Hiroshi Kaito

Rituximab for childhood-onset, complicated, frequently relapsing nephrotic syndrome or steroid-dependent nephrotic syndrome: a multicentre, double-blind, randomised, placebo-controlled trial

BACKGROUND Rituximab could be an effective treatment for childhood-onset, complicated, frequently relapsing nephrotic syndrome (FRNS) and steroid-dependent nephrotic syndrome (SDNS). We investigated the efficacy and safety of rituximab in patients with high disease activity. METHODS We did a multicentre, double-blind, randomised, placebo-controlled trial at nine centres in Japan. We screened patients aged 2 years or older experiencing a relapse of FRNS or SDNS, which had originally been diagnosed as nephrotic syndrome when aged 1-18 years. Patients with complicated FRNS or SDNS who met all other criteria were eligible for inclusion after remission of the relapse at screening. We used a computer-generated sequence to randomly assign patients (1:1) to receive rituximab (375 mg/m(2)) or placebo once weekly for 4 weeks, with age, institution, treatment history, and the intervals between the previous three relapses as adjustment factors. Patients, guardians, caregivers, physicians, and individuals assessing outcomes were masked to assignments. All patients received standard steroid treatment for the relapse at screening and stopped taking immunosuppressive agents by 169 days after randomisation. Patients were followed up for 1 year. The primary endpoint was the relapse-free period. Safety endpoints were frequency and severity of adverse events. Patients who received their assigned intervention were included in analyses. This trial is registered with the University Hospital Medical Information Network clinical trials registry, number UMIN000001405. FINDINGS Patients were centrally registered between Nov 13, 2008, and May 19, 2010. Of 52 patients who underwent randomisation, 48 received the assigned intervention (24 were given rituximab and 24 placebo). The median relapse-free period was significantly longer in the rituximab group (267 days, 95% CI 223-374) than in the placebo group (101 days, 70-155; hazard ratio: 0·27, 0·14-0·53; p<0·0001). Ten patients (42%) in the rituximab group and six (25%) in the placebo group had at least one serious adverse event (p=0·36). INTERPRETATION Rituximab is an effective and safe treatment for childhood-onset, complicated FRNS and SDNS. FUNDING Japanese Ministry of Health, Labour and Welfare.

Molecular analysis of digenic inheritance in Bartter syndrome with sensorineural deafness

Background: Bartter syndrome (BS) is a genetic disorder accompanied by hypokalaemic metabolic alkalosis. BS with sensorineural deafness (SND, OMIM602522) is a newly identified phenotype caused by mutations in the BSND gene that encodes barttin, a β-subunit for chloride channel ClC-Ka and ClC-Kb and classified as type IV BS. Type IV BS features the most severe phenotype entailing life-threatening neonatal volume depletion and chronic renal failure developing during infancy. A recent report described a case of BS with SND from a consanguineous family who showed homozygous mutations in the CLCNKA and CLCNKB genes. This case indicated the possibility of the occurrence of digenic inheritance in BS with SND resulting from double mutations in the CLCNKA and CLCNKB genes. Subject and results: The current report concerns a 2-year-old girl from a non-consanguineous family with BS accompanied by SND. In our case, four loss-of-function mutations, consisting of mutations in both parental alleles in both CLCNKA and CLCNKB, were identified. The paternal allele had a nonsense mutation (Q260X) in CLCNKA and a splicing site mutation (IVS17+1 g>a) in CLCNKB. The maternal allele had a large deletion mutation (about 12 kbp) extending from CLCNKA to CLCNKB. Our case provides clear evidence that loss-of-function alleles in both alleles of both CLCNKA and CLCNKB results in a phenotype indistinguishable from that of mutations in BSND (type IV BS). Conclusions: Recent advances in genetics have resulted in a better understanding of many human inherited diseases, but most of them are monogenic disorders and more complex inheritance patterns remain unresolved. Our case provides clear evidence of digenic inheritance outside the scope of Mendelian inheritance disorders.

