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Introduction: Unlike idiopathic nephrotic syndrome (NS), hereditary podocytopathies are not expected to recur after kidney transplantation. However, some reports of posttransplant recurrence of NS in patients carrying variants in the NPHS2 gene have been described, notably with the p.Arg138Gln variant, which is more prevalent in Europe. The objective of this study was to assess the risk of recurrence after kidney transplantation in a large cohort of patients with biallelic NPHS2 pathogenic variants. Methods: Since January 2010, 61 patients identified at Necker-Enfants Malades Hospital and 56 enrolled in the PodoNet Registry with biallelic variants in the NPHS2 gene were transplanted and were compared with 44 transplanted children with steroid-resistant NS (SRNS) without any identified pathogenic variant. Results: Of the 117 patients, 23 carried the p.Arg138Gln variant in the homozygous state and 16 in the compound heterozygous state. The other 78 patients carried different variants in the homozygous (n = 44) or compound heterozygous state. Only 1 patient with NPHS2-related SRNS experienced posttransplant recurrence (median follow-up of cohort 8.5 years [2.5-15]). Conversely, 7 of 44 patients (16%) without any identified pathogenic variant recurred within a maximum of 7 days after transplantation (median follow-up 8.9 years [0.6-13.9]). Conclusion: In this large cohort, the risk of patients with causative variants in the NPHS2 gene to develop NS recurrence after kidney transplantation was extremely low. This is coherent with the pathophysiology of intrinsic slit-diaphragm disease. These data are reassuring and should be considered when counselling patients, making living kidney donation, whether related or not, a safe choice.
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The human genome comprises approximately 3% of tandem repeats with variable length (VNTR), a few of which have been linked to human rare diseases. Autosomal dominant tubulointerstitial kidney disease-MUC1 (ADTKD-MUC1) is caused by specific frameshift variants in the coding VNTR of the MUC1 gene. Calling variants from VNTR using short-read sequencing (SRS) is challenging due to poor read mappability. We developed a computational pipeline, VNtyper, for reliable detection of MUC1 VNTR pathogenic variants and demonstrated its clinical utility in two distinct cohorts: (1) a historical cohort including 108 families with ADTKD and (2) a replication naive cohort comprising 2,910 patients previously tested on a panel of genes involved in monogenic renal diseases. In the historical cohort all cases known to carry pathogenic MUC1 variants were re-identified, and a new 25bp-frameshift insertion in an additional mislaid family was detected. In the replication cohort, we discovered and validated 30 new patients.
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X-linked Alport syndrome (XLAS) is an inherited kidney disease caused exclusively by pathogenic variants in the COL4A5 gene. In 10-20% of cases, DNA sequencing of COL4A5 exons or flanking regions cannot identify molecular causes. Here, our objective was to use a transcriptomic approach to identify causative events in a group of 19 patients with XLAS without identified mutation by Alport gene panel sequencing. Bulk RNAseq and/or targeted RNAseq using a capture panel of kidney genes was performed. Alternative splicing events were compared to those of 15 controls by a developed bioinformatic score. When using targeted RNAseq, COL4A5 coverage was found to be 23-fold higher than with bulk RNASeq and revealed 30 significant alternative splicing events in 17 of the 19 patients. After computational scoring, a pathogenic transcript was found in all patients. A causative variant affecting COL4A5 splicing and absent in the general population was identified in all cases. Altogether, we developed a simple and robust method for identification of aberrant transcripts due to pathogenic deep-intronic COL4A5 variants. Thus, these variants, potentially targetable by specific antisense oligonucleotide therapies, were found in a high percentage of patients with XLAS in whom pathogenic variants were missed by conventional DNA sequencing.
