Kiyoshi Kawakami

The landscape of somatic mutations in Down syndrome–related myeloid disorders

Transient abnormal myelopoiesis (TAM) is a myeloid proliferation resembling acute megakaryoblastic leukemia (AMKL), mostly affecting perinatal infants with Down syndrome. Although self-limiting in a majority of cases, TAM may evolve as non-self-limiting AMKL after spontaneous remission (DS-AMKL). Pathogenesis of these Down syndrome–related myeloid disorders is poorly understood, except for GATA1 mutations found in most cases. Here we report genomic profiling of 41 TAM, 49 DS-AMKL and 19 non-DS-AMKL samples, including whole-genome and/or whole-exome sequencing of 15 TAM and 14 DS-AMKL samples. TAM appears to be caused by a single GATA1 mutation and constitutive trisomy 21. Subsequent AMKL evolves from a pre-existing TAM clone through the acquisition of additional mutations, with major mutational targets including multiple cohesin components (53%), CTCF (20%), and EZH2, KANSL1 and other epigenetic regulators (45%), as well as common signaling pathways, such as the JAK family kinases, MPL, SH2B3 (LNK) and multiple RAS pathway genes (47%).

Expression of vascular endothelial growth factor in sera and lymph nodes of the plasma cell type of Castleman's disease

To evaluate the possible involvement of vascular endothelial growth factor (VEGF) in the pathogenesis of Castlemans disease, we studied VEGF levels in sera and supernatants of cultured lymph nodes from two patients with the plasma cell type of Castlemans disease, and analysed the expression of VEGF immunohistochemically in the lymph nodes. Clinically, one patient was classified as the localized type and the other as the multicentric type. Histologically, mature plasma cells and hyalinized vessels were prominent in the interfollicular region. The VEGF levels of the sera and the supernatants of cultured lymph nodes of both patients were higher than those of normal controls. VEGF was strongly expressed in plasma cells in the interfollicular region of the lymph nodes of both patients, but rarely in normal lymph nodes. Our results suggest that VEGF may be involved in the marked vascular proliferation in the interfollicular region of the lymph nodes of the plasma cell type of Castlemans disease.

Down syndrome and GATA1 mutations in transient abnormal myeloproliferative disorder: mutation classes correlate with progression to myeloid leukemia

Twenty percent to 30% of transient abnormal myelopoiesis (TAM) observed in newborns with Down syndrome (DS) develop myeloid leukemia of DS (ML-DS). Most cases of TAM carry somatic GATA1 mutations resulting in the exclusive expression of a truncated protein (GATA1s). However, there are no reports on the expression levels of GATA1s in TAM blasts, and the risk factors for the progression to ML-DS are unidentified. To test whether the spectrum of transcripts derived from the mutant GATA1 genes affects the expression levels, we classified the mutations according to the types of transcripts, and investigated the modalities of expression by in vitro transfection experiments using GATA1 expression constructs harboring mutations. We show here that the mutations affected the amount of mutant protein. Based on our estimates of GATA1s protein expression, the mutations were classified into GATA1s high and low groups. Phenotypic analyses of 66 TAM patients with GATA1 mutations revealed that GATA1s low mutations were significantly associated with a risk of progression to ML-DS (P < .001) and lower white blood cell counts (P = .004). Our study indicates that quantitative differences in mutant protein levels have significant effects on the phenotype of TAM and warrants further investigation in a prospective study.

Different genomic and metabolic patterns between mass screening-positive and mass screening-negative later-presenting neuroblastomas

The mass screening of neuroblastoma has been undertaken in Japan by measuring urinary vanillylmandelic acid (VMA) and homovanillic acid (HVA) in all infants at the age of 6 months. This program may not only improve the prognosis but also provide important insights into the biology and evolution of human neuroblastoma. The authors studied and discuss the clinical significance of the N‐myc oncogene, catecholamine metabolism, and other tumor markers in 43 patients with neuroblastoma who underwent the urinary screening test at 6 months of age. Thirty patients were found by the screening, and 13 were negative at the screening but later had a tumor. In the former group, the tumors were mostly in early stages (Stage I, 12; Stage II, 11; Stage III, seven), no amplification of N‐myc was observed, and all patients are alive without disease. Although two patients whose urine at the screening showed elevated VMA and HVA levels and accidentally were not treated for 13 and 17 months, there was no change in the values of VMA and HVA during that time. However, in the latter group, the tumors were mostly in advanced stages (Stage I, one; Stage III, four; Stage IV, eight), and N‐myc amplification was observed in seven of 13. Only two of these 13 are alive without disease. The age at diagnosis of the screening‐negative group was 23 months compared with 8 months in the patients identified by screening, and the pattern of catecholamine metabolites in the screening‐negative group tended to be dopaminergic with a low VMA–HVA ratio, especially in cases with N‐myc amplification. These data suggest that the screening‐positive patients with neuroblastoma may have favorable characteristics, and the biology of these tumors may be different from that of screening‐negative later‐presenting tumors. They also suggest that there may be at least two distinct subsets of neuroblastoma. For the early detection of the poor prognostic neuroblastomas, the measurement of urinary dopamine with VMA and HVA at later ages, such as 1 to 2 years, should be considered.

