Understanding the Current Landscape of Kidney Disease in Canada to Advance Precision Medicine Guided Personalized Care.

Purpose of Review: To understand the impact of kidney disease in Canada and the priority areas of kidney research that can benefit from patient-oriented, precision medicine research using novel technologies. Sources of Information: Information was collected through discussions between health care professionals, researchers, and patient partners. Literature was compiled using search engines (PubMed, PubMed central, Medline, and Google) and data from the Canadian Organ Replacement Register. Methods: We reviewed the impact, prevalence, economic burden, causes of kidney disease, and priority research areas in Canada. After reviewing the priority areas for kidney research, potential avenues for future research that can integrate precision medicine initiatives for patient-oriented research were outlined. Key Findings: Chronic kidney disease (CKD) remains among the top causes of morbidity and mortality in the world and exerts a large financial strain on the health care system. Despite the increasing number of people with CKD, funding for basic kidney research continues to trail behind other diseases. Current funding strategies favor existing clinical treatment and patient educational strategies. The identification of genetic factors for various forms of kidney disease in the adult and pediatric populations provides mechanistic insight into disease pathogenesis. Allocation of resources and funding toward existing high-yield personalized research initiatives have the potential to significantly affect patient-oriented research outcomes but will be difficult due to a constant decline of funding for kidney research. Limitations: This is an overview primarily focused on Canadian-specific literature rather than a comprehensive systematic review of the literature. The scope of our findings and conclusions may not be applicable to health care systems in other countries.

Justification: Cette étude visait à mieux comprendre l'impact de l'insuffisance rénale au Canada et à déterminer les domaines de recherche prioritaires en santé rénale qui pourraient tirer profit des recherches axées sur le patient qui sont menées en médecine de précision avec de nouvelles technologies. Sources: L'information provient de discussions entre les professionnels de la santé, les chercheurs et les patients partenaires. La littérature a été compilée à l'aide de moteurs de recherche (PubMed, PubMed central, Medline et Google) et de données provenant du Registre canadien des insuffisances et des transplantations d'organes. Méthodologie: Nous avons examiné les répercussions, la prévalence, le fardeau économique et les causes des maladies rénales, ainsi que les domaines prioritaires de la recherche au Canada. Après avoir examiné les domaines de recherche prioritaires en santé rénale, nous avons décrit les possibles orientations de recherche pouvant intégrer des initiatives de la recherche axée sur le patient menée en médecine de précision. Principaux résultats: L'insuffisance rénale chronique (IRC) demeure l'une des principales causes de morbidité et de mortalité dans le monde et elle exerce une forte pression financière sur le système de santé. Malgré le nombre croissant de personnes atteintes d'IRC, le financement de la recherche fondamentale en santé rénale reste à la traîne des autres maladies; et le financement actuel favorise les stratégies existantes pour le traitement clinique et l'éducation des patients. L'identification de facteurs génétiques liés aux diverses formes de néphropathies dans les populations adultes et pédiatriques fournit une compréhension mécanistique de la pathogenèse de la maladie. L'attribution des ressources et du financement à des initiatives de recherche personnalisées existantes et connues pour leur rendement élevé pourrait avoir une incidence considérable sur les résultats de la recherche axée sur le patient, mais elle sera difficile compte tenu de la diminution constante du financement de la recherche en santé rénale. Limites: Il s'agit d'un portrait axé principalement sur la littérature canadienne et non d'un examen systématique complet de la littérature. La portée des résultats et conclusions pourrait ne pas s'appliquer aux systèmes de santé d'autres pays.

Primary cilia and actin regulatory pathways in renal ciliopathies.

