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ImportanceNewborn genome sequencing (NBSeq) can detect infants at risk for treatable disorders currently undetected by conventional newborn screening. Despite broad stakeholder support for NBSeq, the perspectives of rare disease experts regarding which diseases should be screened have not been ascertained.ObjectiveTo query rare disease experts about their perspectives on NBSeq and which gene-disease pairs they consider appropriate to evaluate in apparently healthy newborns.Design, Setting, and ParticipantsThis survey study, designed between November 2, 2021, and February 11, 2022, assessed experts’ perspectives on 6 statements related to NBSeq. Experts were also asked to indicate whether they would recommend including each of 649 gene-disease pairs associated with potentially treatable conditions in NBSeq. The survey was administered between February 11 and September 23, 2022, to 386 experts, including all 144 directors of accredited medical and laboratory genetics training programs in the US.ExposuresExpert perspectives on newborn screening using genome sequencing.Main Outcomes and MeasuresThe proportion of experts indicating agreement or disagreement with each survey statement and those who selected inclusion of each gene-disease pair were tabulated. Exploratory analyses of responses by gender and age were conducted using t and χ2 tests.ResultsOf 386 experts invited, 238 (61.7%) responded (mean [SD] age, 52.6 [12.8] years [range 27-93 years]; 126 [52.9%] women and 112 [47.1%] men). Among the experts who responded, 161 (87.9%) agreed that NBSeq for monogenic treatable disorders should be made available to all newborns; 107 (58.5%) agreed that NBSeq should include genes associated with treatable disorders, even if those conditions were low penetrance; 68 (37.2%) agreed that actionable adult-onset conditions should be sequenced in newborns to facilitate cascade testing in parents, and 51 (27.9%) agreed that NBSeq should include screening for conditions with no established therapies or management guidelines. The following 25 genes were recommended by 85% or more of the experts: OTC, G6PC, SLC37A4, CYP11B1, ARSB, F8, F9, SLC2A1, CYP17A1, RB1, IDS, GUSB, DMD, GLUD1, CYP11A1, GALNS, CPS1, PLPBP, ALDH7A1, SLC26A3, SLC25A15, SMPD1, GATM, SLC7A7, and NAGS. Including these, 42 gene-disease pairs were endorsed by at least 80% of experts, and 432 genes were endorsed by at least 50% of experts.Conclusions and RelevanceIn this survey study, rare disease experts broadly supported NBSeq for treatable conditions and demonstrated substantial concordance regarding the inclusion of a specific subset of genes in NBSeq.

Background Galloway-Mowat syndrome (GAMOS) is characterized by neurodevelopmental defects and a progressive nephropathy, which typically manifests as steroid-resistant nephrotic syndrome. The prognosis of GAMOS is poor, and the majority of children progress to renal failure. The discovery of monogenic causes of GAMOS has uncovered molecular pathways involved in the pathogenesis of disease. Methods Homozygosity mapping, whole-exome sequencing, and linkage analysis were used to identify mutations in four families with a GAMOS-like phenotype, and high-throughput PCR technology was applied to 91 individuals with GAMOS and 816 individuals with isolated nephrotic syndrome. In vitro and in vivo studies determined the functional significance of the mutations identified. Results Three biallelic variants of the transcriptional regulator PRDM15 were detected in six families with proteinuric kidney disease. Four families with a variant in the protein's zinc-finger (ZNF) domain have additional GAMOS-like features, including brain anomalies, cardiac defects, and skeletal defects. All variants destabilize the PRDM15 protein, and the ZNF variant additionally interferes with transcriptional activation. Morpholino oligonucleotide-mediated knockdown of Prdm15 in Xenopus embryos disrupted pronephric development. Human wild-type PRDM15 RNA rescued the disruption, but the three PRDM15 variants did not. Finally, CRISPR-mediated knockout of PRDM15 in human podocytes led to dysregulation of several renal developmental genes. Conclusions Variants in PRDM15 can cause either isolated nephrotic syndrome or a GAMOS-type syndrome on an allelic basis. PRDM15 regulates multiple developmental kidney genes, and is likely to play an essential role in renal development in humans.

Introduction Steroid-resistant nephrotic syndrome (SRNS) is the second most common cause of chronic kidney disease during childhood. Identification of 63 monogenic human genes has delineated 12 distinct pathogenic pathways. Methods Here, we generated 2 independent sets of nephrotic syndrome (NS) candidate genes to augment the discovery of additional monogenic causes based on whole-exome sequencing (WES) data from 1382 families with NS. Results We first identified 63 known monogenic causes of NS in mice from public databases and scientific publications, and 12 of these genes overlapped with the 63 known human monogenic SRNS genes. Second, we used a set of 64 genes that are regulated by the transcription factor Wilms tumor 1 (WT1), which causes SRNS if mutated. Thirteen of these WT1-regulated genes overlapped with human or murine NS genes. Finally, we overlapped these lists of murine and WT1 candidate genes with our list of 120 candidate genes generated from WES in 1382 NS families, to identify novel candidate genes for monogenic human SRNS. Using this approach, we identified 7 overlapping genes, of which 3 genes were shared by all datasets, including SYNPO. We show that loss-of-function of SYNPO leads to decreased CDC42 activity and reduced podocyte migration rate, both of which are rescued by overexpression of wild-type complementary DNA (cDNA), but not by cDNA representing the patient mutation. Conclusion Thus, we identified 3 novel candidate genes for human SRNS using 3 independent, nonoverlapping hypotheses, and generated functional evidence for SYNPO as a novel potential monogenic cause of NS., Graphical abstract

