BRIEF COMMUNICATION www.jasn.org Nephrin Mutations Can Cause Childhood-Onset Steroid-Resistant Nephrotic Syndrome Aure´lie Philippe,*† Fabien Nevo,*† Ernie L. Esquivel,*† Dalia Reklaityte,*† ʈ Olivier Gribouval,*† Marie-Jose`phe Teˆte,*‡ Chantal Loirat,§ Jacques Dantal, Michel Fischbach,¶ Claire Pouteil-Noble,** Ste´phane Decramer,†† Martin Hoehne,‡‡ Thomas Benzing,‡‡ Marina Charbit,‡ Patrick Niaudet,*†‡ and Corinne Antignac*†§§ † *Inserm U574, Hoˆpital Necker-Enfants Malades, Universite´ Paris Descartes, Faculte´deMe´ decine Rene´ Descartes, BRIEF COMMUNICATION ‡Pediatric Nephrology and §§Department of Genetics, Hoˆpital Necker-Enfants Malades, Assistance Publique-Hoˆpitaux de Paris, and §Pediatric Nephrology Department, Universite´ Paris VII, Assistance Publique-Hoˆpitaux de Paris, Hoˆpital ʈ Robert Debre´, Paris, ITERT, Department of Nephrology and Clinical Immunology, CHU Nantes, Nantes, ¶Nephrology Dialysis Transplantation Children’s Unit, Hoˆpital de Hautepierre, Strasbourg, **Transplantation and Nephrology Unit, Centre Hospitalier Lyon-Sud, Pierre-Be´nite, and ††Department of Pediatric Nephrology, Hoˆpital des Enfants, and Inserm, U858/I2MR, Department of Renal and Cardiac Remodeling, Toulouse, France; and ‡‡Department of Medicine IV, University of Cologne, Cologne, Germany ABSTRACT Classically, infants with mutations in NPHS1, which encodes nephrin, present with but has since been described in other nephrotic syndrome within the first 3 mo of life (congenital nephrotic syndrome of populations.3–5 Nephrin is a single-pass the Finnish-type), and children with mutations in NPHS2, which encodes podocin, transmembrane protein consisting of present later with steroid-resistant nephrotic syndrome. Recently, however, eight extracellular Ig-like modules, a fi- NPHS2 mutations have been identified in children with congenital nephrotic syn- bronectin type III–like motif, and a cy- drome. Whether NPHS1 mutations similarly account for some cases of childhood tosolic C-terminal tail. Homodimers of steroid-resistant nephrotic syndrome is unknown. In this study, 160 patients who nephrin and heterodimers with the glo- belonged to 142 unrelated families and presented with nephrotic syndrome at merular protein NEPH1 constitute the least 3 mo after birth were screened for NPHS1 variants once mutations in NPHS2 structural basis of the slit diaphragm.6,7 had been excluded. Compound heterozygous NPHS1 mutations were identified in In addition to its structural function, one familial case and nine sporadic cases. Mutations included protein-truncating nephrin is involved in podocyte signal- nonsense and frameshift mutations, as well as splice-site and missense variants. ing events.8 Mutations were classified as “severe” or “mild” using prediction algorithms and Mutations in the NPHS2 gene en- functional assays. Most missense variants trafficked normally to the plasma mem- coding podocin were, thereafter, de- brane and maintained the ability to form nephrin homodimers and to heterodimer- scribed in patients presenting with au- ize with NEPH1, suggesting retained function. The presence of at least one “mild” tosomal recessive SRNS with onset mutation in these patients likely explains the later onset and milder course of typically between 3 mo and 5 yr of age.9 disease. These results broaden the spectrum of renal disease related to nephrin NPHS2 mutations account for 42% of mutations. Received January 17, 2008. Accepted May 14, J Am Soc Nephrol 19: 1871–1878, 2008. doi: 10.1681/ASN.2008010059 2008. Published online ahead of print. Publication date available at www.jasn.org. Idiopathic nephrotic syndrome (NS) improved understanding of the heredi- A.P., F.N., and E.L.E. contributed equally to this represents a heterogeneous group of tary basis of NS.1 Mutations in the work. glomerular disorders occurring mainly NPHS1 gene, encoding the podocyte- Correspondence: Dr. Corinne Antignac, Inserm in children and may be classified as ste- expressed protein nephrin, lead to the U574, 6e`mee´ tage, Tour Lavoisier, Hoˆpital Necker- Enfants Malades, 147, rue de Se`vres, 75015 Paris, roid-sensitive (SSNS) or steroid-resis- congenital NS of the Finnish-type France. Phone: ϩ33-1-44-49-50-98; Fax: ϩ33-1- tant (SRNS) on the basis of response to (CNF), which is inherited in an autoso- 44-49-02-90; E-mail: [email protected] corticosteroid therapy. Gene discovery mal recessive manner. It affects approx- Copyright ᮊ 2008 by the American Society of efforts in the past decade have led to an imately 1:10,000 newborns in Finland2 Nephrology J Am Soc Nephrol 19: 1871–1878, 2008 ISSN : 1046-6673/1910-1871 1871 BRIEF COMMUNICATION www.jasn.org familial and 10% of sporadic cases of and who presented with noncongenital volvement was reported. At the end of SRNS.10 Proper assembly of nephrin and onset and a more protracted course of follow-up, only