Ciliopathies are a group of rare genetic disorders caused by defects to the structure or function of the primary cilium. They often affect multiple organs, leading to brain malformations, congenital heart defects, and anomalies of the retina or skeletal system. Kidney abnormalities are among the most frequent ciliopathic phenotypes manifesting as smaller, dysplastic, and cystic kidneys that are often accompanied by renal fibrosis. Many renal ciliopathies cause chronic kidney disease and often progress to end-stage renal disease, necessitating replacing therapies. There are more than 35 known ciliopathies; each is a rare hereditary condition, yet collectively they account for a significant proportion of chronic kidney disease worldwide. The primary cilium is a tiny microtubule-based organelle at the apex of almost all vertebrate cells. It serves as a "cellular antenna" surveying environment outside the cell and transducing this information inside the cell to trigger multiple signaling responses crucial for tissue morphogenesis and homeostasis. Hundreds of proteins and unique cellular mechanisms are involved in cilia formation. Recent evidence suggests that actin remodeling and regulation at the base of the primary cilium strongly impacts ciliogenesis. In this review, we provide an overview of the structure and function of the primary cilium, focusing on the role of actin cytoskeleton and its regulators in ciliogenesis. We then describe the key clinical, genetic, and molecular aspects of renal ciliopathies. We highlight what is known about actin regulation in the pathogenesis of these diseases with the aim to consider these recent molecular findings as potential therapeutic targets for renal ciliopathies.

Whole exome sequencing identifies potential candidate genes for spina bifida derived from mouse models.

Spina bifida (SB) is the second most common nonlethal congenital malformation. The existence of monogenic SB mouse models and human monogenic syndromes with SB features indicate that human SB may be caused by monogenic genes. We hypothesized that whole exome sequencing (WES) allows identification of potential candidate genes by (i) generating a list of 136 candidate genes for SB, and (ii) by unbiased exome-wide analysis. We generated a list of 136 potential candidate genes from three categories and evaluated WES data of 50 unrelated SB cases for likely deleterious variants in 136 potential candidate genes, and for potential SB candidate genes exome-wide. We identified 6 likely deleterious variants in 6 of the 136 potential SB candidate genes in 6 of the 50 SB cases, whereof 4 genes were derived from mouse models, 1 gene was derived from human nonsyndromic SB, and 1 gene was derived from candidate genes known to cause human syndromic SB. In addition, by unbiased exome-wide analysis, we identified 12 genes as potential candidates for SB. Identification of these 18 potential candidate genes in larger SB cohorts will help decide which ones can be considered as novel monogenic causes of human SB.

ITSN1: a novel candidate gene involved in autosomal dominant neurodevelopmental disorder spectrum.

ITSN1 plays an important role in brain development. Recent studies in large cohorts of subjects with neurodevelopmental disorders have identified de novo variants in ITSN1 gene thereby suggesting that this gene is involved in the development of such disorders. The aim of this study is to provide further proof of such a link. We performed trio exome sequencing in a patient presenting autism, intellectual disability, and severe behavioral difficulties. Additional affected patients with a neurodevelopmental disorder harboring a heterozygous variant in ITSN1 (NM_003024.2) were collected through a worldwide collaboration. All patients underwent detailed phenotypic and genetic assessment and data was collected and shared by healthcare givers. We identified ten novel patients from eight families with heterozygous truncating or missense variants in ITSN1 gene. In addition, four previously published patients from large meta-analysis studies were included. In total, 7/14 patients presented a de novo variant in ITSN1. All patients showed neurodevelopmental disorders from autism spectrum disorders (90%), intellectual disability (86%), and epilepsy (30%). We demonstrated that truncating variants are in the first half of ITSN1 whereas missense variants are clustered in C-terminal region. We suggest ITSN1 gene is involved in development of an autism spectrum disorder with variable additional neurodevelopmental deficiency, thus confirming the hypothesis that ITSN1 is important for brain development.

Exome survey of individuals affected by VATER/VACTERL with renal phenotypes identifies phenocopies and novel candidate genes.

The acronym VATER/VACTERL refers to the rare nonrandom association of the following component features (CFs): vertebral defects (V), anorectal malformations (ARM) (A), cardiac anomalies (C), tracheoesophageal fistula with or without esophageal atresia (TE), renal malformations (R), and limb anomalies (L). For the clinical diagnosis, the presence of at least three CFs is required, individuals presenting with only two CFs have been categorized as VATER/VACTERL-like. The majority of VATER/VACTERL individuals displays a renal phenotype. Hitherto, variants in FGF8, FOXF1, HOXD13, LPP, TRAP1, PTEN, and ZIC3 have been associated with the VATER/VACTERL association; however, large-scale re-sequencing could only confirm TRAP1 and ZIC3 as VATER/VACTERL disease genes, both associated with a renal phenotype. In this study, we performed exome sequencing in 21 individuals and their families with a renal VATER/VACTERL or VATER/VACTERL-like phenotype to identify potentially novel genetic causes. Exome analysis identified biallelic and X-chromosomal hemizygous potentially pathogenic variants in six individuals (29%) in B9D1, FREM1, ZNF157, SP8, ACOT9, and TTLL11, respectively. The online tool GeneMatcher revealed another individual with a variant in ZNF157. Our study suggests six biallelic and X-chromosomal hemizygous VATER/VACTERL disease genes implicating all six genes in the expression of human renal malformations.