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Nefritis Hereditaria , Humanos , Nefritis Hereditaria/diagnóstico , Nefritis Hereditaria/genética , Nefritis Hereditaria/patología , Colágeno Tipo IV/genética , Colágeno Tipo IV/metabolismo , Mutación , Exones , Empalme del ARNRESUMEN
Nephronophthisis (NPH) is an autosomal-recessive ciliopathy representing one of the most frequent causes of kidney failure in childhood characterized by a broad clinical and genetic heterogeneity. Applied to one of the worldwide largest cohorts of patients with NPH, genetic analysis encompassing targeted and whole exome sequencing identified disease-causing variants in 600 patients from 496 families with a detection rate of 71%. Of 788 pathogenic variants, 40 known ciliopathy genes were identified. However, the majority of patients (53%) bore biallelic pathogenic variants in NPHP1. NPH-causing gene alterations affected all ciliary modules defined by structural and/or functional subdomains. Seventy six percent of these patients had progressed to kidney failure, of which 18% had an infantile form (under five years) and harbored variants affecting the Inversin compartment or intraflagellar transport complex A. Forty eight percent of patients showed a juvenile (5-15 years) and 34% a late-onset disease (over 15 years), the latter mostly carrying variants belonging to the Transition Zone module. Furthermore, while more than 85% of patients with an infantile form presented with extra-kidney manifestations, it only concerned half of juvenile and late onset cases. Eye involvement represented a predominant feature, followed by cerebellar hypoplasia and other brain abnormalities, liver and skeletal defects. The phenotypic variability was in a large part associated with mutation types, genes and corresponding ciliary modules with hypomorphic variants in ciliary genes playing a role in early steps of ciliogenesis associated with juvenile-to-late onset NPH forms. Thus, our data confirm a considerable proportion of late-onset NPH suggesting an underdiagnosis in adult chronic kidney disease.
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Ciliopatías , Enfermedades Renales Quísticas , Fallo Renal Crónico , Enfermedades Renales Poliquísticas , Adulto , Humanos , Fallo Renal Crónico/diagnóstico , Enfermedades Renales Poliquísticas/complicaciones , Enfermedades Renales Quísticas/genética , Enfermedades Renales Quísticas/patología , Mutación , Ciliopatías/genéticaRESUMEN
Integrin Subunit Alpha 8 gene (ITGA8) encodes an integrin chain that is known to be critical in the early stage of the kidney development. Bi-allelic pathogenic variants in ITGA8 are associated with bilateral renal agenesis, as well as anomalies involving urogenital system. Here, we report two unrelated patients presenting with slowly progressing chronic kidney disease associated with bilateral renal hypodysplasia carrying homozygous loss of function variants in the ITGA8 gene. These results broaden the clinical and genotypic spectrum of ITGA8 defects, revealing the high and unexpected degree of phenotypic heterogeneity of this autosomal recessive disease. Our study emphasizes the usefulness of Next-Generation Sequencing in unraveling the genetic cause of chronic kidney disease of unknown etiology, and raises the question of genetic modifiers involved in the variation of the phenotypes associated with autosomal recessive ITGA8 pathogenic variants.
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Cadenas alfa de Integrinas , Enfermedades Renales , Humanos , Cadenas alfa de Integrinas/genética , Enfermedades Renales/genéticaRESUMEN
We report the screening of a large panel of genes in a series of 100 fetuses (98 families) affected with severe renal defects. Causative variants were identified in 22% of cases, greatly improving genetic counseling. The percentage of variants explaining the phenotype was different according to the type of phenotype. The highest diagnostic yield was found in cases affected with the ciliopathy-like phenotype (11/15 families and, in addition, a single heterozygous or a homozygous Class 3 variant in PKHD1 in three unrelated cases with autosomal recessive polycystic kidney disease). The lowest diagnostic yield was observed in cases with congenital anomalies of the kidney and urinary tract (9/78 families and, in addition, Class 3 variants in GREB1L in three unrelated cases with bilateral renal agenesis). Inheritance was autosomal recessive in nine genes (PKHD1, NPHP3, CEP290, TMEM67, DNAJB11, FRAS1, ACE, AGT, and AGTR1), and autosomal dominant in six genes (PKD1, PKD2, PAX2, EYA1, BICC1, and MYOCD). Finally, we developed an original approach of next-generation sequencing targeted RNA sequencing using the custom capture panel used for the sequencing of DNA, to validate one MYOCD heterozygous splicing variant identified in two male siblings with megabladder and inherited from their healthy mother.
