Nineteen novel NPHS1 mutations in a worldwide cohort of patients with congenital nephrotic syndrome (CNS).
ABSTRACT: Recessive mutations in the NPHS1 gene encoding nephrin account for approximately 40% of infants with congenital nephrotic syndrome (CNS). CNS is defined as steroid-resistant nephrotic syndrome (SRNS) within the first 90 days of life. Currently, more than 119 different mutations of NPHS1 have been published affecting most exons.We here performed mutational analysis of NPHS1 in a worldwide cohort of 67 children from 62 different families with CNS.We found bi-allelic mutations in 36 of the 62 families (58%) confirming in a worldwide cohort that about one-half of CNS is caused by NPHS1 mutations. In 26 families, mutations were homozygous, and in 10, they were compound heterozygous. In an additional nine patients from eight families, only one heterozygous mutation was detected. We detected 37 different mutations. Nineteen of the 37 were novel mutations (approximately 51.4%), including 11 missense mutations, 4 splice-site mutations, 3 nonsense mutations and 1 small deletion. In an additional patient with later manifestation, we discovered two further novel mutations, including the first one affecting a glycosylation site of nephrin.Our data hereby expand the spectrum of known mutations by 17.6%. Surprisingly, out of the two siblings with the homozygous novel mutation L587R in NPHS1, only one developed nephrotic syndrome before the age of 90 days, while the other one did not manifest until the age of 2 years. Both siblings also unexpectedly experienced an episode of partial remission upon steroid treatment.
Project description:Congenital nephrotic syndrome (CNS) is defined as nephrotic syndrome that manifests within the first 3 months of life. Mutations in the NPHS1 gene encoding nephrin, are a major cause for CNS. Currently, more than 173 different mutations of NPHS1 have been published as causing CNS, affecting most exons.We performed mutation analysis of NPHS1 in a worldwide cohort of 20 families (23 children) with CNS. All 29 exons of the NPHS1 gene were examined using direct sequencing. New mutations were confirmed by demonstrating their absence in 96 healthy control individuals.We detected disease-causing mutations in 9 of 20 families (45%). Seven of the families showed a homozygous mutation, while two were compound heterozygous. In another 2 families, single heterozygous NPHS1 mutations were detected. Out of 10 different mutations discovered, 3 were novel, consisting of 1 splice site mutation and 2 missense mutations.Our data demonstrate that the spectrum of NPHS1 mutations is still expanding, involving new exons, in patients from a diverse ethnic background.
Project description:Mutations in NPHS1, which encodes nephrin, are the main causes of congenital nephrotic syndrome (CNS) in Finnish patients, whereas mutations in NPHS2, which encodes podocin, are typically responsible for childhood-onset steroid-resistant nephrotic syndrome in European populations. Genotype-phenotype correlations are not well understood in non-Finnish patients. We evaluated the clinical presentation, kidney histology, and disease progression in non-Finnish CNS cases by mutational screening in 107 families (117 cases) by sequencing the entire coding regions of NPHS1, NPHS2, PLCE1, WT1, LAMB2, PDSS2, COQ2, and NEPH1. We found that CNS describes a heterogeneous group of disorders in non-Finnish populations. We identified nephrin and podocin mutations in most families and only rarely found mutations in genes implicated in other hereditary forms of NS. In approximately 20% of cases, we could not identify the underlying genetic cause. Consistent with the major role of nephrin at the slit diaphragm, NPHS1 mutations associated with an earlier onset of disease and worse renal outcomes than NPHS2 mutations. Milder cases resulting from mutant NPHS1 had either two mutations in the cytoplasmic tail or two missense mutations in the extracellular domain, including at least one that preserved structure and function. In addition, we extend the spectrum of known NPHS1 mutations by describing long NPHS1 deletions. In summary, these data demonstrate that CNS is not a distinct clinical entity in non-Finnish populations but rather a clinically and genetically heterogeneous group of disorders.
Project description:Congenital nephrotic syndrome (CNS) is de- fined as nephrotic syndrome that manifests at birth or within the first 3 months of life. Most patients develop end-stage renal disease (ESRD) within 2 to 3 years of life. CNS of the Finnish-type (CNF) features a rather specific renal histology and is caused by recessive mutations in the NPHS1 gene encoding nephrin, a major structural protein of the glomerular slit-diaphragm. So far, more than 80 different mutations of NPHS1 causing CNF have been published.Here, we performed mutation analysis of NPHS1 by exon sequencing in a worldwide cohort of 32 children with CNS from 29 different families.Sixteen of the 29 families (55%) were found to have two disease-causing alleles in NPHS1. Two additional patients had a single heterozygous mutation in NPHS1. Thirteen of a total of 20 different mutations detected were novel (65%). These were five missense mutations, one nonsense mutation, three deletions, one insertion and three splice-site mutations.Our data expand the spectrum of known NPHS1 mutations by >15% in a worldwide cohort. Surprisingly, two patients with disease-causing mutations showed a relatively mild phenotype, as one patient had a partial remission with steroid treatment and one patient had normal renal function 1 year after the onset of disease. The increased number of known mutations will facilitate future studies into genotype/phenotype correlations.