Recessive NOS1AP variants impair actin remodeling and cause glomerulopathy in humans and mice.

Nephrotic syndrome (NS) is a leading cause of chronic kidney disease. We found recessive NOS1AP variants in two families with early-onset NS by exome sequencing. Overexpression of wild-type (WT) NOS1AP, but not cDNA constructs bearing patient variants, increased active CDC42 and promoted filopodia and podosome formation. Pharmacologic inhibition of CDC42 or its effectors, formin proteins, reduced NOS1AP-induced filopodia formation. NOS1AP knockdown reduced podocyte migration rate (PMR), which was rescued by overexpression of WT Nos1ap but not by constructs bearing patient variants. PMR in NOS1AP knockdown podocytes was also rescued by constitutively active CDC42Q61L or the formin DIAPH3 Modeling a NOS1AP patient variant in knock-in human kidney organoids revealed malformed glomeruli with increased apoptosis. Nos1apEx3-/Ex3- mice recapitulated the human phenotype, exhibiting proteinuria, foot process effacement, and glomerulosclerosis. These findings demonstrate that recessive NOS1AP variants impair CDC42/DIAPH-dependent actin remodeling, cause aberrant organoid glomerulogenesis, and lead to a glomerulopathy in humans and mice.

Generation of Monogenic Candidate Genes for Human Nephrotic Syndrome Using 3 Independent Approaches.

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.

Recessive Mutations in SYNPO2 as a Candidate of Monogenic Nephrotic Syndrome.

INTRODUCTION: Most of the approximately 60 genes that if mutated cause steroid-resistant nephrotic syndrome (SRNS) are highly expressed in the glomerular podocyte, rendering SRNS a "podocytopathy." METHODS: We performed whole-exome sequencing (WES) in 1200 nephrotic syndrome (NS) patients. RESULTS: We discovered homozygous truncating and homozygous missense mutation in SYNPO2 (synaptopodin-2) (p.Lys1124∗ and p.Ala1134Thr) in 2 patients with childhood-onset NS. We found SYNPO2 expression in both podocytes and mesangial cells; however, notably, immunofluorescence staining of adult human and rat kidney cryosections indicated that SYNPO2 is localized mainly in mesangial cells. Subcellular localization studies reveal that in these cells SYNPO2 partially co-localizes with α-actinin and filamin A-containing F-actin filaments. Upon transfection in mesangial cells or podocytes, EGFP-SYNPO2 co-localized with α-actinin-4, which gene is mutated in autosomal dominant SRNS in humans. SYNPO2 overexpression increases mesangial cell migration rate (MMR), whereas shRNA knockdown reduces MMR. Decreased MMR was rescued by transfection of wild-type mouse Synpo2 cDNA but only partially by cDNA representing mutations from the NS patients. The increased mesangial cell migration rate (MMR) by SYNPO2 overexpression was inhibited by ARP complex inhibitor CK666. SYNPO2 shRNA knockdown in podocytes decreased active Rac1, which was rescued by transfection of wild-type SYNPO2 cDNA but not by cDNA representing any of the 2 mutant variants. CONCLUSION: We show that SYNPO2 variants may lead to Rac1-ARP3 dysregulation, and may play a role in the pathogenesis of nephrotic syndrome.

Mutations in PRDM15 Are a Novel Cause of Galloway-Mowat Syndrome.

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.

De novo TRIM8 variants impair its protein localization to nuclear bodies and cause developmental delay, epilepsy, and focal segmental glomerulosclerosis.