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Enfermedades Renales , Riñón Poliquístico Autosómico Dominante , Antígenos de Neoplasias , Proteínas de Ciclo Celular/genética , Proteínas del Citoesqueleto/genética , Femenino , Feto/anomalías , Secuenciación de Nucleótidos de Alto Rendimiento , Homocigoto , Humanos , Riñón/anomalías , Enfermedades Renales/congénito , Enfermedades Renales/diagnóstico , Enfermedades Renales/genética , Masculino , Mutación , Riñón Poliquístico Autosómico Dominante/genéticaRESUMEN
BACKGROUND: Co-occurrence of polycystic kidney disease and hyperinsulinemic hypoglycemia has been reported in children in a few families associated with a variant in the promotor of the PMM2 gene, at position -167 upstream of the coding sequence. PMM2 encodes phosphomannomutase 2, a key enzyme in N-glycosylation. While biallelic coding PMM2 mutations are involved in congenital disorder of glycosylation CDG1A, that particular variant in the promoter of the gene, either in the homozygous state or associated with a mutation in the coding exons of the gene, is thought to restrict the N-glycosylation defect to the kidney and the pancreas. METHODS: Targeted exome sequencing of a panel of genes involved in monogenic kidney diseases. RESULTS: We identified a PMM2 variant at position -167 associated with a pathogenic PMM2 variant in the coding exons in 3 families, comprising 6 cases affected with a cystic kidney disease. The spectrum of phenotypes was very broad, from extremely enlarged fetal cystic kidneys in the context of a COACH-like syndrome, to isolated cystic kidney disease with small kidneys, slowly progressing toward kidney failure in adulthood. Hypoglycemia was reported only in one case. CONCLUSION: These data show that the PMM2 promotor variation, in trans of a PMM2 coding mutation, is associated with a wide spectrum of kidney phenotypes, and is not always associated with extra-renal symptoms. When present, extra-renal defects may include COACH-like syndrome. These data prompt screening of PMM2 in unresolved cases of fetal hyperechogenic/cystic kidneys as well as in cystic kidney disease in children and adults. Graphical Abstract.
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Enfermedades Renales Poliquísticas , Hiperinsulinismo Congénito , Humanos , Mutación , Fenotipo , Fosfotransferasas (Fosfomutasas) , Regiones Promotoras Genéticas , SíndromeRESUMEN
DNAJB11 (DnaJ Heat Shock Protein Family (Hsp40) Member B11) heterozygous loss of function variations have been reported in autosomal dominant cystic kidney disease with extensive fibrosis, associated with maturation and trafficking defect involving both the autosomal dominant polycystic kidney disease protein polycystin-1 and the autosomal dominant tubulointerstitial kidney disease protein uromodulin. Here we show that biallelic pathogenic variations in DNAJB11 lead to a severe fetal disease including enlarged cystic kidneys, dilation and proliferation of pancreatic duct cells, and liver ductal plate malformation, an association known as Ivemark II syndrome. Cysts of the kidney were developed exclusively from uromodulin negative tubular segments. In addition, tubular cells from the affected kidneys had elongated primary cilia, a finding previously reported in ciliopathies. Thus, our data show that the recessive disease associated with DNAJB11 variations is a ciliopathy rather than a disease of the autosomal dominant tubulointerstitial kidney disease spectrum, and prompt screening of DNAJB11 in fetal hyperechogenic/cystic kidneys.
