End-stage ADPKD with a low-frequency PKD1 mosaic variant accelerated by chemoradiotherapy.

Autosomal dominant polycystic kidney disease (ADPKD) is commonly caused by PKD1, and mosaic PKD1 variants result in milder phenotypes. We present the case of a 32 year-old male with chronic active Epstein-Barr virus who underwent bone marrow transplantation with chemoradiotherapy at age 9. Despite a low-frequency mosaic splicing PKD1 variant, he developed severe renal cysts and end-stage renal disease in his 30 s. This case highlights how environmental factors may contribute to the genetic predisposition to ADPKD.

Comprehensive genetic analysis using next-generation sequencing for the diagnosis of nephronophthisis-related ciliopathies in the Japanese population.

Nephronophthisis is an autosomal-recessive kidney disease that is caused by abnormalities in primary cilia. Nephronophthisis-related ciliopathies (NPHP-RCs) are a common cause of end-stage kidney disease (ESKD) in children and adolescents. NPHP-RCs are often accompanied by extrarenal manifestations, including intellectual disability, retinitis pigmentosa, or polydactyly. Although more than 100 causative genes have been identified, its diagnosis is difficult because the clinical features of each mutation often overlap. From September 2010 to August 2021, we performed genetic analysis, including next-generation sequencing (NGS), in 574 probands with kidney dysfunction and retrospectively studied cases genetically diagnosed with NPHP-RCs. RESULTS: We detected mutations related to NPHP-RCs in 93 patients from 83 families. Members of 60 families were diagnosed using NGS, and the mutations and the corresponding number of families are as follows: NPHP1 (24), NPHP3 (10), OFD1 (7), WDR35 (5), SDCCAG8 (4), BBS10 (3), TMEM67 (3), WDR19 (3), BBS1 (2), BBS2 (2), IFT122 (2), IFT140 (2), IQCB1 (2), MKKS (2), SCLT1 (2), TTC21B (2), ALMS1 (1), ANKS6 (1), BBS4 (1), BBS12 (1), CC2D2A (1), DYNC2H1 (1), IFT172 (1), and MAPKBP1 (1). A total of 39 cases (41.9%) progressed to ESKD at the time of genetic analysis, whereas 58 cases (62.3%) showed extrarenal manifestations, the most common being developmental delay, intellectual disability, and autism spectrum disorder in 44 patients. Comprehensive genetic analysis using NGS is useful for diagnosing patients with NPHP-RCs.

A woman with a dual genetic diagnosis of autosomal dominant tubulointerstitial kidney disease and KBG syndrome.

We present a female patient with a dual genetic diagnosis of autosomal dominant tubulointerstitial kidney disease and KBG syndrome. The proband was an 18-year-old woman presenting with intellectual disability, renal insufficiency, and hyperuricemia. Abdominal ultrasonography did not reveal any abnormalities. The patient's father had been diagnosed with chronic kidney disease and hyperuricemia in his twenties; however, he had no intellectual disability. Her mother and two younger siblings were not affected. Next generation sequencing (NGS) identified mutations in UMOD (c.796T > C) of the proband and her father, and in ANKRD11 (c.1903_1907del) of the proband. Renal insufficiency and intellectual disability were attributed to mutations in UMOD and ANRKD11, respectively. When making genetic diagnoses, the presence of multiple mutations in an individual should be considered, particularly when not all symptoms could be attributed to a single disease. The number of patients with dual genetic diagnosis is expected to increase as NGS becomes more readily available; thus, making it necessary to undertake a careful and robust assessment of the clinical symptoms and the related genotypes, to ensure an accurate diagnosis.

Genotype-phenotype correlations influence the response to angiotensin-targeting drugs in Japanese patients with male X-linked Alport syndrome.

Early kidney failure in the hereditary type IV collagen disease, Alport syndrome, can be delayed by renin-angiotensin inhibitors. However, whether all patients and all different genotypes respond equally well to this kidney-protective therapy remains unclear. Here, we performed a retrospective study on 430 patients with male X-linked Alport syndrome to examine the relationships among kidney prognosis, genotype, and treatment effect in a large cohort of Japanese patients. We analyzed the clinical features, genotype-phenotype correlation, and kidney survival period for patients treated with or without renin-angiotensin inhibitors. As a result, the median kidney survival period of patients in this cohort was found to be at 35 years with a strong genotype-phenotype correlation. The median age at the onset of end stage kidney disease (ESKD) significantly differed between patients treated with and without renin-angiotensin inhibitors (over 50 years versus 28 years, respectively). Moreover, these drugs delayed the onset of ESKD in patients with truncating variants for 12 years, extending the median age from 16 years to 28 years. Thus, our results confirmed a strong genotype-phenotype correlation in patients with male X-linked Alport syndrome. Additionally, it was suggested that renin-angiotensin inhibitors could significantly delay ESKD progression. Despite these therapies, patients with truncating variants developed ESKD at the median age of 28 years.

