about these dystrophies in our population, we retrospectively studied the medical records of 1,246 Brazilian patients with hereditary retinopathies during 20 years of specialized outpatient clinic care. Of these patients, 559 had undergone at least one genetic test. In this cohort, the most prevalent dystrophies were non-syndromic retinitis pigmentosa (35%), Stargardt disease (21%), Leber congenital amaurosis (9%), and syndromic inherited retinal dystrophies (12%). Most patients had never undergone genetic testing (55%), and among the individuals with molecular test results, 28.4% had negative or inconclusive results compared to 71.6% with a conclusive molecular diagnosis. ABCA4 was the most frequent disease-causing gene, accounting for 20% of the positive cases. Pathogenic variants also occurred frequently in the CEP290, USH2A, CRB1, RPGR, and CHM genes. The relative frequency rates of different inherited retinal dystrophies in Brazil are similar to those found globally. Although mutations in more than 250 genes lead to hereditary retinopathies, only 66 genes were responsible for 70% of the cases, which indicated that smaller and cheaper gene panels can be just as effective and provide more affordable solutions for implementation by the Brazilian public health system.