Focal segmental glomerulosclerosis (FSGS) is the main pathology underlying steroid-resistant nephrotic syndrome (SRNS) and a leading cause of chronic kidney disease. Monogenic forms of pediatric SRNS are predominantly caused by recessive mutations, while the contribution of de novo variants (DNVs) to this trait is poorly understood. Using exome sequencing (ES) in a proband with FSGS/SRNS, developmental delay, and epilepsy, we discovered a nonsense DNV in TRIM8, which encodes the E3 ubiquitin ligase tripartite motif containing 8. To establish whether TRIM8 variants represent a cause of FSGS, we aggregated exome/genome-sequencing data for 2,501 pediatric FSGS/SRNS-affected individuals and 48,556 control subjects, detecting eight heterozygous TRIM8 truncating variants in affected subjects but none in control subjects (p = 3.28 × 10-11). In all six cases with available parental DNA, we demonstrated de novo inheritance (p = 2.21 × 10-15). Reverse phenotyping revealed neurodevelopmental disease in all eight families. We next analyzed ES from 9,067 individuals with epilepsy, yielding three additional families with truncating TRIM8 variants. Clinical review revealed FSGS in all. All TRIM8 variants cause protein truncation clustering within the last exon between residues 390 and 487 of the 551 amino acid protein, indicating a correlation between this syndrome and loss of the TRIM8 C-terminal region. Wild-type TRIM8 overexpressed in immortalized human podocytes and neuronal cells localized to nuclear bodies, while constructs harboring patient-specific variants mislocalized diffusely to the nucleoplasm. Co-localization studies demonstrated that Gemini and Cajal bodies frequently abut a TRIM8 nuclear body. Truncating TRIM8 DNVs cause a neuro-renal syndrome via aberrant TRIM8 localization, implicating nuclear bodies in FSGS and developmental brain disease.

Mutations in transcription factor CP2-like 1 may cause a novel syndrome with distal renal tubulopathy in humans.

BACKGROUND: An underlying monogenic cause of early-onset chronic kidney disease (CKD) can be detected in ∼20% of individuals. For many etiologies of CKD manifesting before 25 years of age, >200 monogenic causative genes have been identified to date, leading to the elucidation of mechanisms of renal pathogenesis. METHODS: In 51 families with echogenic kidneys and CKD, we performed whole-exome sequencing to identify novel monogenic causes of CKD. RESULTS: We discovered a homozygous truncating mutation in the transcription factor gene transcription factor CP2-like 1 (TFCP2L1) in an Arabic patient of consanguineous descent. The patient developed CKD by the age of 2 months and had episodes of severe hypochloremic, hyponatremic and hypokalemic alkalosis, seizures, developmental delay and hypotonia together with cataracts. We found that TFCP2L1 was localized throughout kidney development particularly in the distal nephron. Interestingly, TFCP2L1 induced the growth and development of renal tubules from rat mesenchymal cells. Conversely, the deletion of TFCP2L1 in mice was previously shown to lead to reduced expression of renal cell markers including ion transporters and cell identity proteins expressed in different segments of the distal nephron. TFCP2L1 localized to the nucleus in HEK293T cells only upon coexpression with its paralog upstream-binding protein 1 (UBP1). A TFCP2L1 mutant complementary DNA (cDNA) clone that represented the patient's mutation failed to form homo- and heterodimers with UBP1, an essential step for its transcriptional activity. CONCLUSION: Here, we identified a loss-of-function TFCP2L1 mutation as a potential novel cause of CKD in childhood accompanied by a salt-losing tubulopathy.

Rho GTPase regulatory proteins in podocytes.

The Rho family of small GTPases (Rho GTPases) are the master regulators of the actin cytoskeleton and consist of 22 members. Previous studies implicated dysregulation of Rho GTPases in podocytes in the pathogenesis of proteinuric glomerular diseases. Rho GTPases are primarily regulated by the three families of proteins; guanine nucleotide exchange factors (GEFs; 82 members), GTPase-activating proteins (GAPs; 69 members), and GDP dissociation inhibitors (GDIs; 3 members). Since the regulatory proteins far outnumber their substrate Rho GTPases and act in concert in a cell/context-dependent manner, the upstream regulatory mechanism directing Rho GTPases in podocytes is largely unknown. In this review, we summarize recent advances in the understanding of the role of Rho GTPase regulatory proteins in podocytes, including the known mutations of these proteins that cause proteinuria in humans. We also provide critical appraisal of the in vivo and in vitro studies and identify the knowledge gap in the field that will require further studies.

