The most frequent causative genes in podocytopathies are NPHS1 and NPHS2 that encode the main slit diaphragm components nephrin and podocin, respectively. The significance of the nephrin-podocin interaction has remained elusive. The NPHS2 R229Q variant is unique in human genetics as it is only pathogenic when trans-associated to specific 3' variants causing R229Q podocin to form distorted heterooligomers. To examine the effect of podocin on the distance between nephrin molecules, nephrin constructs labeled with YPet or mCherry/mRuby were transitionally coexpressed in HEK-293 cells. We assessed the nephrin-nephrin distance based on the Förster Resonance Energy Transfer (FRET) efficiency between YPet and mCherry/mRuby in living cells. Podocin markedly increased the FRET efficiency between nephrin molecules, reflecting reduced and ordered nephrin-nephrin distances. Its effect was abolished by pathogenic podocin variants. Pathogenic R229Q heterooligomers exhibited increased FRET efficiency between their PHB domains that correlated with a larger nephrin-nephrin distance, thereby explaining the associated mild phenotype. The effect of pathogenic and benign R229Q heterooligomers separated sharply in their effect on nephrin-nephrin spacing (P = 1.19E-33). Based on an intermediate effect on the nephrin-nephrin distance and five families with late-onset FSGS, we reconsider the R229Q-R286Tfs*17 association as pathogenic, but via a mechanism different from other R229Q associations and with incomplete penetrance. In conclusion, podocin provides regularly spaced intracellular anchor points for nephrin chains. Podocin homo-oligomerization affects the nephrin-nephrin spacing, thereby providing an explanation for albuminuria in patients with podocin dysfunction and for the interallelic interactions of R229Q.

Abstract Background ADTKD- MUC1 is one of the major entities of ADTKD caused by frameshift variants in the MUC1 VNTR that standard short-read sequencing fails to detect. Existing 59dupC-targeted probe-extension assays do not allow for broad screening and cannot detect atypical non-dupC variants. Recently, VNtyper, a Kestrel-based genotyping pipeline with optional code-adVNTR cross-validation for MUC1 VNTR genotyping from short-read sequencing data allowed to circumvent this diagnostic limitation, but needed further development for easy access and rapid sample processing. Methods We developed VNtyper 2, by refactoring VNtyper into a modular, production-grade tool with a companion web platform, VNtyper-Online ( https://vntyper.org ), for freely available browser-based analysis with short turnaround time and without local bioinformatics infrastructure. We validated VNtyper 2 on 400 simulated samples generated with MucOneUp and 142 clinical exomes with independently confirmed genotypes. Results In simulation, VNtyper 2 detected the canonical 59dupC variant with 96% sensitivity and 100% specificity. Reference-standard validation on 142 samples yielded 90.6% sensitivity and 98.2% specificity overall, with cohort-dependent performance across the Twist Exome v2 French-German cohort (98% sensitivity, 87.5% specificity) and the KAPA HyperExome V2 (Roche) Czech-US cohort (79.4% sensitivity, 100% specificity). Screening of 3582 exomes and targeted panels from international CKD referral programmes identified 51 positive individuals, including 9 with atypical non-dupC frameshift variants that would have been missed by 59dupC-targeted probe-extension assays. In unselected CKD cohorts, a descriptive random-effects summary estimated a detection rate of 1.4% (95% CI 0.6 to 3.1%). Conclusions VNtyper 2 and VNtyper-Online are open-source tools for MUC1 VNTR genotyping from short-read data and can support locally validated workflows when VNTR coverage is adequate. By improving accessibility and turnaround time, these tools democratize MUC1 diagnostics at global scale. For its integration into routine diagnostics, we propose an expert-informed two-pathway workflow developed through European ADTKD-Net consortium consensus.