Pathogenic variants in the LRP2 gene, encoding the multiligand receptor megalin, cause a rare autosomal recessive syndrome: Donnai-Barrow/Facio-Oculo-Acoustico-Renal (DB/FOAR) syndrome. Because of the rarity of the syndrome, the long-term consequences of the tubulopathy on human renal health have been difficult to ascertain, and the human clinical condition has hitherto been characterized as a benign tubular condition with asymptomatic low-molecular-weight proteinuria. We investigated renal function and morphology in a murine model of DB/FOAR syndrome and in patients with DB/FOAR. We analyzed glomerular filtration rate in mice by FITC-inulin clearance and clinically characterized six families, including nine patients with DB/FOAR and nine family members. Urine samples from patients were analyzed by Western blot analysis and biopsy materials were analyzed by histology. In the mouse model, we used histological methods to assess nephrogenesis and postnatal renal structure and contrast-enhanced magnetic resonance imaging to assess glomerular number. In megalin-deficient mice, we found a lower glomerular filtration rate and an increase in the abundance of injury markers, such as kidney injury molecule-1 and N-acetyl-β-d-glucosaminidase. Renal injury was validated in patients, who presented with increased urinary kidney injury molecule-1, classical markers of chronic kidney disease, and glomerular proteinuria early in life. Megalin-deficient mice had normal nephrogenesis, but they had 19% fewer nephrons in early adulthood and an increased fraction of nephrons with disconnected glomerulotubular junction. In conclusion, megalin dysfunction, as present in DB/FOAR syndrome, confers an increased risk of progression into chronic kidney disease.

Pathogenic variants in the LRP2 gene, encoding the multiligand receptor megalin, cause a rare autosomal recessive syndrome: Donnai-Barrow/Facio-Oculo-Acoustico-Renal (DB/FOAR) syndrome. Because of the rarity of the syndrome, the long-term consequences of the tubulopathy on human renal health have been difficult to ascertain, and the human clinical condition has hitherto been characterized as a benign tubular condition with asymptomatic low-molecular-weight proteinuria. We investigated renal function and morphology in a murine model of DB/FOAR syndrome and in patients with DB/FOAR. We analyzed glomerular filtration rate in mice by FITC-inulin clearance and clinically characterized six families, including nine patients with DB/FOAR and nine family members. Urine samples from patients were analyzed by Western blot analysis and biopsy materials were analyzed by histology. In the mouse model, we used histological methods to assess nephrogenesis and postnatal renal structure and contrast-enhanced magnetic resonance imaging to assess glomerular number. In megalin-deficient mice, we found a lower glomerular filtration rate and an increase in the abundance of injury markers, such as kidney injury molecule-1 and N-acetyl-β-d-glucosaminidase. Renal injury was validated in patients, who presented with increased urinary kidney injury molecule-1, classical markers of chronic kidney disease, and glomerular proteinuria early in life. Megalin-deficient mice had normal nephrogenesis, but they had 19% fewer nephrons in early adulthood and an increased fraction of nephrons with disconnected glomerulotubular junction. In conclusion, megalin dysfunction, as present in DB/FOAR syndrome, confers an increased risk of progression into chronic kidney disease.

Steroid-resistant nephrotic syndrome (SRNS) is a genetically heterogeneous kidney disease that is the second most frequent cause of kidney failure in the first 2 decades of life. Despite the identification of mutations in more than 39 genes as causing SRNS, and the localization of its pathogenesis to glomerular podocytes, the disease mechanisms of SRNS remain poorly understood and no universally safe and effective therapy exists to treat patients with this condition. Recently, genetic research has identified a subgroup of SRNS patients whose kidney pathology is caused by primary coenzyme Q10 (CoQ10) deficiency due to recessive mutations in genes that encode proteins in the CoQ10 biosynthesis pathway. Clinical and preclinical studies show that primary CoQ10 deficiency may be responsive to treatment with CoQ10 supplements bypassing the biosynthesis defects. Coenzyme Q10 is an essential component of the mitochondrial respiratory chain, where it transports electrons from complexes I and II to complex III. Studies in yeast and mammalian model systems have recently identified the molecular functions of the individual CoQ10 biosynthesis complex proteins, validated these findings, and provided an impetus for developing therapeutic compounds to replenish CoQ10 levels in the tissues/organs and thus prevent the destruction of tissues due to mitochondrial OXPHOS deficiencies. In this review, we will summarize the clinical findings of the kidney pathophysiology of primary CoQ10 deficiencies and discuss recent advances in the development of therapies to counter CoQ10 deficiency in tissues.

