Coexisting presentation of two rare genetic variants of autosomal dominant polycystic kidney disease and Alport syndrome.

Alport syndrome and autosomal dominant polycystic kidney disease are monogenic causes of chronic kidney disease and end-stage kidney failure. We present a case of a man in his 60s with progressive chronic kidney disease, bilateral sensorineural hearing loss and multiple renal cysts. Genetic analysis revealed a heterozygous variant in COL4A3 (linked to Alport syndrome) and in the GANAB gene (associated with a milder form of autosomal dominant polycystic kidney disease). Although each variant confers a mild risk of developing end-stage kidney disease, the patient presented a pronounced and accelerated progression of chronic kidney disease, which goes beyond what would be predicted by adding up their individual effects. This suggests a potential synergic effect of both variants, which warrants further investigation.

[Precision diagnosis and therapeutic intervention of Alport syndrome].

Alport syndrome is one of the most common inherited kidney diseases caused by mutations in the type Ⅳ collagen genes. It has a complex pattern of inheritance and diverse clinical manifestations, and severe cases will rapidly progress to end-stage kidney disease. With the rapid development of genetic testing technology, there is a deeper understanding of the genetic spectrum of Alport syndrome, the effectiveness of clinical therapies, and the prediction of disease prognosis. Therefore, the purpose of the article is to introduce the advances in the diagnosis and treatment of Alport syndrome, aiming to improve the early diagnosis and standardized treatment of this disease.

Alport syndrome and Alport kidney diseases - elucidating the disease spectrum.

PURPOSE OF REVIEW: With the latest classification, variants in three collagen IV genes, COL4A3 , COL4A4 , and COL4A5 , represent the most prevalent genetic kidney disease in humans, exhibiting diverse, complex, and inconsistent clinical manifestations. This review breaks down the disease spectrum and genotype-phenotype correlations of kidney diseases linked to genetic variants in these genes and distinguishes "classic" Alport syndrome (AS) from the less severe nonsyndromic genetically related nephropathies that we suggest be called "Alport kidney diseases". RECENT FINDINGS: Several research studies have focused on the genotype-phenotype correlation under the latest classification scheme of AS. The historic diagnoses of "benign familial hematuria" and "thin basement membrane nephropathy" linked to heterozygous variants in COL4A3 or COL4A4 are suggested to be obsolete, but instead classified as autosomal AS by recent expert consensus due to a significant risk of disease progression. SUMMARY: The concept of Alport kidney disease extends beyond classic AS. Patients carrying pathogenic variants in any one of the COL4A3/A4/A5 genes can have variable phenotypes ranging from completely normal/clinically unrecognizable, hematuria without or with proteinuria, or progression to chronic kidney disease and kidney failure, depending on sex, genotype, and interplays of other genetic as well as environmental factors.

Clinicopathological Features of Hereditary Nephritis in the Iranian Population: Analysis of a 14-Year Survey in Kidney Biopsies From a Large Referral Center.

BACKGROUND: Hereditary nephritis (HN), including Alport syndrome (AS) and thin basement membrane nephropathy (TBMN), is a rare genetic cause of hematuria. A definitive diagnosis requires electron microscopy (EM). Therefore, the clinical characteristics of these conditions are less known. This study aimed to determine the percentage and clinicopathological features of HN in patients from a referral center in Iran. METHODS: We checked kidney biopsy reports from 2007 to 2021 and extracted cases with HN. Fresh specimens of the cases diagnosed in the last two years were stained by immunofluorescence (IF) for collagen type IV alpha chains. EM findings in these cases were re-evaluated and categorized as diffuse glomerular basement membrane (GBM) thinning, definite, and suspicious features of AS. RESULTS: We analyzed 3884 pathology reports of kidney biopsies from 2007 to 2021 and identified 210 patients (5.4%) with HN, with a mean age of 13.78±12.42 years old. Hematuria with proteinuria (53.3%), isolated hematuria (44.2%), and proteinuria with hematuria and increased creatinine (2.5%) were found in these patients. The re-evaluation of EM findings revealed GBM thinning, definite, and suspicious findings of AS in 37.5%, 43.8%, and 18.8% cases, respectively. The most common diagnosis in 32 cases after the IF study was X-linked AS (71.9%), and 6.2% of cases were autosomal recessive AS. TBMN and autosomal dominant AS remained the differential diagnoses in 21.9%. CONCLUSION: It was found that EM is helpful for the primary diagnosis of patients with definite AS. Immunostaining improves the diagnostic sensitivity for the differentiation of those with suspicious EM findings and determines the inheritance pattern. However, a multidisciplinary approach for a subset of cases is required for the best diagnosis and management.