Abstract N 6 -threonyl-carbamoylation of adenosine 37 of ANN-type tRNAs (t 6 A) is a universal modification essential for translational accuracy and efficiency. The t 6 A pathway uses two sequentially acting enzymes, YRDC and OSGEP, the latter being a subunit of the multiprotein KEOPS complex. Structures of the subunits and subcomplexes of human KEOPS are known, but knowledge on the detailed interactions with tRNA is lacking. We present here the first structure of complete hKEOPS and of its complex with a substrate tRNA by cryo-electron microscopy. The CAA tail of tRNA is bound to the TPRKB subunit and the anti-codon loop is positioned at the entrance of the catalytic site of OSGEP subunit. The flexibility of the OSGEP-TP53RK interface allows hKEOPS to fit the surface of the tRNA elbow. We recently identified mutations in all genes encoding for proteins of the t 6 A pathway in children with Galloway-Mowat syndrome (GAMOS), a clinically heterogeneous recessive disease characterized by early-onset steroid-resistant nephrotic syndrome and microcephaly. We here expressed and characterized the majority of the Galloway Mowat mutants. All mutants could be purified at high yields and seem to be stable in vitro . The t 6 A activity for most of the mutants is above 40% of the WT. Using CRISPR-Cas9 technology we replaced the genes encoding the t 6 A pathway proteins in yeast by their human homologues. This yeast construct was perfectly viable and produced WT levels of t 6 A modified tRNA. Using this tool, we observed that most of the GAMOS mutants were viable in yeast and yielded comparable t 6 A modified tRNA levels. Our data indicate that healthy human cellular development depends on an optimized level of t 6 A tRNA modification and is not compatible with a total loss of function of the t 6 A machinery.

Rationale & Objective: Molecular diagnosis of autosomal dominant tubulointerstitial kidney disease (ADTKD) due to variants in the MUC1 gene has long been challenging because variants lie in a large variable number of tandem repeat (VNTR) region, making identification impossible using standard short-read techniques. Previously, we addressed this diagnostic limitation by developing a computational pipeline named VNtyper for easier reliable detection of MUC1 VNTR pathogenic variants from short-read sequences. This led to unexpected diagnoses of ADTKD-MUC1 among patients with kidney disease referred for genetic testing, which we report here. Study Design: Cross-sectional observational study. Setting & Participants: 4,040 patients referred to Necker Enfants-Malades Hospital from 2017 to 2023 for genetic testing for (1) glomerular disease, (2) ciliopathy, (3) congenital anomalies of the kidneys and urinary tracts (CAKUT), (4) ADTKD, or (5) chronic kidney disease (CKD) of unknown origin, in whom MUC1 had not been previously tested by SNaPshot minisequencing. Exposure: Clinical suspicion of ADTKD. Outcome: ADTKD-MUC1 diagnosed using VNtyper. Analytical Approach: Data were collected from patients in whom ADTKD-MUC1 was newly diagnosed and patients in whom ADTKD was clinically suspected were compared with those in whom ADTKD was not. Results: We identified 40 patients with MUC1 variants by VNtyper, including 33 new index patients and 7 relatives. Of the 33 index cases, 20 had been suspected of having ADTKD based on clinical features, and in the other 13 ADTKD had not been considered. In patients in whom ADTKD had not been considered clinically, the detection rate was 0.05% (1 of 1,895) among patients with glomerular disease, 1.2% (4 of 329) among patients with ciliopathy, 0.09% (1 of 1,099) among patients with CAKUT and 2.5% (7 of 285) among patients with CKD of unknown origin. In 6 patients there was no family history of kidney disease, and we confirmed de novo presentation in 2 patients by segregation studies. Limitations: Observational study and selected referral population (may not represent the prevalence or phenotypes in the general kidney disease population). Conclusions: With VNtyper, we were able to diagnose new cases of ADTKD-MUC1 in a large cohort of patients with various phenotypes. Some patients had atypical phenotypes due to a variant in another gene, and some had no family history of kidney disease, suggesting de novo disease, which was confirmed in 2 patients.

Kidney organoids are an emerging tool for disease modeling, especially genetic diseases. Among them, X-linked Alport syndrome (XLAS) is a hematuric nephropathy affecting the glomerular basement membrane (GBM) secondary to pathogenic variations in the COL4A5 gene encoding the α5 subunit of type IV collagen [α5(IV)]. In patients carrying pathogenic variations affecting splicing, the use of antisense oligonucleotides (ASOs) offers immense therapeutic hope. In this study, we develop a framework combining the use of patient-derived cells and kidney organoids to provide evidence of the therapeutic efficacy of ASOs in XLAS patients. Using multiomics analysis, we describe the development of GBM in wild-type and mutated human kidney organoids. We show that GBM maturation is a dynamic process, which requires long organoid culture. Then, using semi-automated quantification of α5(IV) at basement membranes in organoids carrying the splicing variants identified in patients, we demonstrate the efficacy of ASO treatment for α5(IV) restoration. These data contribute to our understanding of the development of GBM and pave the way for a therapeutic screening platform for patients. ### Competing Interest Statement The authors have declared no competing interest. Orphan Kidney Diseases (ORKiD) Agence Nationale de la Recherche, ANR-10-IAHU-01 France 2030 program, DOS0212694