BACKGROUND: Galloway-Mowat syndrome (GAMOS) is characterized by neurodevelopmental defects and a progressive nephropathy, which typically manifests as steroid-resistant nephrotic syndrome. The prognosis of GAMOS is poor, and the majority of children progress to renal failure. The discovery of monogenic causes of GAMOS has uncovered molecular pathways involved in the pathogenesis of disease. METHODS: Homozygosity mapping, whole-exome sequencing, and linkage analysis were used to identify mutations in four families with a GAMOS-like phenotype, and high-throughput PCR technology was applied to 91 individuals with GAMOS and 816 individuals with isolated nephrotic syndrome. In vitro and in vivo studies determined the functional significance of the mutations identified. RESULTS: Three biallelic variants of the transcriptional regulator PRDM15 were detected in six families with proteinuric kidney disease. Four families with a variant in the protein’s zinc-finger (ZNF) domain have additional GAMOS-like features, including brain anomalies, cardiac defects, and skeletal defects. All variants destabilize the PRDM15 protein, and the ZNF variant additionally interferes with transcriptional activation. Morpholino oligonucleotide-mediated knockdown of Prdm15 in Xenopus embryos disrupted pronephric development. Human wild-type PRDM15 RNA rescued the disruption, but the three PRDM15 variants did not. Finally, CRISPR-mediated knockout of PRDM15 in human podocytes led to dysregulation of several renal developmental genes. CONCLUSIONS: Variants in PRDM15 can cause either isolated nephrotic syndrome or a GAMOS-type syndrome on an allelic basis. PRDM15 regulates multiple developmental kidney genes, and is likely to play an essential role in renal development in humans.

Item does not contain fulltext Steroid-resistant nephrotic syndrome (SRNS) almost invariably progresses to end-stage renal disease. Although more than 50 monogenic causes of SRNS have been described, a large proportion of SRNS remains unexplained. Recently, it was discovered that mutations of NUP93 and NUP205, encoding 2 proteins of the inner ring subunit of the nuclear pore complex (NPC), cause SRNS. Here, we describe mutations in genes encoding 4 components of the outer rings of the NPC, namely NUP107, NUP85, NUP133, and NUP160, in 13 families with SRNS. Using coimmunoprecipitation experiments, we showed that certain pathogenic alleles weakened the interaction between neighboring NPC subunits. We demonstrated that morpholino knockdown of nup107, nup85, or nup133 in Xenopus disrupted glomerulogenesis. Re-expression of WT mRNA, but not of mRNA reflecting mutations from SRNS patients, mitigated this phenotype. We furthermore found that CRISPR/Cas9 knockout of NUP107, NUP85, or NUP133 in podocytes activated Cdc42, an important effector of SRNS pathogenesis. CRISPR/Cas9 knockout of nup107 or nup85 in zebrafish caused developmental anomalies and early lethality. In contrast, an in-frame mutation of nup107 did not affect survival, thus mimicking the allelic effects seen in humans. In conclusion, we discovered here that mutations in 4 genes encoding components of the outer ring subunits of the NPC cause SRNS and thereby provide further evidence that specific hypomorphic mutations in these essential genes cause a distinct, organ-specific phenotype.

Galloway-Mowat syndrome (GAMOS) is a phenotypically heterogeneous disorder characterized by neurodevelopmental defects combined with renal-glomerular disease, manifesting with proteinuria. To identify additional monogenic disease causes, we here performed whole exome sequencing (WES), linkage analysis, and homozygosity mapping in three affected siblings of an Indian family with GAMOS. Applying established criteria for variant filtering, we identify a novel homozygous splice site mutation in the gene WDR4 as the likely disease-causing mutation in this family. In line with previous reports, we observe growth deficiency, microcephaly, developmental delay, and intellectual disability as phenotypic features resulting from WDR4 mutations. However, the newly identified allele additionally gives rise to proteinuria and nephrotic syndrome, a phenotype that was never reported in patients with WDR4 mutations. Our data thus expand the phenotypic spectrum of WDR4 mutations by demonstrating that, depending on the specific mutated allele, a renal phenotype may be present. This finding suggests that GAMOS may occupy a phenotypic spectrum with other microcephalic diseases. Furthermore, WDR4 is an additional example of a gene that encodes a tRNA modifying enzyme and gives rise to GAMOS, if mutated. Our findings thereby support the recent observation that, like neurons, podocytes of the renal glomerulus are particularly vulnerable to cellular defects resulting from altered tRNA modifications.

Midaortic syndrome (MAS) is a rare cause of severe childhood hypertension characterized by narrowing of the abdominal aorta in children and is associated with extensive vascular disease. It may occur as part of a genetic syndrome, such as neurofibromatosis, or as consequence of a pathological inflammatory disease. However, most cases are considered idiopathic. We hypothesized that in a high percentage of these patients, a monogenic cause of disease may be detected by evaluating whole exome sequencing data for mutations in 1 of 38 candidate genes previously described to cause vasculopathy. We studied a cohort of 36 individuals from 35 different families with MAS by exome sequencing. In 15 of 35 families (42.9%), we detected likely causal dominant mutations. In 15 of 35 (42.9%) families with MAS, whole exome sequencing revealed a mutation in one of the genes previously associated with vascular disease ( NF1 , JAG1 , ELN , GATA6 , and RNF213 ). Ten of the 15 mutations have not previously been reported. This is the first report of ELN , RNF213 , or GATA6 mutations in individuals with MAS. Mutations were detected in NF1 (6/15 families), JAG1 (4/15 families), ELN (3/15 families), and one family each for GATA6 and RNF213 . Eight individuals had syndromic disease and 7 individuals had isolated MAS. Whole exome sequencing can provide conclusive molecular genetic diagnosis in a high fraction of individuals with syndromic or isolated MAS. Establishing an etiologic diagnosis may reveal genotype/phenotype correlations for MAS in the future and should, therefore, be performed routinely in MAS.