A multicenter randomized trial indicates initial prednisolone treatment for childhood nephrotic syndrome for two months is not inferior to six-month treatment

In this multicenter, open-label, randomized controlled trial, we determined whether 2-month prednisolone therapy for steroid-sensitive nephrotic syndrome was inferior or not to 6-month therapy despite significantly less steroid exposure. The primary end point was time from start of initial treatment to start of frequently relapsing nephrotic syndrome. The pre-specified non-inferiority margin was a hazard ratio of 1.3 with one-sided significance of 5%. We randomly assigned 255 children with an initial episode of steroid-sensitive nephrotic syndrome to either 2 - or 6-month treatment of which 246 were eligible for final analysis. The total prednisolone exposure counted both initial and relapse prednisolone treatment administered over 24 months. Median follow-up in months was 36.7 in the 2-month and 38.2 in the 6-month treatment group. Time to frequent relaps was similar in both groups; however, the median was reached only in the 6-month group (799 days). The hazard ratio was 0.86 (90% confidence interval, 0.64–1.16) and met the non-inferior margin. Time to first relapse was also similar in both groups: median day 242 (2-month) and 243 (6-month). Frequency and severity of adverse events were similar in both groups. Most adverse events were transient and occurred during initial or relapse therapy. Thus, 2 months of initial prednisolone therapy for steroid-sensitive nephrotic syndrome, despite less prednisolone exposure, is not inferior to 6 months of initial therapy in terms of time to onset of frequently relapsing nephrotic syndrome.

The Pharmacological Characteristics of Molecular-Based Inherited Salt-Losing Tubulopathies

CONTEXT Our understanding of inherited salt-losing tubulopathies has improved with recent advances in molecular genetics. However, the terminology of Bartter syndrome and Gitelman syndrome does not always accurately reflect their pathophysiological basis or clinical presentation, and some patients are difficult to diagnose from their clinical presentations. OBJECTIVE In the present study, we conducted molecular analysis and diuretic tests for patients with inherited salt-losing tubulopathies to clarify the pharmacological characteristics of these disorders. PATIENTS We detected mutations and subsequently conducted diuretic tests using furosemide and thiazide for 16 patients with salt-losing tubulopathies (two with SLC12A1; two with KCNJ1; nine with CLCNKB; and three with SLC12A3). RESULTS Patients with SLC12A1 mutations showed no response to furosemide, whereas those with SLC12A3 mutations showed no response to thiazide. However, patients with CLCNKB mutations showed no response to thiazide and a normal response to furosemide, and those with KCNJ1 mutations showed a good response to both diuretics. This study revealed the following characteristics of these disorders: 1) subjects with CLCNKB mutations showed one or more biochemical features of Gitelman syndrome (including hypomagnesemia, hypocalciuria, and fractional chloride excretion insensitivity to thiazide administration); and 2) subjects with KCNJ1 mutations appeared to show normal fractional chloride excretion sensitivity to furosemide and thiazide administration. CONCLUSIONS These results indicate that these disorders are difficult to distinguish in some patients, even when using diuretic challenge. This clinical report provides important findings that can improve our understanding of inherited salt-losing tubulopathies and renal tubular physiology.

Molecular analysis of patients with type III bartter syndrome : Picking up large heterozygous deletions with semiquantitative PCR

Type III Bartter syndrome (BS) (OMIM607364) is caused by mutations in the basolateral chloride channel ClC-Kb gene (CLCNKB). The CLCNKB gene is sometimes reported as having a large deletion mutation, but all cases reported previously were large homozygous deletions and a large heterozygous deletion is impossible to detect by direct sequencing. This report concerns a genetic analysis of five Japanese patients with type III BS. To identify the mutations, we used polymerase chain reaction (PCR) and direct sequencing. To detect large heterozygous deletion mutations of the CLCNKB gene, we conducted semiquantitative PCR amplification using capillary electrophoresis. The result was that four mutations were identified, comprising one novel 2-bp deletion mutation, an entire heterozygous deletion, and a heterozygous deletion mutation of exons 1 and 2. The nonsense mutation W610X was detected in all patients, and this mutation is likely to constitute a founder effect in Japan. Capillary electrophoresis is a new method and extremely useful for detecting large heterozygous deletions, and should be used to examine type III BS cases in whom only a heterozygous mutation has been detected by direct sequencing. This is the first report to identify large heterozygous deletion mutations in the CLCNKB gene in patients with type III BS.