Glomerular nephropathy resulting from the genetic defects in COL4A3/4/5 genes including the classical Alport syndrome is the second most common hereditary kidney disease characterized by persistent haematuria progressing to the need for kidney replacement therapy, frequently associated with sensorineural deafness, and occasionally with ocular anomalies. Diagnosis and management of COL4A3/4/5 glomerulopathy is a great challenge due to its phenotypic heterogeneity, multiple modes of inheritance, variable expressivity, and disease penetrance of individual variants as well as imperfect prognostic and progression factors and scarce and limited clinical trials, especially in children. As a joint initiative of the European Rare Kidney disease reference Network (ERKNet), European Renal Association (ERA Genes&Kidney), and European Society for Paediatric Nephrology (ESPN) Inherited renal disorders working group, a team of experts including adult and paediatric nephrologists, kidney geneticists, audiologists, ophthalmologists, and a kidney pathologist were selected to perform a systematic literature review on 21 clinically relevant PICO (Patient or Population covered, Intervention, Comparator, Outcome) questions. The experts formulated recommendations and formally graded them at a consensus meeting with input from patient representatives and a voting panel of nephrologists representing all regions of the world. Genetic diagnostics comprising joint analysis of COL4A3/4/5 genes is already the key diagnostic test during the initial evaluation of an individual presenting with persistent haematuria, proteinuria, kidney failure of unknown origin, focal segmental sclerosis of unknown origin, and possibly cystic kidney disease. Early renin-angiotensin system blockade is the standard of care therapy; sodium-glucose cotransporter-2 inhibitors may be added in adults with proteinuria and chronic kidney disease. Relatives with heterozygous COL4A3/4/5 variants should only be considered as the last possible resource for living kidney donation. This guideline provides guidance for the diagnosis and management of individuals with pathogenic variants in COL4A3/4/5 genes.

KEY POINTS:Exome sequencing solved 26% of nephronophthisis cases, identifying nephropathy and extrarenal disease genes beyond classic ciliopathy panels. Exome sequencing uncovered GN and tubular nephropathy genes misdiagnosed as ciliopathy-associated nephropathy, underscoring diagnostic overlap in kidney diseases. Patients with nonciliary genetic variants may present with ciliopathy-like extrarenal symptoms, showing phenocopies in kidney ciliopathy diagnostics. BACKGROUND:Nephronophthisis (NPH) is an autosomal recessive tubulointerstitial kidney disease and a leading genetic cause of chronic kidney failure in children and young adults. As a ciliopathy, NPH is caused by biallelic variants in genes encoding proteins involved in the structure and function of primary cilia. The broad clinical spectrum of NPH results in a clinically and genetically heterogeneous disease, posing diagnostic challenges and leaving approximately 30% of cases unresolved with current gene panels. METHODS:After targeted gene panel for ciliopathy-associated genes failed to identify diagnostic variants, exome sequencing (ES) was conducted on 42 unrelated index patients with a clinical diagnosis of NPH, defined as cystic nephropathy progressing to kidney failure within the first two decades of life, or by unspecific CKD accompanied by extrarenal features indicative of a ciliopathy. RESULTS:Pathogenic or likely pathogenic variants were identified in 11 of the 42 patients (26%). Variants were detected in known nephropathy genes ( LAMB2 , COQ8B , COL4A3 , MUC1 ) and a multisystem disease gene with secondary kidney involvement ( AGXT ). In addition, ES elucidated deleterious variants explaining extrarenal phenotypes without corresponding kidney disease in six patients ( APTX , TUBB3 , DHX38 , IQCE, CRX , RPGR ). Variants of unknown significance were identified in three patients, while heterozygous variants in genes associated with recessive disease were observed in three others. A potential candidate gene for syndromic tubulointerstitial nephropathy, SSBP1 , was also identified, suggesting a novel pathway involving mitochondrial dysfunction. CONCLUSIONS:ES enabled the identification of pathogenic variants in known genes associated with kidney diseases, nonkidney conditions, and multisystem disorders with secondary kidney involvement, thereby improving diagnosis accuracy, even in incomplete or atypical cases, and guiding specific diagnostic and therapeutic approaches. The identification of SSBP1 in association with tubulointerstitial nephropathy may provide new insights into the pathogenesis of ciliopathies.