Development of an exon skipping therapy for X-linked Alport syndrome with truncating variants in COL4A5.

Currently, there are no treatments for Alport syndrome, which is the second most commonly inherited kidney disease. Here we report the development of an exon-skipping therapy using an antisense-oligonucleotide (ASO) for severe male X-linked Alport syndrome (XLAS). We targeted truncating variants in exon 21 of the COL4A5 gene and conducted a type IV collagen α3/α4/α5 chain triple helix formation assay, and in vitro and in vivo treatment efficacy evaluation. We show that exon skipping enabled trimer formation, leading to remarkable clinical and pathological improvements including expression of the α5 chain on glomerular and the tubular basement membrane. In addition, the survival period was clearly prolonged in the ASO treated mice group. This data suggests that exon skipping may represent a promising therapeutic approach for treating severe male XLAS cases.

Bardet-Biedl syndrome in two unrelated patients with identical compound heterozygous SCLT1 mutations.

Bardet-Biedl syndrome (BBS) is a rare autosomal recessive ciliopathy characterized by retinitis pigmentosa (RP), truncal obesity, cognitive impairment, hypogonadism in men, polydactyly, and renal abnormalities with severe renal dysfunction. Twenty-two causative genes have already been reported for this disorder. In this study, we identified two unrelated Japanese patients with clinical diagnoses of BBS associated with compound heterozygous SCLT1 mutation. Patient 1 was a 10-year-old girl, and patient 2 was a 22-year-old man. Both the patients showed severe renal dysfunction in childhood, RP, mild intellectual disability, short stature, and truncal obesity, without oral aberrations and polydactyly. Patient 2 also had hypogonadism. We identified two missense variants in SCLT1, c.[1218G > A] and [1631A > G], in both the patients by next-generation sequencing. Subsequent cDNA analysis revealed that c.1218G > A affected exon 14 skipping in SCLT1. To date, SCLT1 has been reported as the causative gene of oral-facial-digital syndrome type IX, and Senior-Løken syndrome. The phenotypes of both the present patients were compatible with BBS. These results highlight SCLT1 as an additional candidate for BBS phenotype in an autosomal recessive manner.

A girl with CLOVES syndrome with a recurrent PIK3CA somatic mutation and pancreatic steatosis.

CLOVES syndrome is characterized by congenital lipomatous overgrowth, vascular malformation, epidermal nevi, and scoliosis/spinal malformation. It is caused by somatic mosaicism of gain-of-function variants of PIK3CA. Here, we describe a novel case of a 5-year-old Japanese girl with CLOVES and concurrent pancreatic steatosis. She had a recurrent somatic mutation in PIK3CA (NM_006218.3: c.1357G>A, p.Glu453Lys), elevated HbA1c levels, and pancreatic steatosis. This case indicates that pancreatic screening is critical for PIK3CA-related disorders.

Clinical spectrum of male patients with OFD1 mutations.

Oral-facial-digital syndrome type 1 (OFD1) is a ciliopathy characterized by oral, facial, and digital malformations that are often accompanied by polycystic lesion of the kidney and central nervous involvement. OFD1 shows an X-linked recessive inheritance caused by mutation in the OFD1 gene (Xp22.2). The disease is generally considered embryonic lethal for hemizygous males. However, males with OFD1 mutations were recently reported. Here, we report four additional Japanese male patients with OFD1 variants and describe the variable clinical manifestation and disease severity among the four patients. Patient 1 with pathogenic indels including a 19-bp deletion and 4-bp insertion (c.2600-18_2600delinsACCT) had end-stage renal disease (ESRD) with bilateral cystic kidneys and sensory hearing loss. He showed neither intellectual disability nor facial or digital dysmorphism. Patient 2 with a missense variant in exon 7 (c.539 A > T, p.Asp180Val) presented head circumference enlargement, brachydactyly, high-arched palate, micropenis, severe global developmental delay, and ESRD. Patient 3 had a single base substitution at the splice donor site of intron 16 (c.2260 + 2 T > G) causing a 513-bp deletion at the transcript level. The patient had chronic kidney disease and speech delay, but no oral, facial, or digital dysmorphism. His uncle (patient 4) carried the same OFD1 variant and showed ESRD with extra-renal malformations including obesity and micropenis, which was previously diagnosed as Bardet-Biedl syndrome. The OFD1 mutations were not lethal in these four male patients, likely because the three mutations were in-frame or missense. This report provided insights into the onset mechanism and phenotype-genotype association in patients with OFD1 mutations.