Background Alport syndrome (AS) and atypical hemolytic-uremic syndrome (aHUS) are rare forms of chronic kidney disease (CKD) that can lead to a severe decline of renal function. Steroid-resistant nephrotic syndrome (SRNS) is more common than AS and aHUS and causes 10% of childhood-onset CKD. In recent years, multiple monogenic causes of AS, aHUS and SRNS have been identified, but their relative prevalence has yet to be studied together in a typical pediatric cohort of children with proteinuria and hematuria. We hypothesized that identification of causative mutations by whole exome sequencing (WES) in known monogenic nephritis and nephrosis genes would allow distinguishing nephritis from nephrosis in a typical pediatric group of patients with both proteinuria and hematuria at any level. Methods We therefore conducted an exon sequencing (WES) analysis for 11 AS, aHUS and thrombotic thrombocytopenic purpura-causing genes in an international cohort of 371 patients from 362 families presenting with both proteinuria and hematuria before age 25 years. In parallel, we conducted either WES or high-throughput exon sequencing for 23 SRNS-causing genes in all patients. Results We detected pathogenic mutations in 18 of the 34 genes analyzed, leading to a molecular diagnosis in 14.1% of families (51 of 362). Disease-causing mutations were detected in 3 AS-causing genes (4.7%), 3 aHUS-causing genes (1.4%) and 12 NS-causing genes (8.0%). We observed a much higher mutation detection rate for monogenic forms of CKD in consanguineous families (35.7% versus 10.1%). Conclusions We present the first estimate of relative frequency of inherited AS, aHUS and NS in a typical pediatric cohort with proteinuria and hematuria. Important therapeutic and preventative measures may result from mutational analysis in individuals with proteinuria and hematuria.

Item does not contain fulltext BACKGROUND: Nephrotic syndrome (NS), a chronic kidney disease, is characterized by significant loss of protein in the urine causing hypoalbuminemia and edema. In general, approximately 15% of childhood-onset cases do not respond to steroid therapy and are classified as steroid-resistant NS (SRNS). In approximately 30% of cases with SRNS, a causative mutation can be detected in one of 44 monogenic SRNS genes. The gene LAMA5 encodes laminin-alpha5, an essential component of the glomerular basement membrane. Mice with a hypomorphic mutation in the orthologous gene Lama5 develop proteinuria and hematuria. METHODS: To identify additional monogenic causes of NS, we performed whole exome sequencing in 300 families with pediatric NS. In consanguineous families we applied homozygosity mapping to identify genomic candidate loci for the underlying recessive mutation. RESULTS: In three families, in whom mutations in known NS genes were excluded, but in whom a recessive, monogenic cause of NS was strongly suspected based on pedigree information, we identified homozygous variants of unknown significance (VUS) in the gene LAMA5. While all affected individuals had nonsyndromic NS with an early onset of disease, their clinical outcome and response to immunosuppressive therapy differed notably. CONCLUSION: We here identify recessive VUS in the gene LAMA5 in patients with partially treatment-responsive NS. More data will be needed to determine the impact of these VUS in disease management. However, familial occurrence of disease, data from genetic mapping and a mouse model that recapitulates the NS phenotypes suggest that these genetic variants may be inherited factors that contribute to the development of NS in pediatric patients.

The incidence of nephrolithiasis continues to rise. Previously, we showed that a monogenic cause could be detected in 11.4% of individuals with adult-onset nephrolithiasis or nephrocalcinosis and in 16.7-20.8% of individuals with onset before 18 years of age, using gene panel sequencing of 30 genes known to cause nephrolithiasis/nephrocalcinosis. To overcome the limitations of panel sequencing, we utilized whole exome sequencing in 51 families, who presented before age 25 years with at least one renal stone or with a renal ultrasound finding of nephrocalcinosis to identify the underlying molecular genetic cause of disease. In 15 of 51 families, we detected a monogenic causative mutation by whole exome sequencing. A mutation in seven recessive genes ( AGXT, ATP6V1B1, CLDN16, CLDN19, GRHPR, SLC3A1, SLC12A1 ), in one dominant gene ( SLC9A3R1 ), and in one gene ( SLC34A1 ) with both recessive and dominant inheritance was detected. Seven of the 19 different mutations were not previously described as disease-causing. In one family, a causative mutation in one of 117 genes that may represent phenocopies of nephrolithiasis-causing genes was detected. In nine of 15 families, the genetic diagnosis may have specific implications for stone management and prevention. Several factors that correlated with the higher detection rate in our cohort were younger age at onset of nephrolithiasis/nephrocalcinosis, presence of multiple affected members in a family, and presence of consanguinity. Thus, we established whole exome sequencing as an efficient approach toward a molecular genetic diagnosis in individuals with nephrolithiasis/nephrocalcinosis who manifest before age 25 years.