Kidney organoids are an emerging tool for disease modeling, especially genetic diseases. Among these diseases, X-linked Alport syndrome (XLAS) is a hematuric nephropathy affecting the glomerular basement membrane (GBM) secondary to pathogenic variations in the COL4A5 gene encoding the α5 subunit of type IV collagen [α5(IV)]. In patients carrying pathogenic variations affecting splicing, the use of antisense oligonucleotides (ASOs) offers immense therapeutic hope. In this study, we develop a framework combining the use of patient-derived cells and kidney organoids to provide evidence of the therapeutic efficacy of ASOs in XLAS patients. Using multiomics analysis, we describe the development of GBM in WT and mutated human kidney organoids. We show that GBM maturation is a dynamic process, which requires long organoid culture. Then, using semi-automated quantification of α5(IV) at basement membranes in organoids carrying the splicing variants identified in patients, we demonstrate the efficacy of ASO treatment for α5(IV) restoration. These data contribute to our understanding of the development of GBM in kidney organoids and pave the way for a therapeutic screening platform for patients.

Background:The origin of chronic kidney disease (CKD) remains unknown in ≈16% of patients at the time of renal replacement therapy. The aim of this study was to assess the proportion of monogenic kidney diseases in kidney transplant candidates with kidney disease of unknown cause. Methods:Transplant candidates, referred to a nephrogenetic outpatient clinic, had a molecular investigation and were included if they met the following inclusion criteria: absence of diagnosis (including presumed hypertensive nephropathy or vascular or focal segmental glomerulosclerosis lesions) and a glomerular filtration rate <30 ml/min/1.73 m2 before 50 years of age and/or renal morphology abnormality (including renal hypotrophy, cysts and congenital anomalies of the kidney and urinary tract) and/or extrarenal involvement and/or family history of CKD. Results:Eighty-nine patients were evaluated at the nephrogenetic consultation and 84 patients met the inclusion criteria and were tested and included. Half had a family history of CKD. Almost half of the patients (46.4%) had a morphological abnormality of the kidney. Twenty-eight (33.3%) had been biopsied: 21% had focal and segmental hyalinosis lesions and 25% had chronic interstitial nephropathy. Thirty patients (36%) had a positive genetic diagnosis. Of these, 9/30 (30%) had APOL1 high-risk alleles and 21/30 (70%) had monogenic nephropathy. Patients with a positive genetic diagnosis were significantly more likely to have a family history of kidney disease (70% versus 37%; P = .004). Conclusions:Genetic testing enables a diagnosis to be established in 36% of patients, allowing genetic counselling and may help potential living donor evaluations.

Renal tubular dysgenesis (RTD) is a severe kidney disease characterized by poor development of proximal tubules and persistent fetal anuria leading to oligohydramnios. It can be acquired during fetal life or inherited as an autosomal recessive disease associated with bi-allelic pathogenic variants in one of the genes encoding the renin-angiotensin system (RAS) components, AGT, REN, ACE, or AGTR1. Few cases of RTD remain unsolved despite the lack of fetal cause and comprehensive screening of RAS genes. We investigated a case of unsolved RTD with low renin expression by whole genome sequencing, and then screened a series of unsolved RTD by sequencing of a targeted gene panel of genes coding mitochondrial proteins. Oxidative phosphorylation complexes were studied by SDS-PAGE and immunoblotting. We identified a rare homozygous variant in RMND1, a gene known to be responsible for an autosomal recessive mitochondrial disease, in a case presenting with RTD-like phenotype with low renin expression but without identified RAS disease-causing variant. We demonstrate a severe reduction of combined oxidative phosphorylation complexes I and IV subunits in this case. Next, we identified another RMND1 homozygous variant in another unsolved RTD case belonging to a consanguineous family with recurrent fetal demise. Our study shows that biallelic RMND1 pathogenic variants likely cause severe prenatal kidney disease presenting with RTD-like phenotype, and prompts to screen RMND1 in unelucidated severe fetal nephropathies to provide diagnosis and, ultimately, genetic counselling. In addition, these data confirm a still poorly understood link between RMND1-associated mitochondrial dysfunction and renin expression.