DAAM2 Variants Cause Nephrotic Syndrome via Actin Dysregulation.

The discovery of >60 monogenic causes of nephrotic syndrome (NS) has revealed a central role for the actin regulators RhoA/Rac1/Cdc42 and their effectors, including the formin INF2. By whole-exome sequencing (WES), we here discovered bi-allelic variants in the formin DAAM2 in four unrelated families with steroid-resistant NS. We show that DAAM2 localizes to the cytoplasm in podocytes and in kidney sections. Further, the variants impair DAAM2-dependent actin remodeling processes: wild-type DAAM2 cDNA, but not cDNA representing missense variants found in individuals with NS, rescued reduced podocyte migration rate (PMR) and restored reduced filopodia formation in shRNA-induced DAAM2-knockdown podocytes. Filopodia restoration was also induced by the formin-activating molecule IMM-01. DAAM2 also co-localizes and co-immunoprecipitates with INF2, which is intriguing since variants in both formins cause NS. Using in vitro bulk and TIRF microscopy assays, we find that DAAM2 variants alter actin assembly activities of the formin. In a Xenopus daam2-CRISPR knockout model, we demonstrate actin dysregulation in vivo and glomerular maldevelopment that is rescued by WT-DAAM2 mRNA. We conclude that DAAM2 variants are a likely cause of monogenic human SRNS due to actin dysregulation in podocytes. Further, we provide evidence that DAAM2-associated SRNS may be amenable to treatment using actin regulating compounds.

Mutations of the Transcriptional Corepressor ZMYM2 Cause Syndromic Urinary Tract Malformations.

Congenital anomalies of the kidney and urinary tract (CAKUT) constitute one of the most frequent birth defects and represent the most common cause of chronic kidney disease in the first three decades of life. Despite the discovery of dozens of monogenic causes of CAKUT, most pathogenic pathways remain elusive. We performed whole-exome sequencing (WES) in 551 individuals with CAKUT and identified a heterozygous de novo stop-gain variant in ZMYM2 in two different families with CAKUT. Through collaboration, we identified in total 14 different heterozygous loss-of-function mutations in ZMYM2 in 15 unrelated families. Most mutations occurred de novo, indicating possible interference with reproductive function. Human disease features are replicated in X. tropicalis larvae with morpholino knockdowns, in which expression of truncated ZMYM2 proteins, based on individual mutations, failed to rescue renal and craniofacial defects. Moreover, heterozygous Zmym2-deficient mice recapitulated features of CAKUT with high penetrance. The ZMYM2 protein is a component of a transcriptional corepressor complex recently linked to the silencing of developmentally regulated endogenous retrovirus elements. Using protein-protein interaction assays, we show that ZMYM2 interacts with additional epigenetic silencing complexes, as well as confirming that it binds to FOXP1, a transcription factor that has also been linked to CAKUT. In summary, our findings establish that loss-of-function mutations of ZMYM2, and potentially that of other proteins in its interactome, as causes of human CAKUT, offering new routes for studying the pathogenesis of the disorder.

COL4A1 mutations as a potential novel cause of autosomal dominant CAKUT in humans.

Congenital anomalies of the kidney and urinary tract (CAKUT) are the most common cause of chronic kidney disease (~ 45%) that manifests before 30 years of age. The genetic locus containing COL4A1 (13q33-34) has been implicated in vesicoureteral reflux (VUR), but mutations in COL4A1 have not been reported in CAKUT. We hypothesized that COL4A1 mutations cause CAKUT in humans. We performed whole exome sequencing (WES) in 550 families with CAKUT. As negative control cohorts we used WES sequencing data from patients with nephronophthisis (NPHP) with no genetic cause identified (n = 257) and with nephrotic syndrome (NS) due to monogenic causes (n = 100). We identified a not previously reported heterozygous missense variant in COL4A1 in three siblings with isolated VUR. When examining 549 families with CAKUT, we identified nine additional different heterozygous missense mutations in COL4A1 in 11 individuals from 11 unrelated families with CAKUT, while no COL4A1 mutations were identified in a control cohort with NPHP and only one in the cohort with NS. Most individuals (12/14) had isolated CAKUT with no extrarenal features. The predominant phenotype was VUR (9/14). There were no clinical features of the COL4A1-related disorders (e.g., HANAC syndrome, porencephaly, tortuosity of retinal arteries). Whereas COL4A1-related disorders are typically caused by glycine substitutions in the collagenous domain (84.4% of variants), only one variant in our cohort is a glycine substitution within the collagenous domain (1/10). We identified heterozygous COL4A1 mutations as a potential novel autosomal dominant cause of CAKUT that is allelic to the established COL4A1-related disorders and predominantly caused by non-glycine substitutions.