Item does not contain fulltext BACKGROUND: Congenital anomalies of the kidney and urinary tract (CAKUT) are the most prevalent cause of kidney disease in the first three decades of life. Previous gene panel studies showed monogenic causation in up to 12% of patients with CAKUT. METHODS: We applied whole-exome sequencing to analyze the genotypes of individuals from 232 families with CAKUT, evaluating for mutations in single genes known to cause human CAKUT and genes known to cause CAKUT in mice. In consanguineous or multiplex families, we additionally performed a search for novel monogenic causes of CAKUT. RESULTS: In 29 families (13%), we detected a causative mutation in a known gene for isolated or syndromic CAKUT that sufficiently explained the patient's CAKUT phenotype. In three families (1%), we detected a mutation in a gene reported to cause a phenocopy of CAKUT. In 15 of 155 families with isolated CAKUT, we detected deleterious mutations in syndromic CAKUT genes. Our additional search for novel monogenic causes of CAKUT in consanguineous and multiplex families revealed a potential single, novel monogenic CAKUT gene in 19 of 232 families (8%). CONCLUSIONS: We identified monogenic mutations in a known human CAKUT gene or CAKUT phenocopy gene as the cause of disease in 14% of the CAKUT families in this study. Whole-exome sequencing provides an etiologic diagnosis in a high fraction of patients with CAKUT and will provide a new basis for the mechanistic understanding of CAKUT. 01 september 2018

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 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.

Steroid-resistant nephrotic syndrome (SRNS) is the second most frequent cause of end-stage renal disease (ESRD) among patients manifesting at under 25 years of age. We performed mutation analysis using a high-throughput PCR-based microfluidic technology in 24 single-gene causes of SRNS in a cohort of 72 families, who presented with SRNS before the age of 25 years. Within an 18-month interval, we obtained DNA samples, pedigree information, and clinical information from 77 consecutive children with SRNS from 72 different families seen at Boston Children’s Hospital (BCH). Mutation analysis was completed by combining high-throughput multiplex PCR with next-generation sequencing. We analyzed the sequences of 18 recessive and 6 dominant genes of SRNS in all 72 families for disease-causing variants. We identified the disease-causing mutation in 8 out of 72 (11.1%) families. Mutations were detected in the six genes: NPHS1 (2 out of 72), WT1 (2 out of 72), NPHS2, MYO1E, TRPC6, and INF2. Median age at onset was 4.1 years in patients without a mutation (range 0.5–18.8), and 3.2 years in those in whom the causative mutation was detected (range 0.1–14.3). Mutations in dominant genes presented with a median onset of 4.5 years (range 3.2–14.3). Mutations in recessive genes presented with a median onset of 0.5 years (range 0.1–3.2). Our molecular genetic diagnostic study identified underlying monogenic causes of steroid-resistant nephrotic syndrome in ~11% of patients with SRNS using a cost-effective technique. We delineated some of the therapeutic, diagnostic, and prognostic implications. Our study confirms that genetic testing is indicated in pediatric patients with SRNS.

The aim of this study was to elucidate whether genetic screening test results of pediatric patients with steroid-resistant nephrotic syndrome (SRNS) vary with ethnicity.Using high-throughput DNA sequencing, 28 nephrotic syndrome-related genes were analyzed in 110 chil-dren affected by SRNS and 10 children with isolated proteinuria enrolled by 5 centers in China (67 boys, 53 girls). Their age at disease onset ranged from 1 day to 208 months (median, 48.8 months). Patients were excluded if their age at onset of disease was over 18 years or if they were diagnosed as having Alport syndrome.A genetic etiology was identified in 28.3% of our cohort and the likelihood of establishing a genetic diagnosis decreased as the age at onset of nephrotic syndrome increased. The most common mutated genes were ADCK4 (6.67%), NPHS1 (5.83%), WT1 (5.83%), and NPHS2 (3.33%), and the difference in the frequencies of ADCK4 and NPHS2 mutations between this study and a study on monogenic causes of SRNS in the largest international cohort of 1,783 different families was significant. A case of congenital nephrotic syndrome was attributed to a homozygous missense mutation in ADCK4, and a de novo missense mutation in TRPC6 was detected in a case of infantile nephrotic syndrome.Our results showed that, in the first and the largest multicenter cohort of Chinese pediatric SRNS reported to date, ADCK4 is the most common causative gene, whereas there is a low prevalence of NPHS2 mutations. Our data indicated that the genetic testing results for pediatric SRNS patients vary with different ethnicities, and this information will help to improve management of the disease in clinical practice.