INTRODUCTION:Autosomal dominant tubulointerstitial kidney disease (ADTKD) is a common monogenic kidney disease leading to kidney failure usually during mid adulthood. It is due to pathogenic variants in at least five genes. However, despite thorough screening of UMOD, MUC1, REN, HNF1B and SEC61A1, 25 to 50% of families remain without molecular diagnosis. METHODS:Here, we investigated a cohort of 203 families with ADTK, as well as sporadic cases of kidney disease of unknown etiology and cases of chronic kidney disease stage 5 from the Genomics England 100,000 Genomes Project. Expression of JAG1 in kidney and/or urinary epithelial cell (UREC) lines from patients carrying a pathogenic JAG1 variant associated with isolated ADTKD was studied using immunolabelling, Western blotting, targeted RNA-seq and quantitative RT-PCR. Endoplasmic reticulum (ER) stress was tested by analyzing ER protein BiP expression levels in URECs. RESULTS:A pathogenic or likely pathogenic variant in JAG1, the gene associated with Alagille syndrome, was identified in three large families with unsolved ADTKD, and additional rare variants were identified in sporadic cases. In two of the families, the diagnosis of Alagille syndrome was further established in one infant in the fourth or fifth generation; however, none of the 23 adult patients affected with isolated kidney failure (and tubulointerstitial nephritis in individuals with available kidney biopsy) had overt sign of liver, bile duct, heart, eye, or skeletal defect. JAG1 expression studies as well as ER stress analysis suggests that, despite a noteworthy expression of the JAG1-mutated RNAs, the tubulointerstitial renal disease was not due to cell toxicity of an abnormal protein, but rather to haploinsufficiency and loss of function. CONCLUSIONS:JAG1 pathogenic variants can be associated with isolated tubulointerstitial nephropathy which, according to the KDIGO guidelines, should be classified as ADTKD-JAG1 when JAG1 variants lead to isolated chronic kidney disease that fulfills the criteria for ADTKD.

Abstract Background and Aims The human genome includes tandem repeats with variable length (VNTR) and a subset of these repeats have been associated with rare human diseases. Specific frameshift variants in the coding-VNTR of the MUC1 gene cause autosomal dominant tubulointerstitial kidney disease—MUC1 (ADTKD-MUC1). Calling variants from VNTR using short-read sequencing (SRS) is challenging due to poor read mappability, motif complexity (34*60-mer motifs are known up to now), variable repetition, and enormous motif sequence similarity. We have recently developed a computational pipeline called VNtyper, tailored for the precise detection of disease-causing variants within the MUC1 VNTR region using short-read sequencing data. This advancement allowed us to identify overlooked cases in a hereditary renal disease registry, leading to the diagnosis of at least 40 patients. Given that regular exome sequencing with low VNTR coverage proved inefficient for VNtyper, our focus shifted to boost the capture of MUC1 VNTR in exome. This improvement aims to enable the application of VNtyper in ADTKD-MUC1 diagnosis through exome sequencing. Method VNtyper utilizes two independent genotyping algorithms (Kestrel and code-adVNTR) along with MUC1 VNTR-specific reference sequence for the variant detection and is sensitive to the VNTR coverage. We employ Twist custom panels with v1 protocol for target enrichment in both our panel and exome sequencing. This procedure allows for the inclusion of spike-in probes without interfering with other targets. We designed an NTI panel that specifically targets 6 genes associated with ADTKD, including UMOD, MUC1, HNF1B, REN, SEC61A1, and DNAJB11. These genes were captured using a 1x tiling approach, whereas the MUC1 VNTR region was captured with 4x tiling approach. During the exome target enrichment process, we included the NTI probes as a spike-in to boost VNTR coverage. We conducted a routine exome sequencing on a total of 6 samples, consisting of 3 positive controls and 3 negative controls. Additionally, we conducted 16 boosted exome, using 3 negative controls and 13 MUC1 positive samples. Prior to using VNtyper on both exomes, we conducted initial quality controls to verify that there were no alterations in the coverages of other genes. Results The mean coverage of MUC1 VNTR in the regular exome was 72.1x, while in the boosted exome, it exhibited a significant increase to 144x. Adding spike in probes did not have significant effect on the coverage of the contig and there was no alteration in ratio of heterozygosity. As anticipated, in the regular exome, VNtyper was unsuccessful in identifying pathogenic variation in 2 out of 3 true positives. However, all negative controls tested negative. When we used the enhanced exome, we accurately identified all 13 true positive cases, while the negative control cases stayed negative. Through downsampling experiments (reducing read depth from 50% to 5% of the total) on panel data with a mean coverage of 700x, we determined that coverage below 100 should be considered as low coverage for VNTR genotyping using our pipeline. This underscores the critical importance of maintaining adequate coverage for accurate and reliable VNTR genotyping. Conclusion We have achieved successful enhancements in the accuracy and sensitivity of ADTKD diagnosis through the application of VNtyper on clinically boosted exome data. While VNtyper could identify MUC1-positive patients in regular exome data, the effectiveness is influenced by the number of repeats in both alleles and the specific motif in which the variation is present, which significantly decreases the sensitivity. In summary, the VNtyper pipeline demonstrates its ability to detect pathogenic variations in ADTKD-MUC1 on panel data (100% sensitivity), and the improvement in VNTR capture in exome significantly enhances the sensitivity of the ADTKD diagnosis.