Detection of Splicing Abnormalities and Genotype-Phenotype Correlation in X-linked Alport Syndrome.

BACKGROUND: X-linked Alport syndrome (XLAS) is a progressive hereditary nephropathy caused by mutations in the COL4A5 gene. Genotype-phenotype correlation in male XLAS is relatively well established; relative to truncating mutations, nontruncating mutations exhibit milder phenotypes. However, transcript comparison between XLAS cases with splicing abnormalities that result in a premature stop codon and those with nontruncating splicing abnormalities has not been reported, mainly because transcript analysis is not routinely conducted in patients with XLAS. METHODS: We examined transcript expression for all patients with suspected splicing abnormalities who were treated at one hospital between January of 2006 and July of 2017. Additionally, we recruited 46 males from 29 families with splicing abnormalities to examine genotype-phenotype correlation in patients with truncating (n=21, from 14 families) and nontruncating (n=25, from 15 families) mutations at the transcript level. RESULTS: We detected 41 XLAS families with abnormal splicing patterns and described novel XLAS atypical splicing patterns (n=14) other than exon skipping caused by point mutations in the splice consensus sequence. The median age for developing ESRD was 20 years (95% confidence interval, 14 to 23 years) among patients with truncating mutations and 29 years (95% confidence interval, 25 to 40 years) among patients with nontruncating mutations (P=0.001). CONCLUSIONS: We report unpredictable atypical splicing in the COL4A5 gene in male patients with XLAS and reveal that renal prognosis differs significantly for patients with truncating versus nontruncating splicing abnormalities. Our results suggest that splicing modulation should be explored as a therapy for XLAS with truncating mutations.

Clinically diverse phenotypes and genotypes of patients with branchio-oto-renal syndrome.

Branchio-oto-renal (BOR) syndrome is a rare autosomal dominant disorder characterized by branchiogenic anomalies, hearing loss, and renal anomalies. The aim of this study was to reveal the clinical phenotypes and their causative genes in Japanese BOR patients. Patients clinically diagnosed with BOR syndrome were analyzed by direct sequencing, multiplex ligation-dependent probe amplification (MLPA), array-based comparative genomic hybridization (aCGH), and next-generation sequencing (NGS). We identified the causative genes in 38/51 patients from 26/36 families; EYA1 aberrations were identified in 22 families, SALL1 mutations were identified in two families, and SIX1 mutations and a 22q partial tetrasomy were identified in one family each. All patients identified with causative genes suffered from hearing loss. Second branchial arch anomalies, including a cervical fistula or cyst, preauricular pits, and renal anomalies, were frequently identified (>60%) in patients with EYA1 aberrations. Renal hypodysplasia or unknown-cause renal insufficiency was identified in more than half of patients with EYA1 aberrations. Even within the same family, renal phenotypes often varied substantially. In addition to direct sequencing, MLPA and NGS were useful for the genetic analysis of BOR patients.

Detection of copy number variations by pair analysis using next-generation sequencing data in inherited kidney diseases.

BACKGROUND: Comprehensive genetic approaches for diagnosing inherited kidney diseases using next-generation sequencing (NGS) have recently been established. However, even with these approaches, we are still failing to detect gene defects in some patients who appear to suffer from genetic diseases. One of the reasons for this is the difficulty of detecting copy number variations (CNVs) using our current approaches. For such cases, we can apply methods of array-based comparative genomic hybridization (aCGH) or multiplex ligation and probe amplification (MLPA); however, these are expensive and laborious and also often fail to identify CNVs. Here, we report seven cases with CNVs in various inherited kidney diseases screened by NGS pair analysis. METHODS: Targeted sequencing analysis for causative genes was conducted for cases with suspected inherited kidney diseases, for some of which a definitive genetic diagnosis had not been achieved. We conducted pair analysis using NGS data for those cases. When CNVs were detected by pair analysis, they were confirmed by aCGH and/or MLPA. RESULTS: In seven cases, CNVs in various causative genes of inherited kidney diseases were detected by pair analysis. With aCGH and/or MLPA, pathogenic CNV variants were confirmed: COL4A5 or HNF1B in two cases each, and EYA1, CLCNKB, or PAX2 in one each. CONCLUSION: We presented seven cases with CNVs in various genes that were screened by pair analysis. The NGS-based CNV detection method is useful for comprehensive screening of CNVs, and our results revealed that, for a certain proportion of cases, CNV analysis is necessary for accurate genetic diagnosis.

