Yusuke Mitani

Paternal uniparental isodisomy of chromosome 22 in a patient with metachromatic leukodystrophy

Metachromatic leukodystrophy (MLD) is an autosomal recessive lysosomal storage disease caused by deficiency of the enzyme arylsulfatase A encoded by the ARSA gene located on 22q13.33. Typically, in autosomal recessive disease, a patient inherits two mutations from both parents who are heterozygous carriers. However, in some instances, it is possible to develop the disease by uniparental isodisomy (UPiD), in which two copies of the same mutated allele are inherited from only one carrier parent. Here, we report the first patient with MLD caused by UPiD of chromosome 22. The patient has a homozygous missense mutation, P136T, on ARSA. Family study of the ARSA gene and leukocyte enzyme activity revealed that his father and sister were heterozygous carriers, but his mother possessed only wild-type alleles and normal enzyme activity. Karyotypes of the patient and the parents were normal. Microsatellite analysis showed no discrepancy of parentage, and paternal UPiD of chromosome 22 was indicated. Finally, genome-wide single-nucleotide polymorphism array confirmed the region of UPiD was extended to the entire chromosome 22 of the patient.

Applying and testing the conveniently optimized enzyme mismatch cleavage method to clinical DNA diagnosis

Establishing a simple and effective mutation screening method is one of the most compelling problems with applying genetic diagnosis to clinical use. Because there is no reliable and inexpensive screening system, amplifying by PCR and performing direct sequencing of every coding exon is the gold standard strategy even today. However, this approach is expensive and time consuming, especially when gene size or sample number is large. Previously, we developed CEL nuclease mediated heteroduplex incision with polyacrylamide gel electrophoresis and silver staining (CHIPS) as an ideal simple mutation screening system constructed with only conventional apparatuses and commercially available reagents. In this study, we evaluated the utility of CHIPS technology for genetic diagnosis in clinical practice by applying this system to screening for the COL2A1, WRN and RPS6KA3 mutations in newly diagnosed patients with Stickler syndrome (autosomal dominant inheritance), Werner syndrome (autosomal recessive inheritance) and Coffin-Lowry syndrome (X-linked inheritance), respectively. In all three genes, CHIPS detected all DNA variations including disease causative mutations within a day. Direct sequencing of all coding exons of these genes confirmed 100% sensitivity and specificity. We demonstrate high sensitivity, high cost performance and reliability of this simple system, with compatibility to all inheritance modes. Because of its low technology, CHIPS is ready to use and potentially disseminate to any laboratories in the world.

A girl with infantile neuronal ceroid lipofuscinosis caused by novel PPT1 mutation and paternal uniparental isodisomy of chromosome 1

BACKGROUND Infantile neuronal ceroid lipofuscinosis (INCL) is an autosomal recessive disorder starting in infancy as early as 12-month-old, caused by PPT1 (palmitoyl-protein thioesterase 1) mutations, and characterized by progressive psychomotor deterioration, brain atrophy, myoclonic jerk and visual impairment. INCL can be diagnosed by brain magnetic resonance image (MRI) prior to rapid deterioration stage. To date, there is no INCL patient whose manifestation was caused by uniparental isodisomy (UPiD). PATIENT We reported a girl diagnosed with INCL. Genetic analysis revealed a novel PPT1 mutation c.20_47del28:p.Leu7Hisfs*21. Only the father of the patient was found as a carrier of this mutation. SNP array showed the mutation became homozygous by paternal UPiD of chromosome 1. DISCUSSION Although ICNL is a rare disease except in Finland, it is not difficult to diagnose it since the clinical symptoms and MRI findings are characteristic. Genetic testing is useful for definitive diagnosis, and distinction of UPiD is essential for genetic counseling.

Mutational analysis of TSC1 and TSC2 in Japanese patients with tuberous sclerosis complex revealed higher incidence of TSC1 patients than previously reported

Tuberous sclerosis complex (TSC) is an autosomal dominant disorder characterized by multiple hamartias and hamartomas involving throughout the body. To date, many TSC1 and TSC2 mutations have been reported all over the world, however, few TSC mutation studies have been performed in the Japanese population, and genetic characteristics of Japanese TSC patients are not yet clear. In this study, we analyzed TSC1 and TSC2 in 57 Japanese patients with TSC (8 familial and 49 sporadic; 46 definite and 11 suspect TSC) and identified 31 mutations including 11 TSC1 mutations (two familial and nine sporadic; all definite TSC) and 20 TSC2 mutations (2 familial and 18 sporadic; 19 definite and 1 suspect TSC). We also reviewed all Japanese TSC mutations previously reported. Our study demonstrates significantly higher incidence (P=0.007) of TSC1 mutations among sporadic TSC patients in the Japanese population compared with US and European studies. No differences emerged in mutation distributions and types in precedent studies, excepting low frequency of the TSC2 nonsense mutation. Comparing clinical manifestations, developmental delay and/or mental retardation were milder in TSC1 patients than TSC2 patients for its frequency (P=0.032) and severity (P=0.015); however, no other symptoms were clearly different.