five patients had reached other slit diaphragm constituents and renal disease. ESRD, at a mean age of 13.6 yr (range 6 to trafficking to the plasma membrane and Mutation screening was performed in 25 yr), whereas six patients had normal to lipid rafts require interaction with a cohort of familial and sporadic cases of serum creatinine (Table 1). Four patients podocin.11,12 SRNS, for which NPHS2 mutations had successfully received a renal allograft, Classically, mutations in the NPHS1 been excluded by sequencing the exonic with no disease recurrence. and NPHS2 genes have been distin- regions and intronic junctions. Nephrin In total, 14 mutations, 12 of which are guished by their implications in familial mutations were found in one family with novel, were identified in 10 unrelated pa- congenital (onset at birth to 3 mo) and in two affected siblings, among 44 families tients. These included six nonsense and childhood-onset (later than 3 mo) cases, with familial SRNS, using a combination frameshift, two splice-site, and six mis- respectively. Hinkes et al.13 recently con- of linkage analysis and NPHS1 sequencing sense mutations, uniformly distributed firmed the findings of others that pa- (Table 1). Of 98 patients with sporadic throughout the NPHS1 gene (Figure 1). tients with NPHS1 mutations present SRNS, nine individuals were compound Segregation of mutations confirmed re- with NS exclusively during the first 3 mo heterozygotes for NPHS1 mutations (Ta- cessive inheritance. All nonsense and of life.3–5,14 They also identified NPHS2 ble 1). The mean age of onset of NS in these frameshift mutations are predicted to re- mutations in 39% of children with con- 11 patients was 3.0 yr (range 6 mo to 8 yr); sult in a truncated protein. In addition, genital onset of NS,13 as described previ- hence, later than previously described for the c.3720_3735delC (p.L1240fs1286X) ously,15 therein broadening the spec- patients with NPHS1 mutations. The NS mutation is unusual because it involves a trum of NPHS2-associated renal disease. was resistant to corticosteroids in all cases, deletion of the last seven nucleotides of We therefore sought to determine as well as to cyclosporine and cyclophos- the NPHS1 coding and of 9 bp of the 3Ј- whether mutations in the nephrin gene phamide, when additional treatments untranslated regions. The resulting pro- may similarly account for a wider range were attempted. Renal biopsy performed tein lacks the usual terminal valine resi- of disease presentations to include child- at the time of presentation revealed me- due and instead bears an additional 45 hood-onset SRNS. Our studies revealed sangioproliferative lesions in one pa- amino acids with no known sequence that compound heterozygous mutations tient, minimal-change disease in six pa- homology. in the NPHS1 gene were responsible for tients, and FSGS in three patients (Table Six missense mutations were identi- SRNS in a cohort of patients in whom 1). Characteristic tubular lesions of CNF fied in this cohort of patients, and all NPHS2 mutations had been excluded were not observed. No extrarenal in- were ruled out as polymorphisms by se- Table 1. Clinical data of patients who had SRNS and in whom NPHS1 mutations were identifieda Age of Onset Patient Gender Biopsy Therapy Evolution Mutation 1 Severe Mutation 2 Mild Pu (NS) (yr) 1420 F 0.25 (3.00) MCNS CS, CP Normal Cr at 14 yr c.609–2A3C (M) c.319G3A p.A107T (P) 446 F 0.80 (0.80) MCNS CS ESRF at 9 yr c.3720_3735del16 c.1724C3A p.L1240fs1286Xb (P) p.P575Q (M) 1075 F 0.50 (0.50) FSGS Unknown ESRF at 13 yr c.1379G3A c.2928G3T p.R460Qb (?) p.R976S (?) 841 F 3.80 (3.80) FSGS CS, CsA Normal Cr at 6 yr c.468C3G c.2928G3T p.Y156X (P) p.R976S (M) 466 F 0.25 (0.75) MCNS CS, CP Normal Cr at 10 yr c.2479C3T c.2928G3T p.R827X (M) p.R976S (P) 1167 F 3.10 (3.10) FSGS CS, CP ESRF at 15 yr c.516delC c.2928G3T p.T712fs175X (M) p.R976S (P) 693 M 8.00 (8.00) MCNS CS Normal Cr at 16 yr c.1134–1135delGC c.286C3G p.R379fs417X (M) p.L96V (P) 1407 F 5.00 (5.00) MCNS CS, CP ESRF at 25 yr c.516delC c.2928G3T p.T712fs175X (M) p.R976S (P) 634 M 3.00 (3.00) MP Unknown ESRF at 6 yr c.1491delC c.2072–6C3G (P) p.S494fs547X (M) 771c F 2.20 (2.80) MCNS CS, CsA Normal Cr at 9 yr c.2495T3C c.2928G3T 1462c M 2.50 (2.50) Not performed No treatment Normal Cr at 6 yr p.L832P (P) p.R976S (M) aCP, cyclophosphamide; Cr, creatinine; CS, corticosteroids; CsA, cyclosporin A; ESRF, end-stage renal failure; MCNS, minimal-change glomerulonephritis; MP, mesangioproliferative glomerulonephritis; Pu, proteinuria. bThis mutation may potentially be a “mild” one. Mode of transmission of mutant alleles are indicated in parentheses: M, maternal; P, paternal; ?, unknown. cSiblings with the same parents. 1872 Journal of the American Society of Nephrology J Am Soc Nephrol 19: 1871–1878, 2008 www.jasn.org BRIEF COMMUNICATION Figure 1.