Exome Sequencing and Identification of Phenocopies in Patients With Clinically Presumed Hereditary Nephropathies.

RATIONALE & OBJECTIVE: Hereditary nephropathies are clinically and genetically heterogeneous disorders. For some patients, the clinical phenotype corresponds to a specific hereditary disease but genetic testing reveals that the expected genotype is not present (phenocopy). The aim of this study was to evaluate the spectrum and frequency of phenocopies identified by using exome sequencing in a cohort of patients who were clinically suspected to have hereditary kidney disorders. STUDY DESIGN: Cross-sectional cohort study. SETTING & PARTICIPANTS: 174 unrelated patients were recruited for exome sequencing and categorized into 7 disease groups according to their clinical presentation. They included autosomal dominant tubulointerstitial kidney disease, Alport syndrome, congenital anomalies of the kidney and urinary tract, ciliopathy, focal segmental glomerulosclerosis/steroid-resistant nephrotic syndrome, VACTERL association, and "other." RESULTS: A genetic diagnosis (either likely pathogenic or pathogenic variant according to the guidelines of the American College of Medical Genetics) was established using exome sequencing in 52 of 174 (30%) cases. A phenocopy was identified for 10 of the 52 exome sequencing-solved cases (19%), representing 6% of the total cohort. The most frequent phenocopies (n=5) were associated with genetic Alport syndrome presenting clinically as focal segmental glomerulosclerosis/steroid-resistant nephrotic syndrome. Strictly targeted gene panels (<25 kilobases) did not identify any of the phenocopy cases. LIMITATIONS: The spectrum of described phenocopies is small. Selection bias may have altered the diagnostic yield within disease groups in our study population. The study cohort was predominantly of non-Finnish European descent, limiting generalizability. Certain hereditary kidney diseases cannot be diagnosed by using exome sequencing (eg, MUC1-autosomal dominant tubulointerstitial kidney disease). CONCLUSIONS: Phenocopies led to the recategorization of disease and altered clinical management. This study highlights that exome sequencing can detect otherwise occult genetic heterogeneity of kidney diseases.

Cyclosporine A responsive congenital nephrotic syndrome with single heterozygous variants in NPHS1, NPHS2, and PLCE1.

BACKGROUND: Congenital nephrotic syndrome (CNS) is primarily a monogenetic disease, with the majority of cases due to changes in five different genes: the nephrin (NPHS1), podocin (NPHS2), Wilms tumor 1 (WT1), laminin ß2 (LAMB2), and phospholipase C epsilon 1 (PLCE1, NPHS3) gene. Usually CNS is not responsive to immunosuppressive therapy, but treatment with ACE inhibitors, AT1 receptor blockade and/or indomethacin can reduce proteinuria. If the disease progresses to end-stage renal disease, kidney transplantation is the therapy of choice. CASE-DIAGNOSIS: Here, we present the case of a 4-month-old girl with congenital nephrotic syndrome. Upon admission, the patient presented with life-threatening anasarca, hypoalbuminemia, proteinuria, and impaired growth. There was no evidence of an infectious or immunological etiology. The genetic evaluation revealed a heterozygous variant in NPHS1 (p.Arg207Trp), in NPHS2 (p.Ser95Phe) as well as in PLCE1 (p.Ala1045Ser) and did not explain CNS. In addition to daily parenteral albumin infusions plus furosemide, a pharmacological antiproteinuric therapy was started to reduce protein excretion. Based on the genetic results, immunosuppressive therapy with prednisolone was initiated, but without response. However, following cyclosporine A treatment, the patient achieved complete remission and now has good renal function, growth, and development. CONCLUSIONS: A profound search for the cause of CNS is necessary but has its limitations. The therapeutic strategy should be adapted when the etiology remains unclear.

Identification of co-occurrence in a patient with Dent's disease and ADA2-deficiency by exome sequencing.

Patients with co-occurrence of two independent pathologies pose a challenge for clinicians as the phenotype often presents as an unclear syndrome. In these cases, exome sequencing serves as a powerful instrument to determine the underlying genetic causes. Here, we present the case of a 4-year old boy with proteinuria, microhematuria, hypercalciuria, nephrocalcinosis, livedo-like rash, recurrent abdominal pain, anemia and continuously elevated CRP. Single exome sequencing revealed the pathogenic nonsense mutation p.(Arg98*) in the CLCN5 gene causing the X-linked inherited, renal tubular disorder Dent's disease. Furthermore, the two pathogenic and compound heterozygous missense variants p.(Gly47Ala) and p.(Pro251Leu) in the CECR1 gene could be identified. Mutations in the CECR1 gene are associated with a hereditary form of polyarteritis nodosa, called ADA2-deficiency. Both parents were carriers of a single heterozygous variant in CECR1 and the mother was carrier of the CLCN5 variant. This case evidently demonstrates the advantage of whole exome sequencing compared to single gene testing as the pathology in the CECR1 gene might have only been diagnosed after the occurrence of signs of systemic vasculitis like strokes or hemorrhages. Therefore, treatment and prevention can now start early to improve the outcome of these patients.

Identification of 47 novel mutations in patients with Alport syndrome and thin basement membrane nephropathy.

BACKGROUND: Alport syndrome (ATS) is a progressive hereditary nephropathy characterized by hematuria and proteinuria. It can be associated with extrarenal manifestations. In contrast, thin basement membrane nephropathy (TBMN) is characterized by microscopic hematuria, is largely asymptomatic, and is rarely associated with proteinuria and end-stage renal disease. Mutations have been identified in the COL4A5 gene in ATS and in the COL4A3 and COL4A4 genes in ATS and TBMN. To date, more than 1000 different mutations in COL4A5, COL4A3, and COL4A4 are known. METHODS: In this study mutational analysis by exon sequencing and multiplex ligation-dependent probe amplification was performed in a large European cohort of families with ATS and TBMN. RESULTS: Molecular diagnostic testing of 216 individuals led to the detection of 47 novel mutations, thereby expanding the spectrum of known mutations causing ATS and TBMN by up to 10 and 6%, respectively, depending on the database. Remarkably, a high number of ATS patients with only single mutations in COL4A3 and COL4A4 were identified. Additionally, three ATS patients presented with synonymous sequence variants that possible affect correct mRNA splicing, as suggested by in silico analysis. CONCLUSIONS: The results of this study clearly broaden the genotypic spectrum of known mutations for ATS and TBMN, which will in turn now facilitate future studies into genotype-phenotype correlations. Further studies should also examine the significance of single heterozygous mutations in COL4A3 and COL4A4 and of synonymous sequence variants associated with ATS.