Genetic diagnosis of Alport syndrome in 16 Chinese families.

BACKGROUND: Alport syndrome (AS) is a genetically heterogeneous disorder resulting from mutations in the collagen IV genes COL4A3, COL4A4, and COL4A5. The genetic diagnosis of AS is very important to make precise diagnosis and achieve optimal outcomes. METHODS: In this study, 16 Chinese families with suspected AS were recruited after pedigree analysis, and the clinical presentations were analyzed by a nephrologist. The genetic diagnosis was performed by whole-exome sequencing (WES) and the disease-causing variants were confirmed by Sanger sequencing. RESULTS: The cohort of probands included seven men and nine women, with a mean age of 19.9 years. Pathological analysis showed slight-to-moderate mesangial proliferation, and thin basement membrane was the main findings. Pathogenic variants were revealed by WES in each family, and the co-segregation with renal presentation was confirmed by PCR. In addition, RT-PCR analysis showed that the intronic variant led to aberrant splicing. CONCLUSION: Our findings expand the spectrum of AS gene variation, which will inform genetic diagnosis and add to the theoretical basis for the prevention of AS.

Quantitative assessment of glomerular basement membrane collagen IV α chains in paraffin sections from patients with focal segmental glomerulosclerosis and Alport gene variants.

Focal segmental glomerulosclerosis (FSGS) lesions have been linked to variants in COL4A3/A4/A5 genes, which are also mutated in Alport syndrome. Although it could be useful for diagnosis, quantitative evaluation of glomerular basement membrane (GBM) type IV collagen (colIV) networks is not widely used to assess these patients. To do so, we developed immunofluorescence imaging for collagen α5(IV) and α1/2(IV) on kidney paraffin sections with Airyscan confocal microscopy that clearly distinguishes GBM collagen α3α4α5(IV) and α1α1α2(IV) as two distinct layers, allowing quantitative assessment of both colIV networks. The ratios of collagen α5(IV):α1/2(IV) mean fluorescence intensities (α5:α1/2 intensity ratios) and thicknesses (α5:α1/2 thickness ratios) were calculated to represent the levels of collagen α3α4α5(IV) relative to α1α1α2(IV). The α5:α1/2 intensity and thickness ratios were comparable across all 11 control samples, while both ratios were significantly and markedly decreased in all patients with pathogenic or likely pathogenic Alport COL4A variants, supporting validity of this approach. Thus, with further validation of this technique, quantitative measurement of GBM colIV subtype abundance by immunofluorescence, may potentially serve to identify the subgroup of patients with FSGS lesions likely to harbor pathogenic COL4A variants who could benefit from genetic testing.

Multidisciplinary management improves the genetic diagnosis of hereditary kidney diseases in the next generation sequencing (NGS) era.