Nephrolithiasis, a condition in which urinary supersaturation leads to stone formation in the urinary system, affects about 5%-10% of individuals worldwide at some point in their lifetime and results in significant medical costs and morbidity. To date, mutations in more than 30 genes have been described as being associated with nephrolithiasis, and these mutations explain about 15% of kidney stone cases, suggesting that additional nephrolithiasis-associated genes remain to be discovered. To identify additional genes whose mutations are linked to nephrolithiasis, we performed targeted next- generation sequencing of 18 hypothesized candidate genes in 348 unrelated individuals with kidney stones. We detected biallelic mutations in SLC26A1 (solute carrier family 26 member 1) in two unrelated individuals with calcium oxalate kidney stones. We show by immunofluorescence, immunoblotting, and glycosylation analysis that the variant protein mimicking p.Thr185Met has defects in protein folding or trafficking. In addition, by measuring anion exchange activity of SLC26A1, we demonstrate that all the identified mutations in SLC26A1 result in decreased transporter activity. Our data identify SLC26A1 mutations as causing a recessive Mendelian form of nephrolithiasis.

Background Whole-exome sequencing (WES) finds a CKD-related mutation in approximately 20% of patients presenting with CKD before 25 years of age. Although provision of a molecular diagnosis could have important implications for clinical management, evidence is lacking on the diagnostic yield and clinical utility of WES for pediatric renal transplant recipients. Methods To determine the diagnostic yield of WES in pediatric kidney transplant recipients, we recruited 104 patients who had received a transplant at Boston Children’s Hospital from 2007 through 2017, performed WES, and analyzed results for likely deleterious variants in approximately 400 genes known to cause CKD. Results By WES, we identified a genetic cause of CKD in 34 out of 104 (32.7%) transplant recipients. The likelihood of detecting a molecular genetic diagnosis was highest for patients with urinary stone disease (three out of three individuals), followed by renal cystic ciliopathies (seven out of nine individuals), steroid-resistant nephrotic syndrome (nine out of 21 individuals), congenital anomalies of the kidney and urinary tract (ten out of 55 individuals), and chronic glomerulonephritis (one out of seven individuals). WES also yielded a molecular diagnosis for four out of nine individuals with ESRD of unknown etiology. The WES-related molecular genetic diagnosis had implications for clinical care for five patients. Conclusions Nearly one third of pediatric renal transplant recipients had a genetic cause of their kidney disease identified by WES. Knowledge of this genetic information can help guide management of both transplant patients and potential living related donors.

No efficient treatment exists for nephrotic syndrome (NS), a frequent cause of chronic kidney disease. Here we show mutations in six different genes (MAGI2, TNS2, DLC1, CDK20, ITSN1, ITSN2) as causing NS in 17 families with partially treatment-sensitive NS (pTSNS). These proteins interact and we delineate their roles in Rho-like small GTPase (RLSG) activity, and demonstrate deficiency for mutants of pTSNS patients. We find that CDK20 regulates DLC1. Knockdown of MAGI2, DLC1, or CDK20 in cultured podocytes reduces migration rate. Treatment with dexamethasone abolishes RhoA activation by knockdown of DLC1 or CDK20 indicating that steroid treatment in patients with pTSNS and mutations in these genes is mediated by this RLSG module. Furthermore, we discover ITSN1 and ITSN2 as podocytic guanine nucleotide exchange factors for Cdc42. We generate Itsn2-L knockout mice that recapitulate the mild NS phenotype. We, thus, define a functional network of RhoA regulation, thereby revealing potential therapeutic targets., Nephrotic syndrome is the second most common chronic kidney disease but there are no targeted treatment strategies available. Here the authors identify mutations of six genes codifying for proteins involved in cytoskeleton remodelling and modulation of small GTPases in 17 families with nephrotic syndrome.

The amount of energy needed to operate high-performance computing systems increases regularly since some years at a high pace, and the energy consumption has attracted a great deal of attention. Moreover, high energy consumption inevitably contains failures and reduces system reliability. However, there has been considerably less work of simultaneous management of system performance, reliability, and energy consumption on heterogeneous systems. In this paper, we first build the precedence-constrained parallel applications and energy consumption model. Then, we deduce the relation between reliability and processor frequencies and get their parameters approximation value by least squares curve fitting method. Thirdly, we establish a task execution reliability model and formulate this reliability and energy aware scheduling problem as a linear programming. Lastly, we propose a heuristic Reliability-Energy Aware Scheduling (REAS) algorithm to solve this problem, which can get good tradeoff among system performance, reliability, and energy consumption with lower complexity. Our extensive simulation performance evaluation study clearly demonstrates the tradeoff performance of our proposed heuristic algorithm.

Steroid-resistant nephrotic syndrome (SRNS) represents the second most frequent cause of chronic kidney disease in the first three decades of life. It manifests histologically as focal segmental glomerulosclerosis (FSGS) and carries a 33% risk of relapse in a renal transplant. No efficient treatment exists. Identification of single-gene (monogenic) causes of SRNS has moved the glomerular epithelial cell (podocyte) to the center of its pathogenesis. Recently, mutations in >30 recessive or dominant genes were identified as causing monogenic forms of SRNS, thereby revealing the encoded proteins as essential for glomerular function. These findings helped define protein interaction complexes and functional pathways that could be targeted for treatment of SRNS. Very recently, it was discovered that in the surprisingly high fraction of ∼30% of all individuals who manifest with SRNS before 25 years of age, a causative mutation can be detected in one of the ∼30 different SRNS-causing genes. These findings revealed that SRNS and FSGS are not single disease entities but rather are part of a spectrum of distinct diseases with an identifiable genetic etiology. Mutation analysis should be offered to all individuals who manifest with SRNS before the age of 25 years, because (i) it will provide the patient and families with an unequivocal cause-based diagnosis, (ii) it may uncover a form of SRNS that is amenable to treatment (e.g. coenzyme Q(10)), (iii) it may allow avoidance of a renal biopsy procedure, (iv) it will further unravel the puzzle of pathogenic pathways of SRNS and (v) it will permit personalized treatment options for SRNS, based on genetic causation in way of ‘precision medicine’.