Abstract Background and Aims Autosomal dominant tubulointerstitial kidney disease (ADTKD) due to pathogenic variants in the MUC1 gene is difficult to diagnose since these variants lie in a large variable number tandem repeats (VNTR) and require specialized genetic testing, such as SNaPshot minisequencing. We recently developed a computational pipeline, VNtyper, for easier reliable detection of MUC1 VNTR pathogenic variants and have applied this tool to a large cohort of patients with various phenotypes of kidney disease. The aim of this study is to clinically describe patients in whom MUC1 pathogenic variants were unexpectedly detected with the help of VNtyper. Method We applied a computational pipeline, VNtyper, to a large cohort of patients with suspected hereditary kidney disease referred for genetic testing, regardless of their phenotype. This cohort included patients evaluated from 2017 to 2023 in the Molecular Genetics Department of Necker-Enfants Malades Hospital (Paris, France). Clinical characteristics were collected from patients in whom we detected new MUC1 VNTR pathogenic variants. Results 44 out of 3735 patients tested (1.2%) were newly diagnosed with a MUC1 pathogenic variant. Of these patients, ADTKD was clinically suspected in only 31 out of 44 patients (70%). In 2 patients, there was a pathogenic variant in another gene that could explain the phenotype, in addition to the MUC1 pathogenic variant. In fact, one 52-year-old patient had polycystic kidneys, nephrolithiasis, hematuria and normal kidney function. Her mother also had kidney cysts with absence of chronic kidney disease (CKD). This patient had a pathogenic variant in IFT140 gene (c.2399+1G>T) which could explain her autosomal dominant polycystic kidney disease (ADPKD) phenotype. Whether the MUC1 variant contributes to her disease remains unclear, especially seeing as her kidney function is normal at the age of 52 years. On the other hand, one patient had kidney cysts discovered at 3 days of age. A heterozygous HNF1B deletion was detected which could explain the phenotype. The following detection of a pathogenic MUC1 variant was thus an incidental finding and seemed de novo, although parents declined to get tested. These incidental findings can be challenging, especially when it comes to counselling and follow-up. There was no family history of kidney disease in 7 out of 44 patients (16%), and in one patient, we were able to confirm de novo disease since her parents were tested and were negative. It was unfortunately not possible to get DNA from the parents of the other patients. This is the first case of proven de novo ADTKD-MUC1 described to our knowledge. De novo variants in ADTKD-MUC1 are not surprising since the 7C stretch found in VNTR is a hotspot for mutagenesis. Interestingly, these patients seemed to have a more severe clinical presentation with an early onset of disease (median age at diagnosis 28 years [18, 32]). However, this could be due to a selection bias. In fact, young age of onset of CKD may have prompted clinicians to refer these patients for genetic testing despite negative family history, whereas older patients with possible ADTKD-MUC1 and no family history of kidney disease might be presumed to have another etiology of CKD. One way to confirm this would be to test a large cohort of patients with CKD from unknown etiology with no family history of CKD. Considering the nonspecific clinical manifestations of ADTKD-MUC1 and the phenotypic variability, it has previously been suggested that the most important clue to diagnosis is the presence of CKD in first-degree family. The possibility of de novo cases makes the clinical diagnosis of ADKTD-MUC1 even more challenging. Conclusion With VNtyper, we were able to diagnose new cases of ADTKD-MUC1 in a large cohort of patients with various phenotypes, with a significant prevalence of 1.2%. ADTKD-MUC1 was not suspected in 30% of these patients who would have maybe never been diagnosed otherwise. This is also the first study in which we describe a proven de novo case of ADTKD-MUC1. Pathogenic variants in the MUC1 gene should thus be in the differential diagnosis of all unspecified CKD, even if there is no family history since de novo variants are also possible.