NEXT GENERATION SEQUENCING: IMPLICATIONS IN CLINICAL PRACTICE AND DIAGNOSIS OF STEROID-RESISTANT NEPHROTIC SYNDROME

PROVENZANO, ALDESIA;B. Tesi;B. Mazzinghi;E. Contini;V. Cetica;D. Mei;G. Lavoratti;M. Materassi;P. Romagnani;S. Giglio

2011-01-01

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Abstract

During recent years, several podocyte genes have been implicated in severe forms of steroid-resistant nephrotic syndrome (SRNS) progressing to renal failure. To date, at least 14 genes highly expressed in the podocyte have been associated with the syndrome and different mutations in these genes have been identified; it is now known that the phenotypes associated with mutations in these genes display significant variability, rendering genetic testing and counselling a more complex task. Due to this large genetic heterogeneity and missing of adequate gene-diagnostic tools, most patients are not genetically characterized, which would be important for individualized patient care. Currently, next-generation sequencing technologies are revolutionizing genetic, since they are capable to produce billions of bases of sequence information in a single experiment. Accordingly, this powerful technology can now also open avenues for genetic diagnostics. The scope of our work is to illustrate clinical applications of the Next Generation Sequencing (NGS) technology to study patients affected by SRNS, in which the previous analysis of NPHS2 and WT1 genes had not shown pathogenetic mutations. We perform in 4 affected subjects a targeted resequencing of 23 genes including those already known as causative of the disorder and several other genes highly expressed in the podocyte. In the case of a young girl with SRNS and focal segmental glomerulosclerosis (FSGS), which quickly led to end stage kidney disease (ESKD) and dialysis treatment, we found a new heterozygous missense mutation in the ACTN4 gene, which encodes for α-actinin-4, a protein of podocyte cytoskeleton. Mutations in this gene are associated with an autosomal dominant form of FSGS characterized by adult-onset of proteinuria. In another patient who had received at 2 years of age the diagnosis of SRNS and Minimal-change disease (MCD) and currently under dialysis treatment, we discovered a new heterozygous missense mutation in the PLCE1 gene. In literature rare mutations reported in this gene are associated with diffuse mesangial sclerosis. We also detected a variation in the ZHX2 gene, that to date has never been associated with SRNS. This gene encode for a transcriptional factor which regulates the expression of several genes in the podocyte, including WT1, during the development of nephrotic syndrome. The combination of the variations in the known genes for SRNS and the identification of this novel gene combined with the extensive clinical investigations provides an exciting opportunity to reveal further insight into the pathogenic mechanisms that underlie this debilitating disorder.