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Anomalías Múltiples , Enfermedades Renales Poliquísticas , Riñón Poliquístico Autosómico Dominante , Proteínas del Choque Térmico HSP40 , Humanos , Riñón/anomalías , Riñón/diagnóstico por imagen , Hígado/anomalías , Páncreas/anomalías , Riñón Poliquístico Autosómico Dominante/complicaciones , Riñón Poliquístico Autosómico Dominante/genéticaRESUMEN
BACKGROUND: Bi-allelic loss of function variations in genes encoding proteins of the renin-angiotensin system (AGT, ACE, REN, AGTR1) are associated with autosomal recessive renal tubular dysgenesis, a severe disease characterized by the absence of differentiated proximal tubules leading to fetal anuria and neonatal end-stage renal disease. CASE-DIAGNOSIS/TREATMENT: We identified bi-allelic loss of function mutations in ACE, the gene encoding angiotensin-converting enzyme, in 3 unrelated cases displaying progressive chronic renal failure, whose DNAs had been sent for suspicion of juvenile hyperuricemic nephropathy, nephronophthisis, and cystic renal disease, respectively. In all cases, patients were affected with anemia whose severity was unexpected regarding the level of renal failure and with important polyuro-polydipsia. CONCLUSIONS: Bi-allelic loss of function mutation of ACE can have atypical and sometimes late presentation with chronic renal failure, anemia (out of proportion with the level of renal failure), and polyuro-polydipsia. These data illustrate the usefulness of next generation sequencing and "agnostic" approaches to elucidate cases with chronic kidney disease of unknown etiology and to broaden the spectrum of phenotypes of monogenic renal diseases. It also raises the question of genetic modifiers involved in the variation of the phenotypes associated with these mutations.
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Túbulos Renales Proximales/anomalías , Sistema Renina-Angiotensina/genética , Anomalías Urogenitales/diagnóstico , Adolescente , Preescolar , Femenino , Humanos , Recién Nacido , Masculino , Mutación , Insuficiencia Renal Crónica/diagnóstico , Insuficiencia Renal Crónica/genética , Anomalías Urogenitales/genéticaRESUMEN
BACKGROUND: While typical ultrasound patterns of ciliopathy-related cystic kidney diseases have been described in children, ultrasound findings can overlap between different diseases and atypical patterns exist. In this study, we assessed the presence of the "salt and pepper" pattern in different renal ciliopathies and looked for additional ultrasound features. METHODS: This single-center, retrospective study included all patients with a molecular-proven diagnosis of renal ciliopathy, referred to our center between 2007 and 2017. Images from the first and follow-up ultrasound exams were reviewed. Basic ultrasound features were grouped into patterns and compared to genetic diagnoses. The "salt and pepper" aspect was described as enlarged kidneys with heterogeneous, increased parenchymal echogenicity. RESULTS: A total of 41 children with 5 different renal ciliopathies were included (61% male; median age, 6 years [range, 3 days to 17 years]). The "salt and pepper" pattern was present in 14/15 patients with an autosomal recessive polycystic kidney disease (ARPKD). A similar pattern was found in 1/4 patients with an autosomal dominant polycystic kidney disease and in 1/11 patients with HNF1B mutation. Additional signs found were areas of cortical sparing, comet-tail artifacts, and color comet-tail artifacts. CONCLUSION: Although the "salt and pepper" ultrasound pattern is predominantly found in ARPKD, it may be detected in other ciliopathies. The color comet-tail artifact is an interesting sign when suspecting a renal ciliopathy in case of enlarged hyperechoic kidneys with no detectable microcysts on B-mode grayscale ultrasound.