Risk factors for developing severe clinical course in HUS patients: a national survey in Japan

Background: Hemolytic uremic syndrome (HUS) is characterized by acute renal failure, thrombocytopenia and hemolytic anemia. Cases accompanied by prodromal gastrointestinal tract symptoms are referred to as typical HUS. Some severe HUS patients require dialysis or develop central nervous system (CNS) disorders after the onset of HUS.

Milder clinical aspects of X-linked Alport syndrome in men positive for the collagen IV α5 chain

X-linked Alport syndrome is caused by mutations in the COL4A5 gene encoding the type IV collagen α5 chain (α5(IV)). Complete absence of α5(IV) in the renal basal membrane is considered a pathological characteristic in male patients; however, positive α5(IV) staining has been found in over 20% of patients. We retrospectively studied 52 genetically diagnosed male X-linked Alport syndrome patients to evaluate differences in clinical characteristics and renal outcomes between 15 α5(IV)-positive and 37 α5(IV)-negative patients. Thirteen patients in the α5(IV)-positive group had non-truncating mutations consisting of nine missense mutations, three in-frame deletions, and one splice-site mutation resulting in small in-frame deletions of transcripts. The remaining two showed somatic mutations with mosaicism. Missense mutations in the α5(IV)-positive group were more likely to be located before exon 25 compared with missense mutations in the α5(IV)-negative group. Furthermore, urinary protein levels were significantly lower and the age at onset of end-stage renal disease was significantly higher in the positive group than in the negative group. These results help to clarify the milder clinical manifestations and molecular characteristics of male X-linked Alport syndrome patients expressing the α5(IV) chain.

Recurrent EIARF and PRES With Severe Renal Hypouricemia by Compound Heterozygous SLC2A9 Mutation

Renal hypouricemia (RHU) is a hereditary disease that predisposes affected people to exercise-induced acute renal failure (EIARF). In most patients with RHU, the disorder is caused by loss-of-function mutations in SLC22A12 (solute carrier family 22, member 12), which encodes urate transporter 1 (URAT1). Patients with RHU without any mutations in the URAT1 gene were recently found to have a mutation in the glucose transporter 9 (GLUT9) gene (SLC2A9 [solute carrier family 2, member 9]). Central nervous system complications seem to be rare in patients with RHU with SLC22A12 mutations. Here, we report the case of a girl with severe RHU (serum urate: 5.9 μmol/L [0.1 mg/dL]) associated with recurrent EIARF in whom the disease was caused by a compound heterozygous mutation in SLC2A9, a nonsense mutation in the paternal allele (p.G207X in exon 7), and a large duplication (c.1–2981_1204+16502) in the maternal allele detected by reverse-transcription polymerase chain reaction (PCR), semiquantitative PCR, long PCR, and direct sequencing. The episodes of EIARF were complicated by posterior reversible encephalopathy syndrome (PRES), which suggested a relationship between PRES and GLUT9 or severe hypouricemia. This is the second report of mutations of both alleles of SLC2A9 that resulted in severe hypouricemia. Our findings indicate that even a nonsense mutation responsible for the heterozygous status of SLC2A9 did not cause severe hypouricemia, and they lend support to previous speculation that mutations of both SLC2A9 alleles cause severe hypouricemia. Our case shows that GLUT9, unlike URAT1, may play a specific role in exercise-induced PRES.

Somatic mosaicism for a mutation of the COL4A5 gene is a cause of mild phenotype male Alport syndrome