Project description:Hereditary nephrotic syndrome often presents with steroid-resistance and onset within the first year of life. Mutations in genes highly expressed in podocytes have been found in two thirds of these patients, especially NPHS1 and NPHS2 among at least 29 genetic causes that have been discovered. We reported two siblings with steroid-resistant nephrotic syndrome caused by co-inheritance of mutations at NPHS1 (c.1339G>A, p.E447K) and ACDK4 (c.748G>C, p.D250H) genes. The siblings presented with steroid-resistant nephrotic syndrome and pathological lesions of focal segmental glomerulosclerosis (FSGS), while the elder sister also developed hypertension, renal failure and cardiac dysfunction.
Project description:Congenital nephrotic syndrome, a rare and severe disease, is inherited as an autosomal recessive trait. The disease manifests shortly after birth and occurs predominantly in families of Finnish origin but has now been observed in all countries and races. Mutations in the NPHS1 gene, which encodes nephrin, are the main causes of congenital nephrotic syndrome in patients. In this study, we report the first mutational analysis of the NPHS1 gene in three unrelated children from three different Vietnamese families. These patients were examined and determined to be suffering from congenital nephrotic syndrome in the Department of Pediatrics, Vietnam National Hospital of Pediatrics. All 29 exons and exon-intron boundaries of NPHS1 were analyzed by PCR and DNA sequencing. Genetic analysis of the NPHS1 gene revealed one compound heterozygous variant p.Glu117Lys, one heterozygous missense mutation p.Asp310Asn, and one heterozygous frame-shifting mutation (c.3250_3251insG causing p.Val1084Glyfs⁎12) in patient 1. In patient 2, one heterozygous variant p.Glu117Lys and one novel heterozygous missense mutation p.Ser324Ala were identified. Finally, a novel missense mutation p.Arg802Leu and a novel nonsense mutation (c.2442C>G causing p.K792⁎) were identified in patient 3.
Project description:Congenital nephrotic syndrome is defined as nephrotic syndrome which manifests in utero or during the first 3 months of life. The prototype of congenital nephrotic syndrome is congenital nephrotic syndrome of Finnish type (CNF, OMIM #602716), which is caused by loss-of-function mutations of the nephrin gene (NPHS1). There have been few clinical case reports of CNF in Korea, but none of which was confirmed by genetic study. Here, we report two children with congenital nephrotic syndrome. Genetic analysis of the NPHS1 gene revealed compound heterozygous frame-shifting mutations (c.2156_2163 delTGCACTGC causing p.L719DfsX4 and c.3250_3251insG causing p.V1084GfsX12) in one patient and a missense mutation (c.1381G>A causing p.R460Q) and a nonsense mutation (c.2442C>G causing p.Y814X) in the other patient. The nonsense mutation was novel. The clinical courses of the patients were typical of CNF. This is the first report of genetically confirmed CNF in Korea to date. The early genetic diagnosis of CNF is important for proper clinical management of the patients and precise genetic counseling of the families.
Project description:BACKGROUND AND OBJECTIVES:Steroid-resistant nephrotic syndrome overwhelmingly progresses to ESRD. More than 30 monogenic genes have been identified to cause steroid-resistant nephrotic syndrome. We previously detected causative mutations using targeted panel sequencing in 30% of patients with steroid-resistant nephrotic syndrome. Panel sequencing has a number of limitations when compared with whole exome sequencing. We employed whole exome sequencing to detect monogenic causes of steroid-resistant nephrotic syndrome in an international cohort of 300 families. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS:Three hundred thirty-five individuals with steroid-resistant nephrotic syndrome from 300 families were recruited from April of 1998 to June of 2016. Age of onset was restricted to <25 years of age. Exome data were evaluated for 33 known monogenic steroid-resistant nephrotic syndrome genes. RESULTS:In 74 of 300 families (25%), we identified a causative mutation in one of 20 genes known to cause steroid-resistant nephrotic syndrome. In 11 families (3.7%), we detected a mutation in a gene that causes a phenocopy of steroid-resistant nephrotic syndrome. This is consistent with our previously published identification of mutations using a panel approach. We detected a causative mutation in a known steroid-resistant nephrotic syndrome gene in 38% of consanguineous families and in 13% of nonconsanguineous families, and 48% of children with congenital nephrotic syndrome. A total of 68 different mutations were detected in 20 of 33 steroid-resistant nephrotic syndrome genes. Fifteen of these mutations were novel. NPHS1, PLCE1, NPHS2, and SMARCAL1 were the most common genes in which we detected a mutation. In another 28% of families, we detected mutations in one or more candidate genes for steroid-resistant nephrotic syndrome. CONCLUSIONS:Whole exome sequencing is a sensitive approach toward diagnosis of monogenic causes of steroid-resistant nephrotic syndrome. A molecular genetic diagnosis of steroid-resistant nephrotic syndrome may have important consequences for the management of treatment and kidney transplantation in steroid-resistant nephrotic syndrome.