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Riñón Poliquístico Autosómico Dominante/patología , Adolescente , Niño , Preescolar , Bases de Datos Factuales , Femenino , Humanos , Lactante , Recién Nacido , Masculino , Mutación , Riñón Poliquístico Autosómico Dominante/diagnóstico por imagen , Riñón Poliquístico Autosómico Dominante/genética , Estudios Retrospectivos , Ultrasonografía Doppler en ColorRESUMEN
BACKGROUNDProteinuria is considered an unfavorable clinical condition that accelerates renal and cardiovascular disease. However, it is not clear whether all forms of proteinuria are damaging. Mutations in CUBN cause Imerslund-Gräsbeck syndrome (IGS), which is characterized by intestinal malabsorption of vitamin B12 and in some cases proteinuria. CUBN encodes for cubilin, an intestinal and proximal tubular uptake receptor containing 27 CUB domains for ligand binding.METHODSWe used next-generation sequencing for renal disease genes to genotype cohorts of patients with suspected hereditary renal disease and chronic proteinuria. CUBN variants were analyzed using bioinformatics, structural modeling, and epidemiological methods.RESULTSWe identified 39 patients, in whom biallelic pathogenic variants in the CUBN gene were associated with chronic isolated proteinuria and early childhood onset. Since the proteinuria in these patients had a high proportion of albuminuria, glomerular diseases such as steroid-resistant nephrotic syndrome or Alport syndrome were often the primary clinical diagnosis, motivating renal biopsies and the use of proteinuria-lowering treatments. However, renal function was normal in all cases. By contrast, we did not found any biallelic CUBN variants in proteinuric patients with reduced renal function or focal segmental glomerulosclerosis. Unlike the more N-terminal IGS mutations, 37 of the 41 proteinuria-associated CUBN variants led to modifications or truncations after the vitamin B12-binding domain. Finally, we show that 4 C-terminal CUBN variants are associated with albuminuria and slightly increased GFR in meta-analyses of large population-based cohorts.CONCLUSIONCollectively, our data suggest an important role for the C-terminal half of cubilin in renal albumin reabsorption. Albuminuria due to reduced cubilin function could be an unexpectedly common benign condition in humans that may not require any proteinuria-lowering treatment or renal biopsy.FUNDINGATIP-Avenir program, Fondation Bettencourt-Schueller (Liliane Bettencourt Chair of Developmental Biology), Agence Nationale de la Recherche (ANR) Investissements d'avenir program (ANR-10-IAHU-01) and NEPHROFLY (ANR-14-ACHN-0013, to MS), Steno Collaborative Grant 2018 (NNF18OC0052457, to TSA and MS), Heisenberg Professorship of the German Research Foundation (KO 3598/5-1, to AK), Deutsche Forschungsgemeinschaft (DFG) Collaborative Research Centre (SFB) KIDGEM 1140 (project 246781735, to CB), and Federal Ministry of Education and Research (BMB) (01GM1515C, to CB).
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Albuminuria , Anemia Megaloblástica , Túbulos Renales Proximales , Síndromes de Malabsorción , Mutación , Proteinuria , Receptores de Superficie Celular , Deficiencia de Vitamina B 12 , Albuminuria/epidemiología , Albuminuria/genética , Albuminuria/metabolismo , Albuminuria/patología , Anemia Megaloblástica/epidemiología , Anemia Megaloblástica/genética , Anemia Megaloblástica/metabolismo , Anemia Megaloblástica/patología , Femenino , Humanos , Túbulos Renales Proximales/metabolismo , Túbulos Renales Proximales/patología , Síndromes de Malabsorción/epidemiología , Síndromes de Malabsorción/genética , Síndromes de Malabsorción/metabolismo , Síndromes de Malabsorción/patología , Masculino , Proteinuria/epidemiología , Proteinuria/genética , Proteinuria/metabolismo , Proteinuria/patología , Receptores de Superficie Celular/genética , Receptores de Superficie Celular/metabolismo , Deficiencia de Vitamina B 12/epidemiología , Deficiencia de Vitamina B 12/genética , Deficiencia de Vitamina B 12/metabolismo , Deficiencia de Vitamina B 12/patologíaRESUMEN
The prevalence of neurological involvement in patients with a deletion of or a variant in the HNF1B gene remains discussed. The aim of this study was to investigate the neuropsychological outcomes in a large cohort of children carrying either a HNF1B whole-gene deletion or a disease-associated variant, revealed by the presence of kidney anomalies. The neuropsychological development-based on school level-of 223 children included in this prospective cohort was studied. Data from 180 children were available for analysis. Patients mean age was 9.6 years, with 39.9% of girls. Among these patients, 119 carried a HNF1B deletion and 61 a disease-associated variant. In the school-aged population, 12.7 and 3.6% of patients carrying a HNF1B deletion and a disease-associated variant had special educational needs, respectively. Therefore, the presence of a HNF1B deletion increases the risk to present with a neuropsychiatric involvement when compared with the general population. On the other hand, almost 90% of patients carrying a HNF1B disease-associated variant or deletion have a normal schooling in a general educational environment. Even if these findings do not predict the risk of neuropsychiatric disease at adulthood, most patients diagnosed secondary to kidney anomalies do not show a neurological outcome severe enough to impede standard schooling at elementary school. These results should be taken into account in prenatal counseling.