A Say–Barber–Biesecker–Young–Simpson variant of Ohdo syndrome with a KAT6B 10-base pair palindromic duplication: a recurrent mutation causing a severe phenotype mixed with genitopatellar syndrome

The Say‐Barber‐Biesecker‐Young‐Simpson variant of Ohdo syndrome (SBBYSS) (MIM# 603736) and genitopatellar syndrome (GPS) (MIM#606170) are allelic diseases caused by KAT6B mutation. Genotype–phenotype correlation is assumed, but a few patients manifest overlapping features of both syndromes. Here we report the case of a boy with SBBYSS. He had a KAT6B mutation previously reported in typical SBBYSS, but he also manifested severe developmental delay, as well as genital features and laryngomalacia requiring tracheostomy that conformed to GPS.

CHIPS for genetic testing to improve a regional clinical genetic service

In current practice of clinical genetics, molecular diagnosis has become more widely used than ever before. DNA diagnosis is important for appropriate medical care of the patient, and proper genetic counseling to the family. However, genetic testing of orphan disease cannot always be performed easily. In multiple congenital anomalies (MCA) syndromes by monogenic cause, the broad mutational spectrum and large size of responsible genes often make molecular diagnosis expensive and cumbersome. We solve this problem with on‐demand genetic testing by CHIPS (CEL nuclease mediated heteroduplex incision with polyacrylamide gel electrophoresis and silver staining) technology, which is the ultimately conventional and economical mutation screening system. In this article, we show eight patients with MCA syndromes who were recently treated at our hospital, and demonstrate that CHIPS successfully offers efficient and inexpensive genetic testing and facilitates clinical genetic service in our local region.

Deep spontaneous molecular remission in a patient with congenital acute myeloid leukemia expressing a novel MOZ-p300 fusion transcript

Yasuhiro Ikawa , Ryosei Nishimura, Hideaki Maeba, Toshihiro Fujiki , Rie Kuroda, Kazuhiro Noguchi, Masaki Fukuda, Shintaro Mase, Raita Araki, Yusuke Mitani, Tomohiko Sato, Kiminori Terui, Etsurou Ito, Issay Kitabayashi and Akihiro Yachie Department of Pediatrics, School of Medicine, Institute of Medical, Pharmaceutical and Health Sciences, Kanazawa University, Kanazawa, Ishikawa, Japan; Department of Pediatrics, Hirosaki University School of Medicine, Hirosaki, Aomori Prefecture, Japan; Division of Hematological Malignancy, National Cancer Center Research Institute, Tokyo, Japan

A successful case of neoadjuvant chemotherapy and radical hysterectomy during pregnancy for advanced uterine cervical cancer accompanied by neonatal erythroderma: Radical hysterectomy after NAC in pregnancy

Recent reports showed that neoadjuvant chemotherapy (NAC) has been successfully applied to treat advanced uterine cervical cancers during pregnancy. However, its side effects on the fetus remain unclear. Here, we report a 33‐year‐old primipara who underwent four courses of NAC therapy, paclitaxel and cisplatin, from 17 to 27 weeks of gestation due to uterine cervical cancer stage IB2. At 31 weeks of gestation, cesarean section and radical hysterectomy were performed, and a female baby weighing 1446 g was born. Although pre‐ and postnatal courses were uneventful, neonatal erythroderma over the entire body was observed just after delivery. The pathological diagnosis was ichthyosiform erythroderma, which was later demonstrated to be keratitis‐ichthyosis‐deafness syndrome, by exome sequencing analysis. Although her skin disorder was consistent with keratitis‐ichthyosis‐deafness syndrome, the skin condition gradually improved after delivery. These findings suggest that NAC therapy during pregnancy might cause or exacerbate systemic skin lesions in the fetus/neonate.