BACKGROUND AND OBJECTIVE: Hereditary kidney diseases (HKD) are a frequent cause of chronic kidney disease, and their diagnosis has increased since the introduction of next generation sequencing (NGS). In 2018, the Multidisciplinary Unit for Hereditary Kidney Diseases of the Region of Murcia (UMERH-RM) was founded based on the genetic study of HKD. The objective of this study is to analyze the results obtained in the first 3 years of operation, and to analyze the clinical factors associated to a final genetic diagnosis. MATERIALS AND METHODS: All the patients studied with the HKD gene panel were included. The characteristics between those who obtained a final genetic diagnosis and those who did not were compared. RESULTS: A total of 360 patients were studied, detecting genetic variants in 164 not related patients (45.6%). 45 of these were variants of uncertain significance requiring a family co-segregation study, which was facilitated by the multidisciplinary unit. Overall, considering the results obtained with the NGS panel and the extended genomic studies, a final diagnostic yield of HRD of 33.3% (120/360) was achieved, and including incidental findings 35.6% (128/360). Two hundred and twenty-three patients with suspected Alport syndrome were studied. Diagnosis was confirmed in 28.5% (COL4A4 most frequent gene), more frequently women with an obvious compatible family history. They also had frequently microhematuria, although 5 patients without microhematuria confirmed the diagnosis. There were no differences in age, proteinuria, renal function, hearing loss, or ophthalmologic abnormalities. The most frequent finding in the renal biopsy was mesangial proliferation. We estimate that 39 patients avoided renal biopsy. A total of 101 patients with suspected PKD were also studied, 49.5% had a conclusive genetic result (most frequent gene PKD1), more frequently women, with larger kidney sizes (although 9 patients with normal kidney size confirmed diagnosis). Again, the most predictive characteristic of genetic outcome was family history. CONCLUSIONS: The implementation of an NGS panel for HKD, together with the multidisciplinary approach to cases, has improved the diagnostic performance of HKD. In our sample, autosomal dominant Alport syndrome is of highest incidence. Ophthalmological and auditory examinations did not contribute to the diagnosis. We have seen a significant decrease in the indication of renal biopsies thanks to molecular diagnosis. The multidisciplinary approach, with the active participation of nephrologists, paediatricians, clinical and molecular geneticists, with insistence on adequate patient phenotyping and review of their family history, offers a better interpretation of genetic variants, allowing reclassification of the diagnosis of some nephropathies, thus improving their management and genetic advice.

Is there a dominant-negative effect in individuals with heterozygous disease-causing variants in COL4A3/COL4A4?

Alport syndrome (AS) shows a broad phenotypic spectrum ranging from isolated microscopic hematuria (MH) to end-stage kidney disease (ESKD). Monoallelic disease-causing variants in COL4A3/COL4A4 have been associated with autosomal dominant AS (ADAS) and biallelic variants with autosomal recessive AS (ARAS). The aim of this study was to analyze clinical and genetic data regarding a possible genotype-phenotype correlation in individuals with disease-causing variants in COL4A3/COL4A4. Eighty-nine individuals carrying at least one COL4A3/COL4A4 variant classified as (likely) pathogenic according to the American College of Medical Genetics guidelines and current amendments were recruited. Clinical data concerning the prevalence and age of first reported manifestation of MH, proteinuria, ESKD, and extrarenal manifestations were collected. Individuals with monoallelic non-truncating variants reported a significantly higher prevalence and earlier diagnosis of MH and proteinuria than individuals with monoallelic truncating variants. Individuals with biallelic variants were more severely affected than those with monoallelic variants. Those with biallelic truncating variants were more severely affected than those with compound heterozygous non-truncating/truncating variants or individuals with biallelic non-truncating variants. In this study an association of heterozygous non-truncating COL4A3/COL4A4 variants with a more severe phenotype in comparison to truncating variants could be shown indicating a potential dominant-negative effect as an explanation for this observation. The results for individuals with ARAS support the, still scarce, data in the literature.

Alport syndrome: A puzzling case of thrombocytopenia and giant unusual platelets in blood smear.

A puzzling case of thrombocytopenia and giant unusual platelets in blood smear reveals a past diagnosis of Alport syndrome in 44-year-old woman with end-stage renal disease and abnormal CBC values.

A case report and literature study on Alport syndrome featuring nephrotic syndrome as its primary manifestation.

BACKGROUND: Historically, due to the lack of distinct clinical symptoms, Alport syndrome, a hereditary kidney disease prevalent in children and a leading cause of kidney failure, has often been misdiagnosed as other kidney conditions. CASE DESCRIPTION: This article presents a comprehensive review and analysis of clinical data concerning a child diagnosed with Alport syndrome, where nephrotic syndrome served as the primary manifestation. The male child in this case exhibited symptoms starting at the age of 6, initially diagnosed as nephrotic syndrome. Consequently, oral steroid medication was administered, proving ineffective. Due to persistent proteinuria and microscopic hematuria, a renal biopsy was performed. Immunofluorescence staining revealed no abnormal expression of the α3, α4, and α5 chains of type IV collagen. Notably, electron microscopy revealed the basement membrane to be partially torn and arachnoid. Genetic testing indicated a hemizygous COL4A5 acceptor-splice-site mutation c.4707-1(IVS50)G > A, inherited from his mother. CONCLUSION: This specific mutated locus, being the first of its kind reported, adds valuable information to the existing gene mutation spectrum of Alport syndrome. Consequently, it emphasizes the importance for clinicians to deepen their understanding of rare kidney diseases, contributing to enhanced diagnostic accuracy and improved patient care.