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Rendimiento Académico/estadística & datos numéricos , Factor Nuclear 1-beta del Hepatocito/genética , Trastornos del Neurodesarrollo/genética , Adolescente , Niño , Femenino , Eliminación de Gen , Humanos , Riñón/anomalías , Masculino , Trastornos del Neurodesarrollo/epidemiología , SíndromeRESUMEN
BACKGROUND: Recommendations for management of Finnish-type congenital nephrotic syndrome (CNS) followed by many teams include daily albumin infusions, early bilateral nephrectomy, dialysis and transplantation. We aimed to assess the treatment and outcome of patients with CNS in France. METHODS: We conducted a nationwide retrospective study on 55 consecutive children born between 2000 and 2014 treated for non-infectious CNS. RESULTS: The estimated cumulative incidence of CNS was 0.5/100 000 live births. The underlying defect was biallelic mutations in NPHS1 (36/55, 65%), NPHS2 (5/55, 7%), PLCE1 (1/55, 2%), heterozygous mutation in WT1 (4/55, 7%) and not identified in nine children (16%). Fifty-three patients (96%) received daily albumin infusions from diagnosis (median age 14 days), which were spaced and withdrawn in 10 patients. Twenty children (35%) were managed as outpatients. Thirty-nine patients reached end-stage kidney disease (ESKD) at a median age of 11 months. The overall renal survival was 64% and 45% at 1 and 2 years of age, respectively. Thirteen children died during the study period including four at diagnosis, two of nosocomial catheter-related septic shock, six on dialysis and one after transplantation. The remaining 13 patients were alive with normal renal function at last follow-up [median 32 months (range 9-52)]. Renal and patient survivals were longer in patients with NPHS1 mutations than in other patients. The invasive infection rate was 2.41/patient/year. CONCLUSIONS: Our study shows: (i) a survival free from ESKD in two-thirds of patients at 1 year and in one-half at 2 years and (ii) a significant reduction or even a discontinuation of albumin infusions allowing ambulatory care in a subset of patients. These results highlight the need for new therapeutic guidelines for CNS patients.
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Proteínas de la Membrana/genética , Mutación , Nefrectomía/mortalidad , Síndrome Nefrótico/mortalidad , Progresión de la Enfermedad , Femenino , Francia/epidemiología , Humanos , Incidencia , Lactante , Recién Nacido , Masculino , Síndrome Nefrótico/epidemiología , Síndrome Nefrótico/genética , Síndrome Nefrótico/terapia , Estudios Retrospectivos , Tasa de Supervivencia , Resultado del TratamientoRESUMEN
Renal tubular dysgenesis (RTD) is a developmental abnormality of the nephron characterized by fetal anuria, oligohydramnios, and severe postnatal hypotension. Genetic forms have an autosomal recessive inheritance and are caused by mutations in genes encoding key components of the renin-angiotensin pathway. We report three patients from two unrelated families with RTD due to pathogenic variants of the angiotensin-converting enzyme (ACE) gene, in whom RTD was associated with microcolon. We also detail key variations of the renin-angiotensin system in one of these infants. The severe intestinal developmental abnormality culminating in microcolon and early terminal ileum perforation/necrotizing enterocolitis is a novel finding not previously associated with RTD, which points to a role of the renin-angiotensin system in gut development.