Project description:BACKGROUND: Nephrotic syndrome is traditionally classified on the basis of the response to standard steroid treatment. Mutations in more than 24 genes have been associated with nephrotic syndrome in children, although the great majority of steroid-resistant cases have been attributed to mutations in three main genes: NPHS1, NPHS2 and WT1. The aims of this study were to identify mutations in these genes more frequently reported as mutated and to characterize each variation using different in silico prediction algorithms in order to understand their biological functions. METHODS: We performed direct sequence analysis of exons 8 and 9 of WT1, 8 exons of NPHS2 and 29 exons of NPHS1, including NPHS2 and NPHS1 intron-exon boundary sequences, as well as 700 bp of the 5' UTR from both genes in 27 steroid-resistant patients aged between 3 months and 18 years. RESULTS: Analysis of the NPHS2 gene revealed four missense mutations, one frameshift mutation and three variations in the 5' UTR. Four patients presented compound heterozygosis, and four other patients presented one heterozygous alteration only. WT1 and NPHS1 gene analysis did not reveal any mutations. DISCUSSION: This is the first study focusing on genetics of SRNS in Brazilian children. Identification of mutations is important because it could influence physicians' decision on patient treatment, as patients carrying mutations can be spared the side effects of immunosuppressive therapy and ultimately could be considered for kidney transplantation from a living donor. CONCLUSIONS: After molecular analysis of the genes more frequently reported as mutated in 27 steroid-resistant nephrotic syndrome patients, we identified NPHS2 mutations confirming the hereditary character of the kidney disease in only 14.8% of patients. Therefore, the next step is to perform a next generation sequencing based analysis of glomeluropathy-related panel of genes for the remaining patients in order to search for mutations in other genes related to steroid-resistant nephrotic syndrome.
Project description:Mutations in the transmembrane protein nephrin (encoded by NPHS1) underlie nearly half of all cases of congenital nephrotic syndrome (CNS), which is caused by aberrations in the blood filtering function of glomerular podocytes. Nephrin directly contributes to the structure of the filtration barrier, and it also serves as a signaling scaffold in podocytes, undergoing tyrosine phosphorylation on its cytoplasmic tail to recruit intracellular effector proteins. Nephrin phosphorylation is lost in several human and experimental models of glomerular disease, and genetic studies have confirmed its importance in maintenance of the filtration barrier. To date, however, the effect of CNS-associated NPHS1 variants on nephrin phosphorylation remains to be determined, which hampers genotype-phenotype correlations. Here, we have characterized a novel nephrin sequence variant, A419T, which is expressed along with C623F in a patient presenting with CNS. Nephrin localization is altered in kidney biopsies, and we further demonstrate reduced surface expression and ER retention of A419T and C623F in cultured cells. Moreover, we show that both mutations impair nephrin tyrosine phosphorylation, and they exert dominant negative effects on wildtype nephrin signaling. Our findings thus reveal that missense mutations in the nephrin extracellular region can impact nephrin signaling, and they uncover a potential pathomechanism to explain the spectrum of clinical severity seen with mild NPHS1 mutations.
Project description:The aim of our study was to examine NPHS1, NPHS2, WT1 and LAMB2 mutations, previously reported in two thirds of patients with nephrotic syndrome with onset before the age of one year old. Genomic DNA samples from Polish children (n=33) with Steroid-Resistant Nephrotic Syndrome (SRNS) due to focal segmental glomerulosclerosis (FSGS), manifesting before the age of 13 years old, underwent retrospective analysis of NPHS1, NPHS2, WT1 (exons 8, 9 and adjacent exon/intron boundaries) and LAMB2. No pathogenic NPHS1 or LAMB2 mutations were found in our FSGS cohort. SRNS-causing mutations of NPHS2 and WT1 were detected in 7 of 33 patients (21%), including those with nephrotic syndrome manifesting before one year old: five of seven patients. Four patients had homozygous c.413G>A (p.Arg138Gln) NPHS2 mutations; one subject was homozygous for c.868G>A (p.Val290Met) NPHS2. A phenotypic female had C>T transition at position +4 of the WT1 intron 9 (c.1432+4C>T) splice-donor site, and another phenotypic female was heterozygous for G>A transition at position +5 (c.1432+5G>A). Genotyping revealed a female genotypic gender (46, XX) for the first subject and male (46, XY) for the latter. In addition, one patient was heterozygous for c.104dup (p.Arg36Profs*34) NPHS2; two patients carried a c.686G>A (p.Arg229Gln) NPHS2 non-neutral variant. Results indicate possible clustering of causative NPHS2 mutations in FSGS-proven SRNS with onset before age one year old, and provide additional evidence that patients with childhood steroid-resistant nephrotic syndrome due to focal segmental glomerulosclerosis should first undergo analysis of NPHS2 coding sequence and WT1 exons 8 and 9 and surrounding exon/intron boundary sequences, followed by gender genotyping.