[Genetic testing and prenatal diagnosis for two Chinese pedigrees affected with Alport syndrome due to variants of COL4A5 gene].

OBJECTIVE: To analysis variants of COL4A5 gene in two Chinese pedigrees affected with Alport syndrome (AS) and provide prenatal diagnosis for them. METHODS: Two unrelated ethnic Han Chinese pedigrees who had visited the First Affiliated Hospital of Zhengzhou University respectively in September 2018 and January 2020 were selected as the study subjects. Clinical data were collected, and genomic DNA was extracted from peripheral venous blood and amniotic fluid samples for genetic testing. Following next generation sequencing, candidate variants of the COL4A5 gene were verified by Sanger sequencing and bioinformatic analysis. The gender of the fetuses was determined by the presence of sex-determining region on Y (SRY). RESULTS: Genetic testing revealed that the proband and a fetus from pedigree 1 had both harbored a c.2723G>A (p.Gly908Glu) variant in exon 32 of the COL4A5 gene, whilst the proband and a fetus from pedigree 2 had both harbored a c.3817G>A (p.Gly1273Asp) variant in exon 44 of the COL4A5 gene. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), both variants were classified as likely pathogenic (PP2+PM2_Supporting). Following exclusion of maternal contamination, PCR amplification of the SRY region indicated that both fetuses were males. CONCLUSION: The c.2723G>A (p.Gly908Glu) and c.3817G>A (p.Gly1273Asp) variants of the COL4A5 gene probably underlay the AS in the two pedigrees. Detection of the SRY region can reliably identify the fetal sex, which is conducive to the prenatal diagnosis. Above results have also enriched the mutational spectrum of the COL4A5 gene and provided a reference for correlating the genotype and phenotype of the AS.

A comparison of the ocular features in Pierson and Alport syndrome: a case report and literature review.

BACKGROUND: Pierson syndrome and X-linked Alport syndrome result from pathogenic variants in LAMB2 and COL4A5, respectively, and both affect basement membranes in the kidney and the eye. This study describes the ocular features in an individual with a homozygous LAMB2 pathogenic variant and compares the reported abnormalities in Pierson syndrome with those in Alport syndrome. METHODS: A 28-year-old man who developed kidney failure 10 years previously and subsequently had an atrial septal defect repair was suspected of having genetic kidney disease on the basis of his likely diagnosis of Focal and Segmental Glomerulosclerosis (FSGS), his young age at presentation, and his cardiac anomaly. He then underwent Whole Exome Sequencing and a formal ophthalmological examination. RESULTS: The patient was found to have a homozygous Likely Pathogenic missense variant (p.(Arg1719Cys)) in LAMB2 consistent with the diagnosis of Pierson syndrome. He had normal visual acuity, normal optic globe and cornea size, and normal lens appearance on direct examination. Upon further testing, his cornea demonstrated central thinning. There was also increased corneal endothelial pleomorphism, a reduced foveal reflex, and a blunted foveal curvature, similar to the features seen in X-linked Alport syndrome. CONCLUSION: Our patient had a later onset form of Pierson syndrome or "FSGS type 5, with or without ocular abnormalities," consistent with his "milder" LAMB2 missense variant. The resemblance of the ocular features in Pierson syndrome and X-linked Alport syndrome suggests that mutations in LAMB2 and COL4A5 have similar effects on basement membranes and the pathogenesis of ocular damage.

Investigation of the current situation regarding diagnosis and treatment of Alport syndrome in Asian countries: results of survey of the Asian Paediatric Nephrology association (AsPNA) tubular and inherited working group.