The smallest de novo 20q11.2 microdeletion causing intellectual disability and dysmorphic features.

The 20q11.2 microdeletion is a rare chromosomal aberration characterized by intellectual disability (ID), motor developmental delay, neonatal feeding problems, and facial dysmorphism. Here, a 2-year- and 6-month-old Japanese girl with a 1.2 Mb microdeletion of 20q11.2 showed ID, motor developmental delay, and distinctive facial features without feeding problems. The deleted region was identified by array-based comparative genomic hybridization and is the smallest reported for a 20q11.2 microdeletion.

Natural History and Genotype-Phenotype Correlation in Female X-Linked Alport Syndrome.

INTRODUCTION: X-linked Alport syndrome (XLAS) is a hereditary disease characterized by progressive nephritis, hearing loss, and ocular abnormalities. Affected male patients usually progress to end-stage renal disease in early or middle adulthood, and disease severity is strongly correlated with genotype. However, the clinical course in female patients has rarely been reported. METHODS: We conducted a retrospective analysis of females with genetically proven XLAS (n = 275) and their affected female family members (n = 61) from 179 Japanese families. Patients suspected to have Alport syndrome from pathologic findings or a family history who were referred from anywhere in Japan for genetic diagnosis between 2006-2015 were included in this study. Clinical and laboratory data were collected from medical records at the time of registration for genetic analysis. RESULTS: Proteinuria was detected in 175 genetically proven patients (72.6%), and the median age for developing proteinuria was 7.0 years. Fifty-two of 336 patients developed end-stage renal disease with a median renal survival age of 65.0 years. No obvious genotype-phenotype correlation was observed. Additionally, targeted sequencing for podocyte-related genes in patients with severe phenotypes revealed no obvious variants considered to be modifier genes except for 1 patient with a COL4A3 gene variant. DISCUSSION: This study revealed that phenotypes in female XLAS patients may be severe, but genotype does not help to predict the disease severity. Clinicians must therefore pay careful attention to the clinical course and appropriate treatment in females with XLAS.

Genetic, Clinical, and Pathologic Backgrounds of Patients with Autosomal Dominant Alport Syndrome.

BACKGROUND AND OBJECTIVES: Alport syndrome comprises a group of inherited heterogeneous disorders involving CKD, hearing loss, and ocular abnormalities. Autosomal dominant Alport syndrome caused by heterozygous mutations in collagen 4A3 and/or collagen 4A4 accounts for <5% of patients. However, the clinical, genetic, and pathologic backgrounds of patients with autosomal dominant Alport syndrome remain unclear. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: We conducted a retrospective analysis of 25 patients with genetically proven autosomal dominant Alport syndrome and their family members (a total of 72 patients) from 16 unrelated families. Patients with suspected Alport syndrome after pathologic examination who were referred from anywhere in Japan for genetic analysis from 2006 to 2015 were included in this study. Clinical, laboratory, and pathologic data were collected from medical records at the point of registration for genetic diagnosis. Genetic analysis was performed by targeted resequencing of 27 podocyte-related genes, including Alport-related collagen genes, to make a diagnosis of autosomal dominant Alport syndrome and identify modifier genes or double mutations. Clinical data were obtained from medical records. RESULTS: The median renal survival time was 70 years, and the median age at first detection of proteinuria was 17 years old. There was one patient with hearing loss and one patient with ocular lesion. Among 16 patients who underwent kidney biopsy, three showed FSGS, and seven showed thinning without lamellation of the glomerular basement membrane. Five of 13 detected mutations were reported to be causative mutations for autosomal recessive Alport syndrome in previous studies. Two families possessed double mutations in both collagen 4A3 and collagen 4A4, but no modifier genes were detected among the other podocyte-related genes. CONCLUSIONS: The renal phenotype of autosomal dominant Alport syndrome was much milder than that of autosomal recessive Alport syndrome or X-linked Alport syndrome in men. It may, thus, be difficult to make an accurate diagnosis of autosomal dominant Alport syndrome on the basis of clinical or pathologic findings. No modifier genes were identified among the known podocyte-related genes.