BACKGROUND Alport syndrome is the most common form of hereditary nephritis and is mainly caused by mutations in the COL4A5 gene, which shows the X-linked form. It is well known that some male Alport syndrome cases show a relatively mild phenotype, but few molecular investigations have been conducted to clarify the mechanism of this phenotype. Methods and results. This report concerns an 8-year-old male sporadic Alport syndrome patient. While electron microscopy of the glomerular basement membrane showed typical findings for Alport syndrome, however, the immunohistochemical analysis of the glomerulus showed mosaic staining of the type IV collagen alpha 5 chain. The mutational analysis of the COL4A5 gene unexpectedly disclosed two peaks at the intron 43 splicing acceptor site (c. 3998-2 a/t) with direct sequencing. Restriction enzyme analysis demonstrated that the presence of somatic mosaicism was responsible for this mutation. mRNA extracted from the urinary sediments was analysed by RT-PCR and two PCR fragments were amplified, one consisting of a normal sequence and one with skipping of exon 44. CONCLUSIONS Our findings indicate that somatic mosaicism for COL4A5 is responsible for male X-linked Alport syndrome with an alpha 5 mosaic staining pattern. Several cases with somatic mosaicism have previously been reported, however, this is the first case where the presence of this mutation was proved with a comprehensive analysis of genomic DNA, mRNA and alpha 5 expression in the tissues. Somatic mosaicism may thus be one of the causes of the mild phenotype in Alport syndrome.

Clinical practice guideline for pediatric idiopathic nephrotic syndrome 2013: general therapy

Nephrotic syndrome is a disorder characterized by severe proteinuria, hypoproteinemia, and generalized edema resulting from damage to the glomerular basement membrane. In Western countries, nephrotic syndrome affects 2 of 100,000 children per year [1]. In Japan, approximately 1,300 new cases per year of pediatric nephrotic syndrome are reported to the Medical Aid for Specific Chronic Disease of Children and the disease develops in 5 of 100,000 children per year. Approximately 90 % of the cases of pediatric nephrotic syndrome are idiopathic, or of unknown cause. The first-line treatment for an initial episode of pediatric idiopathic nephrotic syndrome is oral steroid therapy, which leads to remission in approximately 80 % of cases (steroidsensitive nephrotic syndrome) [2]. However, 80 % of children with steroid-sensitive nephrotic syndrome experience one or more relapses, [3] and 50 % of these children have frequent relapses [4]. Those with frequently relapsing nephrotic syndrome are prone to suffer steroid-induced side effects such as obesity, growth impairment, hypertension, diabetes mellitus, osteoporosis, and adrenal insufficiency. Many cases of steroid-resistant nephrotic syndrome, where steroids are ineffective, progress to renal failure. Pediatric idiopathic nephrotic syndrome is a very important disease in the field of pediatric nephrology. The Scientific Committee in the Japanese Society for Pediatric Nephrology previously published the ‘‘Clinical Practice Guideline for Pediatric Idiopathic Nephrotic Syndrome’’ (2013). This is the English translation from the ‘‘Medical Therapy’’ portion of the guideline.Nephrotic syndrome is a disorder characterized by severe proteinuria, hypoproteinemia, and generalized edema resulting from damage to the glomerular basement membrane. In Western countries, nephrotic syndrome affects 2 of 100,000 children per year [1]. In Japan, approximately 1,300 new cases per year of pediatric nephrotic syndrome are reported to the Medical Aid for Specific Chronic Disease of Children and the disease develops in 5 of 100,000 children per year. Approximately 90 % of the cases of pediatric nephrotic syndrome are idiopathic, or of unknown cause. The first-line treatment for an initial episode of pediatric idiopathic nephrotic syndrome is oral steroid therapy, which leads to remission in approximately 80 % of cases (steroidsensitive nephrotic syndrome) [2]. However, 80 % of children with steroid-sensitive nephrotic syndrome experience one or more relapses, [3] and 50 % of these children have frequent relapses [4]. Those with frequently relapsing nephrotic syndrome are prone to suffer steroid-induced side effects such as obesity, growth impairment, hypertension, diabetes mellitus, osteoporosis, and adrenal insufficiency. Many cases of steroid-resistant nephrotic syndrome, where steroids are ineffective, progress to renal failure. Pediatric idiopathic nephrotic syndrome is a very important disease in the field of pediatric nephrology. The Scientific Committee in the Japanese Society for Pediatric Nephrology previously published the ‘‘Clinical Practice Guideline for Pediatric Idiopathic Nephrotic Syndrome’’ (2013). This is the English translation from the ‘‘Medical Therapy’’ portion of the guideline.