BACKGROUND: Alport syndrome is one of the most common inherited kidney diseases worldwide. A genetic test or kidney biopsy is necessary for a definite diagnosis of this disease, and an accurate diagnosis system for this disease is highly desired in each country. However, the current situation in Asian countries is not clear. Therefore, the tubular and inherited disease working group of the Asian Pediatric Nephrology Association (AsPNA) aimed to assess the current situation of diagnosis and treatment for Alport syndrome in Asia. METHODS: The group conducted an online survey among the members of AsPNA in 2021-2022. Collected data included the number of patients for each inheritance mode, availability of gene tests or kidney biopsy, and treatment strategies for Alport syndrome. RESULTS: A total of 165 pediatric nephrologists from 22 countries in Asia participated. Gene test was available in 129 institutes (78%), but the cost was still expensive in most countries. Kidney biopsy was available in 87 institutes (53%); however, only 70 can access electron microscopy, and 42 can conduct type IV collagen α5 chain staining. Regarding treatment, 140 centers use renin-angiotensin system (RAS) inhibitors (85%) for Alport syndrome patients. CONCLUSIONS: This study result might suggest that the system is underdeveloped enough to diagnose all Alport syndrome patients in most Asian countries. However, once diagnosed with Alport syndrome, most of them were treated with RAS inhibitors. These survey results can be used to address knowledge, diagnostic system, and treatment strategy gaps and improve the Alport patients' outcomes in Asian countries.

Electroretinographic abnormalities in Alport syndrome with a novel COL4A5 truncated variant (p.Try20GlyfsTer19).

PURPOSE: Alport syndrome comprises a heterogeneous group of inherited kidney diseases that are associated with ocular complications. In this study, we aimed to detail the clinical characteristics of a patient with X-linked Alport syndrome. METHODS: We performed next-generation sequencing (NGS) with hybridization capture to identify the disease-causing variant of Alport syndrome and a comprehensive ophthalmic examination, including full-field electroretinography (FF-ERG). RESULTS: Genetic testing using NGS with hybridization capture revealed a novel hemizygous variant [c.51_52delGA (p.Trp20GlyfsTer19)] in exon 1 of COL4A5. The patient underwent cataract surgery in both eyes because of decreased visual acuity and photophobia. The best-corrected visual acuity improved from 0.9 and 0.7 in the right and left eyes, respectively, to 1.5 in both eyes. Anterior-segment optical coherence tomography (OCT) revealed anterior and posterior lenticonus. Fundus photographs showed central and peripheral fleck retinopathy. Wide-field fundus autofluorescence (AF) imaging showed mottled hyper- and hypo-AF in the peripheral retina, which was consistent with peripheral fleck retinopathy. Furthermore, OCT revealed thinning of the inner retinal layers, especially at the temporal macular, but the outer retinal layers were preserved. Ganglion cell analysis showed no progression for 5 years. FF-ERG was performed at 41 (phakia) and 46 (pseudophakia) years of age. The amplitudes of dark-adapted (DA) and light-adapted (LA) responses showed selective b-wave abnormalities. The b/a-wave ratios of DA 3.0 were 1.22 and 1.16 in the right and left eyes, respectively. The amplitudes of DA 3.0 oscillatory potentials (OP) were reduced. Five years later, the amplitudes of DA and LA responses revealed no remarkable changes, except for an OP wave of DA 3.0, which was substantially reduced. CONCLUSIONS: Our findings revealed electroretinographic abnormalities in a patient with Alport syndrome, which predominantly indicated impairment of the inner retina. Notably, little short-term progression was observed.

Ratio of Urinary Proteins to Albumin Excretion Shifts Substantially during Progression of the Podocytopathy Alport Syndrome, and Spot Urine Is a Reliable Method to Detect These Pathologic Changes.