Monogenic causes of chronic kidney disease in adults.

Approximately 500 monogenic causes of chronic kidney disease (CKD) have been identified, mainly in pediatric populations. The frequency of monogenic causes among adults with CKD has been less extensively studied. To determine the likelihood of detecting monogenic causes of CKD in adults presenting to nephrology services in Ireland, we conducted whole exome sequencing (WES) in a multi-centre cohort of 114 families including 138 affected individuals with CKD. Affected adults were recruited from 78 families with a positive family history, 16 families with extra-renal features, and 20 families with neither a family history nor extra-renal features. We detected a pathogenic mutation in a known CKD gene in 42 of 114 families (37%). A monogenic cause was identified in 36% of affected families with a positive family history of CKD, 69% of those with extra-renal features, and only 15% of those without a family history or extra-renal features. There was no difference in the rate of genetic diagnosis in individuals with childhood versus adult onset CKD. Among the 42 families in whom a monogenic cause was identified, WES confirmed the clinical diagnosis in 17 (40%), corrected the clinical diagnosis in 9 (22%), and established a diagnosis for the first time in 16 families referred with CKD of unknown etiology (38%). In this multi-centre study of adults with CKD, a molecular genetic diagnosis was established in over one-third of families. In the evolving era of precision medicine, WES may be an important tool to identify the cause of CKD in adults.

Whole-Exome Sequencing Enables a Precision Medicine Approach for Kidney Transplant Recipients.

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.

Acute and Chronic Management in an Atypical Case of Ethylmalonic Encephalopathy.

Ethylmalonic encephalopathy (EE) is caused by mutations in the ETHE1 gene. ETHE1 is vital for the catabolism of hydrogen sulfide (H2S). Patients with pathogenic mutations in ETHE1 have markedly increased thiosulfate, which is a reliable index of H2S levels. Accumulation of H2S is thought to cause the characteristic metabolic derangement found in EE. Recently introduced treatment strategies in EE, such as combined use of metronidazole (MNZ) and N-acetylcysteine (NAC), are aimed at lowering chronic H2S load. Experience with treatment strategies directed against acute episodes of metabolic decompensation (e.g., hemodialysis) is limited. Here we present an unusually mild, molecularly confirmed, case of EE in a 19-year-old male on chronic treatment with MNZ and NAC. During an acute episode of metabolic decompensation, we employed continuous renal replacement therapy (CRRT) to regain metabolic control. On continuous treatment with NAC and MNZ during the months preceding the acute event, plasma thiosulfate levels ranged from 1.6 to 4 µg/mL (reference range up to 2 µg/mL) and had a mean value of 2.5 µg/mL. During the acute decompensation, thiosulfate levels were 6.7 µg/mL, with hyperlactatemia and perturbed organic acid, acylglycine, and acylcarnitine profiles. CRRT decreased thiosulfate within 24 h to 1.4 µg/mL. Following discontinuation of CRRT, mean thiosulfate levels were 3.2 µg/mL (range, 2.4-3.7 µg/mL) accompanied by clinical improvement with metabolic stabilization of blood gas, acylcarnitine, organic acid, and acylglycine profiles. In conclusion, CRRT may help to regain metabolic control in patients with EE who have an acute metabolic decompensation on chronic treatment with NAC and MNZ.

Mutations in multiple components of the nuclear pore complex cause nephrotic syndrome.

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.

Whole-Exome Sequencing Identifies Causative Mutations in Families with Congenital Anomalies of the Kidney and Urinary Tract.

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.