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Anomalías Múltiples/genética , Íleon/anomalías , Túbulos Renales/anomalías , Peptidil-Dipeptidasa A/genética , Fenotipo , Anomalías Múltiples/patología , Femenino , Humanos , Recién Nacido , MasculinoRESUMEN
Congenital anomalies of the kidney and urinary tract (CAKUT) occur in three to six of 1000 live births, represent about 20% of the prenatally detected anomalies, and constitute the main cause of CKD in children. These disorders are phenotypically and genetically heterogeneous. Monogenic causes of CAKUT in humans and mice have been identified. However, despite high-throughput sequencing studies, the cause of the disease remains unknown in most patients, and several studies support more complex inheritance and the role of environmental factors and/or epigenetics in the pathophysiology of CAKUT. Here, we report the targeted exome sequencing of 330 genes, including genes known to be involved in CAKUT and candidate genes, in a cohort of 204 unrelated patients with CAKUT; 45% of the patients were severe fetal cases. We identified pathogenic mutations in 36 of 204 (17.6%) patients. These mutations included five de novo heterozygous loss of function mutations/deletions in the PBX homeobox 1 gene (PBX1), a gene known to have a crucial role in kidney development. In contrast, the frequency of SOX17 and DSTYK variants recently reported as pathogenic in CAKUT did not indicate causality. These findings suggest that PBX1 is involved in monogenic CAKUT in humans and call into question the role of some gene variants recently reported as pathogenic in CAKUT. Targeted exome sequencing also proved to be an efficient and cost-effective strategy to identify pathogenic mutations and deletions in known CAKUT genes.
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Proteínas de Unión al ADN/genética , Proteínas Proto-Oncogénicas/genética , Anomalías Urogenitales/genética , Estudios de Cohortes , Análisis Mutacional de ADN , Exoma , Femenino , Humanos , Masculino , Factor de Transcripción 1 de la Leucemia de Células Pre-BRESUMEN
BACKGROUND: Nephronophthisis, an autosomal recessive nephropathy, is responsible for 10% of childhood chronic renal failure. The deletion of its major gene, NPHP1, with a minor allele frequency of 0.24% in the general population, is the most common mutation leading to a monogenic form of childhood chronic renal failure. It is challenging to detect it in the heterozygous state. We aimed to evaluate the sensitivity and the specificity of the quantitative multiplex PCR of short fluorescent fragments (QMPSF) in its detection. METHODS: After setting up the protocol of QMPSF, we validated it on 39 individuals diagnosed by multiplex ligation-dependent probe amplification (MLPA) with normal NPHP1 copy number (n=17), with heterozygous deletion (n=13, seven parents and six patients), or with homozygous deletion (n=9). To assess the rate of the deletions that arise from independent events, deleted alleles were haplotyped. RESULTS: The results of QMPSF and MLPA correlated perfectly in the identification of 76 heterozygously deleted and 56 homozygously deleted exons. The inter-experimental variability of the dosage quotient obtained by QMPSF was low: control, 1.05 (median; range, 0.86-1.33, n = 102 exons); heterozygous deletion, 0.51 (0.42-0.67, n = 76 exons); homozygous deletion, 0 (0-0, n = 56 exons). All patients harboring a heterozygous deletion were found to carry a hemizygous mutation. At least 15 out of 18 deletions appeared on different haplotypes and one deletion appeared de novo. CONCLUSIONS: The cost- and time-effective QMPSF has a 100% sensitivity and specificity in the detection of NPHP1 deletion. The potential de novo appearance of NPHP1 deletions makes its segregation analysis highly recommended in clinical practice.