The urinary albumin- and protein-to-creatinine ratios (UACR and UPCR, respectively) are key endpoints in most clinical trials assessing risk of progression of chronic kidney disease (CKD). For the first time, the current study compares the UACR versus the UPCR head-to-head at early stages of CKD, taking use of the hereditary podocytopathy Alport syndrome (AS) as a model disease for any CKD. Urine samples originated from the prospective randomized, controlled EARLY PRO-TECT Alport trial (NCT01485978). Urine samples from 47 children with confirmed diagnoses of AS at very early stages of CKD were divided according to the current stage of AS: stage 0 (UACR < 30 mg/g), stage 1 (30-300 mg/g) or stage 2 (>300 mg/g). The range of estimated glomerular filtration rate was 75-187.6 mL/min. The mean age was 10.4 ± 4.5 years. In children at stage 0, proteinuria in spot urine, confirmed in 24 h urine, was almost ten times higher than albuminuria (106.4 ± 42.2 vs. 12.5 ± 9.7; p < 0.05); it was "only" about three times higher in stage 1 (328.5 ± 210.1 vs. 132.3 ± 80.5; p < 0.05) and almost equal in stage 2 (1481.9 ± 983.4 vs. 1109.7 ± 873.6; p = 0.36). In 17 children, UACRs and UPCRs were measured simultaneously in 24 h urine and spot urine in the same study visit. Interestingly, the UACR (and UPCR) in 24 h urine vs. in spot urine varied by less than 10% (266.8 ± 426.4 vs. 291.2 ± 530.2). In conclusion, our study provides the first evidence that in patients with normal glomerular filtration rate (GFR) and low amounts of albuminuria, especially in children with podocytopathies such as AS, measuring the UACR and UPCR in spot urine is a reliable and convenient alternative to 24 h urine collection. Our study advocates both the UACR and the UPCR as relevant diagnostic biomarkers in future clinical trials in children with glomerular diseases because the UPCR seems to be a very significant parameter at very early stages of podocytopathies. The German Federal Ministry of Education and Research funded this trial (01KG1104).

[Expert consensus on the diagnosis and treatment of Alport syndrome (version 2023)].

The "Expert Recommendations on the Diagnosis and Treatment of Alport Syndrome" published in 2018 have greatly promoted the standardized management of Alport syndrome in China. In recent years, the rapid advances in researches related to this disorder have provided new insights into the clinical practice of Alport syndrome. To this end, based on the latest research progress at home and abroad, the Alport Syndrome Collaborative Group, the National Clinical Research Center of Kidney Diseases of Jinling Hospital and the Rare Diseases Branch of Beijing Medical Association have jointly organized the experts in relevant fields to revise the 2018 version of the recommendations. The updated version adds new contents related to genetic testing and variant interpretation, and refines the strategies for diagnosis, treatment and follow-up management, thereby providing guidance for the clinical diagnosis and treatment of Alport syndrome.

A wave of deep intronic mutations in X-linked Alport syndrome.

X-linked Alport syndrome (XLAS) is an inherited kidney disease caused exclusively by pathogenic variants in the COL4A5 gene. In 10-20% of cases, DNA sequencing of COL4A5 exons or flanking regions cannot identify molecular causes. Here, our objective was to use a transcriptomic approach to identify causative events in a group of 19 patients with XLAS without identified mutation by Alport gene panel sequencing. Bulk RNAseq and/or targeted RNAseq using a capture panel of kidney genes was performed. Alternative splicing events were compared to those of 15 controls by a developed bioinformatic score. When using targeted RNAseq, COL4A5 coverage was found to be 23-fold higher than with bulk RNASeq and revealed 30 significant alternative splicing events in 17 of the 19 patients. After computational scoring, a pathogenic transcript was found in all patients. A causative variant affecting COL4A5 splicing and absent in the general population was identified in all cases. Altogether, we developed a simple and robust method for identification of aberrant transcripts due to pathogenic deep-intronic COL4A5 variants. Thus, these variants, potentially targetable by specific antisense oligonucleotide therapies, were found in a high percentage of patients with XLAS in whom pathogenic variants were missed by conventional DNA sequencing.

Genomics in the kidney clinic.

Inherited diseases are a frequent cause of end-stage kidney disease and often seen in the kidney clinic. Clinical genomic testing is increasingly available in the UK and eligible patients in England can be referred through the NHS Genomic Medicine Service. Testing is useful for diagnosis, prognostication and management of conditions such as autosomal dominant polycystic kidney disease (ADPKD), Alport syndrome, autosomal dominant tubulointerstitial kidney disease (ADTKD) and focal segmental glomerulosclerosis (FSGS). As more patients undergo genomic testing and newer technologies such as whole genome sequencing are applied, we are developing a greater appreciation of the full phenotypic spectrum of inherited kidney diseases and the challenges associated with the interpretation of clinically significant variants.