The Japanese experience and pharmacokinetics of antenatal maternal high-dose immunoglobulin treatment as a prophylaxis for neonatal hemochromatosis in siblings

Abstract Background: Neonatal hemochromatosis (NH) is a rare but serious disease causing fulminant hepatic failure. The recurrence rate of NH in a subsequent infant of a mother with an affected infant is 70–90%. Recently, antenatal maternal high-dose intravenous immunoglobulin (IVIG) treatment has been reported to be effective for preventing NH recurrence. However, data on the IgG concentrations during this treatment are limited. Objective: We report a Japanese experience and present a pharmacokinetic simulation model of IgG during IVIG treatment. Methods: Women with histories of pregnancy diagnosed with NH were treated with IVIG weekly from the second trimester until the end of gestation. Serum IgG levels during treatment were collected frequently and pharmacokinetics were simulated by a two-compartment model. Results: Six women were included during eight pregnancies. None experienced severe adverse events. Three out of eight infants showed temporary liver dysfunction, but none required any treatment. A simulation study showed that the estimated trough and peak levels of IgG concentrations during IVIG were 2000–3000 and 4000–5000 mg/dl, respectively. Conclusion: This treatment prevented the recurrence of NH in siblings in Japanese women. We examined the details of serum IgG concentrations and introduced a new pharmacokinetic simulation model of IgG concentrations during IVIG treatment.

XL-EDA-ID Presenting with Congenital Duodenal Atresia and Perforations

To the Editor: Clinical features of X-linked anhidrotic ectodermal dysplasia with immunodeficiency (XL-EDA-ID) include hypohidrosis, delayed eruption of teeth, coarse hair, and immunodeficiency associated with frequent bacterial infections [1]. XL-EDA-ID is caused by dysfunction of the IKBKG gene, which encodes NEMO, the NF-kB essential modulator. NEMO participates in activation of the IkB kinase complex, which enables nuclear translocation of NF-kB dimers. It is well known that patients with XL-EDA-ID are susceptible to inflammatory colitis [1]; however, there is no report of XLEDA-ID presenting with congenital duodenal atresia or multiple intestinal perforations. A male infant who was prenatally diagnosed as having duodenal obstruction was born at 39 weeks of gestation via elective cesarean section. His birth weight was 2300 g. There was no family history of ectodermal dysplasia or immunodeficiency. The patient had bilious nasogastric aspirates and abdominal distension. Plain radiographs showed a double-bubble sign, and a contrast study revealed complete duodenal obstruction (Fig. 1a). Laboratory studies revealed a white blood cell count of 15,900/μL, a hemoglobin level of 12.8 g/dL, and a serum Creactive protein level of 0.0 mg/dL. On day 1, exploratory laparotomy showed hemorrhagic ascites, complete duodenal atresia, multiple perforations of the duodenum and proximal jejunum (Fig. 1b), annular pancreas, and intestinal malrotation. Subsequently, duodenojejunostomy was performed. There was no evidence of meconium peritonitis. Histological examination of the duodenum showed mild congestion, bleeding, focal mucosal atrophy, and a defect of the muscle layer (Fig. 1c, d). A few fibrinoid materials and inflammatory cells were also observed. The postoperative period was unremarkable until postoperative day 11, when the patient presented with prolonged fever that did not easily respond to antibiotics treatment and required 2 months of hospitalization. Despite repeated cultures of blood, urine, and stool, no pathogenic organisms were isolated. No cytomegalovirus DNA was detected in his dried umbilical cord by polymerase chain reaction (PCR). Shortly after discharge from the hospital, the patient had frequent episodes of fever, vomiting, and diarrhea. At 5 months of age, gastroduodenal anastomosis was performed because of frequent vomiting, poor weight gain, and stagnation of contrast medium at the anastomotic region. At 6 months, the patient was hospitalized again for gastrointestinal hemorrhage caused by anastomotic ulcer. Because he showed positive cytomegalovirus antigenemia and Pneumocystis jirovecii pneumonia, primary immunodeficiency was clinically suspected. Prophylactic use of antibiotics and antifungals was initiated. However, an immunological evaluation was largely normal. Serum immunoglobulin (Ig) levels were as follows: IgG, 767 mg/dL (normal, 290–950); IgA, 94 mg/dL (normal, 8–50); and IgM, 131 mg/dL (normal, 46–176). There was no lymphopenia, and immunophenotypic analysis of lymphocytes showed a normal percentage of CD3 T (71.0%; normal, 67.5 ± 6.9%) and CD4 T cells (41.0%; normal, 45.6 ± 9.3%), with a slight increase in the percentage of CD8 T cells (26.2%; normal, 15.2 ± 4.2%) and CD20 B cells (28.4%; normal, 16.7 ± 5.4%). Lymphocytes showed normal proliferation in response to phytohemagglutinin. Levels of T cell receptor excision circles and kappa-deleting recombination excision circles were normal. Whole exome sequencing failed to identify any pathogenic mutations in genes related to primary immunodeficiency. Mutations of duodenal atresia-related genes, such as NKX3–2, HNF1B, and RFX6, were also absent. * Taizo Wada taizo@staff.kanazawa-u.ac.jp