Galloway-Mowat syndrome (GAMOS) is a severe autosomal-recessive disease characterized by the combination of early-onset steroid-resistant nephrotic syndrome (SRNS) and microcephaly with brain anomalies. To date, mutations of WDR73 are the only known monogenic cause of GAMOS and in most affected individuals the molecular diagnosis remains elusive. We here identify recessive mutations of OSGEP, TP53RK, TPRKB, or LAGE3, encoding the 4 subunits of the KEOPS complex in 33 individuals of 30 families with GAMOS. CRISPR/Cas9 knockout in zebrafish and mice recapitulates the human phenotype of microcephaly and results in early lethality. Knockdown of OSGEP, TP53RK, or TPRKB inhibits cell proliferation, which human mutations fail to rescue, and knockdown of either gene activates DNA damage response signaling and induces apoptosis. OSGEP and TP53RK molecularly interact and co-localize with the actin-regulating ARP2/3 complex. Furthermore, knockdown of OSGEP and TP53RK induces defects of the actin cytoskeleton and reduces migration rate of human podocytes, an established intermediate phenotype of SRNS. We thus identify 4 novel monogenic causes of GAMOS, describe the first link between KEOPS function and human disease, and delineate potential pathogenic mechanisms.

Steroid-resistant nephrotic syndrome (SRNS) causes 15% of chronic kidney disease cases. A mutation in 1 of over 40 monogenic genes can be detected in approximately 30% of individuals with SRNS whose symptoms manifest before 25 years of age. However, in many patients, the genetic etiology remains unknown. Here, we have performed whole exome sequencing to identify recessive causes of SRNS. In 7 families with SRNS and facultative ichthyosis, adrenal insufficiency, immunodeficiency, and neurological defects, we identified 9 different recessive mutations in SGPL1, which encodes sphingosine-1-phosphate (S1P) lyase. All mutations resulted in reduced or absent SGPL1 protein and/or enzyme activity. Overexpression of cDNA representing SGPL1 mutations resulted in subcellular mislocalization of SGPL1. Furthermore, expression of WT human SGPL1 rescued growth of SGPL1-deficient dpl1. yeast strains, whereas expression of disease-associated variants did not. Immunofluorescence revealed SGPL1 expression in mouse podocytes and mesangial cells. Knockdown of Sgpl1 in rat mesangial cells inhibited cell migration, which was partially rescued by VPC23109, an S1P receptor antagonist. In Drosophila, Sply mutants, which lack SGPL1, displayed a phenotype reminiscent of nephrotic syndrome in nephrocytes. WT Sply, but not the disease-associated variants, rescued this phenotype. Together, these results indicate that SGPL1 mutations cause a syndromic form of SRNS.

PubMed ID: 29058690, Steroid-resistant nephrotic syndrome (SRNS) is a frequent cause of chronic kidney disease. Here, we identified recessive mutations in the gene encoding the actin-binding protein advillin (AVIL) in 3 unrelated families with SRNS. While all AVIL mutations resulted in a marked loss of its actin-bundling ability, truncation of AVIL also disrupted colocalization with F-actin, thereby leading to impaired actin binding and severing. Additionally, AVIL colocalized and interacted with the phospholipase enzyme PLCE1 and with the ARP2/3 actin-modulating complex. Knockdown of AVIL in human podocytes reduced actin stress fibers at the cell periphery, prevented recruitment of PLCE1 to the ARP3-rich lamellipodia, blocked EGF-induced generation of diacylglycerol (DAG) by PLCE1, and attenuated the podocyte migration rate (PMR). These effects were reversed by overexpression of WT AVIL but not by overexpression of any of the 3 patient-derived AVIL mutants. The PMR was increased by overexpression of WT Avil or PLCE1, or by EGF stimulation; however, this increased PMR was ameliorated by inhibition of the ARP2/3 complex, indicating that ARP-dependent lamellipodia formation occurs downstream of AVIL and PLCE1 function. Together, these results delineate a comprehensive pathogenic axis of SRNS that integrates loss of AVIL function with alterations in the action of PLCE1, an established SRNS protein., VE 196/1-1, HE 7456/1-1, Jo 1324/1-1 U.S. Public Health Service: DK-56338 National Institutes of Health: DK076683 National Institute of Diabetes and Digestive and Kidney Diseases: DK-98120 National Research Foundation of Korea, NRF: 2015R1D1A1A01056685 Department of Science and Technology, Government of Kerala NAS LPDS-2015-07 Fudan University, We are grateful to the families and study participants for their contributions. We thank the Yale Center for Mendelian Genomics (U54HG006504) for WES analysis. FH is a William E. Harmon Professor of Pediatrics. This research was supported by the NIH (DK076683, to FH); the Young Scholars Program of Children’s Hospital of Fudan University (to JR); Basic Science Research Program through the National Research Foundation of Korea 2015R1D1A1A01056685 (to HYG); DFG fellowships (VE 196/1-1, to ATvdV; Jo 1324/1-1, to TJS; and HE 7456/1-1, to TH); the German National Academy of Sciences Leopoldina (LPDS-2015-07, to EW); the Egyptian Group for Orphan Renal Diseases (EGORD) (to NAS); the Department of Science and Technology, Government of India (DST-SERB, to MAJ); the National Institute of Diabetes and Digestive and Kidney Diseases (DK-98120, to SK); and the Public Health Service (DK-56338, to SK).