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Proteínas Adaptadoras Transductoras de Señales/genética , Eliminación de Gen , Heterocigoto , Proteínas de la Membrana/genética , Reacción en Cadena de la Polimerasa/métodos , Secuencia de Bases , Proteínas del Citoesqueleto , Exones/genética , Humanos , Enfermedades Renales Quísticas/congénito , Enfermedades Renales Quísticas/genética , Límite de DetecciónRESUMEN
Prenatal forms of autosomal dominant polycystic kidney disease (ADPKD) are rare but can be recurrent in some families, suggesting a common genetic modifying background. Few patients have been reported carrying, in addition to the familial mutation, variation(s) in polycystic kidney disease 1 (PKD1) or HNF1 homeobox B (HNF1B), inherited from the unaffected parent, or biallelic polycystic kidney and hepatic disease 1 (PKHD1) mutations. To assess the frequency of additional variations in PKD1, PKD2, HNF1B, and PKHD1 associated with the familial PKD mutation in early ADPKD, these four genes were screened in 42 patients with early ADPKD in 41 families. Two patients were associated with de novo PKD1 mutations. Forty patients occurred in 39 families with known ADPKD and were associated with PKD1 mutation in 36 families and with PKD2 mutation in two families (no mutation identified in one family). Additional PKD variation(s) (inherited from the unaffected parent when tested) were identified in 15 of 42 patients (37.2%), whereas these variations were observed in 25 of 174 (14.4%, P=0.001) patients with adult ADPKD. No HNF1B variations or PKHD1 biallelic mutations were identified. These results suggest that, at least in some patients, the severity of the cystic disease is inversely correlated with the level of polycystin 1 function.
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Riñón Poliquístico Autosómico Dominante/genética , Canales Catiónicos TRPP/genética , Adolescente , Adulto , Anciano , Niño , Preescolar , Análisis Mutacional de ADN , Padre , Femenino , Factor Nuclear 1-beta del Hepatocito/genética , Humanos , Lactante , Fallo Renal Crónico/etiología , Masculino , Persona de Mediana Edad , Madres , Mutación , Linaje , Riñón Poliquístico Autosómico Dominante/complicaciones , Riñón Poliquístico Autosómico Dominante/diagnóstico por imagen , Receptores de Superficie Celular/genética , Estudios Retrospectivos , Ultrasonografía Prenatal , Adulto JovenRESUMEN
Alport syndrome is an inherited nephropathy associated with mutations in genes encoding type IV collagen chains present in the glomerular basement membrane. COL4A5 mutations are associated with the major X-linked form of the disease, and COL4A3 and COL4A4 mutations are associated with autosomal recessive and dominant forms (thought to be involved in 15% and 1%-5% of the families, respectively) and benign familial hematuria. Mutation screening of these three large genes is time-consuming and expensive. Here, we carried out a combination of multiplex PCR, amplicon quantification, and next generation sequencing (NGS) analysis of three genes in 101 unrelated patients. We identified 88 mutations and 6 variations of unknown significance on 116 alleles in 83 patients. Two additional indel mutations were found only by secondary Sanger sequencing, but they were easily identified retrospectively with the web-based sequence visualization tool Integrative Genomics Viewer. Altogether, 75 mutations were novel. Sequencing the three genes simultaneously was particularly advantageous as the mode of inheritance could not be determined with certainty in many instances. The proportion of mutations in COL4A3 and COL4A4 was notably high, and the autosomal dominant forms of Alport syndrome appear more frequently than reported previously. Finally, this approach allowed the identification of large COL4A3 and COL4A4 rearrangements not described previously. We conclude that NGS is efficient, reduces screening time and cost, and facilitates the provision of appropriate genetic counseling in Alport syndrome.