Widmeier E, Tan W, Airik M, Hildebrandt F. A small molecule screening to detect potential therapeutic targets in human podocytes. Am J Physiol Renal Physiol 312: F157–F171, 2017. First published October 19, 2016; doi:10.1152/ajprenal.00386.2016. Steroid-resistant nephrotic syndrome (SRNS) inevitably progresses to end-stage kidney disease, requiring dialysis or transplantation for survival. However, treatment modalities and drug discovery remain limited. Mutations in over 30 genes have been discovered as monogenic causes of SRNS. Most of these genes are predominantly expressed in the glomerular epithelial cell, the podocyte, placing it at the center of the pathogenesis of SRNS. Podocyte migration rate (PMR) represents a relevant intermediate phenotype of disease in monogenic causes of SRNS. We therefore adapted PMR in a high-throughput manner to screen small molecules as potential therapeutic targets for SRNS. We performed a high-throughput drug screening of a National Institutes of Health Clinical Collection (NCC) library ( n = 725 compounds) measuring PMR by videomicroscopy. We used the Woundmaker to perform individual 96-well scratch wounds and screened compounds using a quantitative kinetic live cell imaging migration assay using IncuCyte ZOOM technology. Using a normal distribution for the average PMR in wild-type podocytes with a vehicle control (DMSO), we applied a 90% confidence interval to define “distinct” compounds (5% faster/slower PMR) and found that 12 of 725 compounds (at 10 μM) reduced PMR. Clusters of drugs that alter PMR included actin/tubulin modulators such as the azole class of antifungals and antineoplastic vinca-alkaloids. We hereby identify compounds that alter PMR. The PMR assay provides a new avenue to test therapeutics for nephrotic syndrome. Positive results may reveal novel pathways in the study of glomerular diseases such as SRNS.

Background and objectives Nephrolithiasis is a prevalent condition that affects 10%–15% of adults in their lifetime. It is associated with high morbidity due to colicky pain, the necessity for surgical intervention, and sometimes progression to CKD. In recent years, multiple monogenic causes of nephrolithiasis and nephrocalcinosis have been identified. However, the prevalence of each monogenic gene in a pediatric renal stone cohort has not yet been extensively studied. Design, setting, participants, & measurements To determine the percentage of cases that can be explained molecularly by mutations in one of 30 known nephrolithiasis/nephrocalcinosis genes, we conducted a high-throughput exon sequencing analysis in an international cohort of 143 individuals n =123) or isolated nephrocalcinosis ( n =20). Over 7 months, all eligible individuals at three renal stone clinics in the United States and Europe were approached for study participation. Results We detected likely causative mutations in 14 of 30 analyzed genes, leading to a molecular diagnosis in 16.8% (24 of 143) of affected individuals; 12 of the 27 detected mutations were not previously described as disease causing (44.4%). We observed that in our cohort all individuals with infantile manifestation of nephrolithiasis or nephrocalcinosis had causative mutations in recessive rather than dominant monogenic genes. In individuals who manifested later in life, causative mutations in dominant genes were more frequent. Conclusions We present the first exclusively pediatric cohort examined for monogenic causes of nephrolithiasis/nephrocalcinosis, and suggest that important therapeutic and preventative measures may result from mutational analysis in individuals with early manifestation of nephrolithiasis or nephrocalcinosis.

Nucleoporins are essential components of the nuclear pore complex (NPC). Only a few diseases have been attributed to NPC dysfunction. Steroid-resistant nephrotic syndrome (SRNS), a frequent cause of chronic kidney disease, is caused by dysfunction of glomerular podocytes. Here we identify in eight families with SRNS mutations in NUP93, its interaction partner NUP205 or XPO5 (encoding exportin 5) as hitherto unrecognized monogenic causes of SRNS. NUP93 mutations caused disrupted NPC assembly. NUP93 knockdown reduced the presence of NUP205 in the NPC, and, reciprocally, a NUP205 alteration abrogated NUP93 interaction. We demonstrate that NUP93 and exportin 5 interact with the signaling protein SMAD4 and that NUP93 mutations abrogated interaction with SMAD4. Notably, NUP93 mutations interfered with BMP7-induced SMAD transcriptional reporter activity. We hereby demonstrate that mutations of NUP genes cause a distinct renal disease and identify aberrant SMAD signaling as a new disease mechanism of SRNS, opening a potential new avenue for treatment.