N-myc oncogene and stage IV-S neuroblastoma : preliminary observations on ten cases

We studied the clinical significance of genomic amplification of N‐myc in Stage IV‐S neuroblastoma, with reference to spontaneous regression. Among 103 neuroblastomas in which N‐myc was measured, ten were Stage IV‐S (eight children were younger than and two were older than 1 year of age). The number of copies of N‐myc was 1 to 3 in five patients, four to ten in one patient, and more than ten in four patients, and the survivors of each group were four, one, and one (recurrent), respectively. Of 41 patients younger than 1 year of age, N‐myc amplification of more than three copies was found only in Stage IV‐S neuroblastoma. Cure with a tendency to regress spontaneously was seen in five of eight patients younger than 1 year of age. However, two patients older than 1 year of age classified as Stage IV‐S (one with N‐myc amplification) died of progressive disease. In two patients (1 and 3 months of age) with a huge hepatic involvement and in whom the tumor had an amplified N‐myc of more than ten copies, tumor regression occurred but there was a relapse to a progressive state later. The overexpression of N‐myc mRNA occurred in nine of ten stage IV‐S tumors and did not correlate with the prognosis. The vanillylmandelic acid (VMA) to homovanillic acid (HVA) ratio was low in tumors with an increased number of copies of N‐myc. Serum lactate dehydrogenase (LDH) levels were increased in Stage IV‐S patients with N‐myc amplification but not in those with regressing tumors and without N‐myc amplification. These data suggest that N‐myc amplification may affect the final outcome in the patient classified as Stage IV‐S, but tumor regression can occur early after birth and appears to be independent of N‐myc amplification.

Mass screening for neuroblastoma at 6 months of age: Difficult to justify

BACKGROUND/PURPOSE A statistical analysis of the mass screening for neuroblastoma in Japan based on a population study rarely has been reported. This study aims to evaluate retrospectively the effectiveness of mass screening at 6 months of age using the available population data. METHODS The data on the neuroblastoma cases registered by the Committee for Pediatric Solid Malignant Tumors in the Kyushu area were analyzed based on both screened and unscreened populations in the Kyushu area. RESULTS From 1988 to 1992, the cumulative incidence of neuroblastoma in children less than 5 years of age was 82 in 484,599 for screened children, and 11 in 92,966 for unscreened children, respectively. Fourteen of the 82 screened patients had negative findings at 6 months of age (MS-negative cases). No significant difference was observed in the cumulative mortality rates from neuroblastoma in children younger than 5 years of age between the screened children and the unscreened children. Six of seven patients who died among the screened children were MS-negative cases with stage III or IV disease. In addition, no significant difference was found in the cumulative mortality rates from the neuroblastoma cases in patients less than 5 years of age between the children screened from 1988 to 1992 (7 of 484,599) and all children from 1980 to 1984 (14 of 668,084). CONCLUSIONS These findings suggests that the majority of the patients detected by mass screening had a favorable prognosis, and, mass screening in Japan for children less than 6 months of age was not observed to reduce the incidence and mortality from neuroblastoma. Therefore, mass screening at 6 months of age was not found to improve substantially the prognosis of patients with unfavorable neuroblastoma identified over 1 year of age, which is the primary purpose of such mass screening for neuroblastoma.

Severe neurotoxicities in a case of Charcot-Marie-Tooth disease type 2 caused by vincristine for acute lymphoblastic leukemia.

We report a 13-year-old male patient with Charcot-Marie-Tooth disease (CMT) type 2 who developed severe neuropathy because of vincristine (VCR) for his acute lymphoblastic leukemia. A clumsy gait, muscle weakness in his fingers, and inverted champagne bottlelike muscle in the lower limbs were noticed after remission induction treatment for acute lymphoblastic leukemia, which included VCR at a total dose of 8 mg/m2. An electrophysiologic study showed an almost normal median motor nerve conduction velocity (approximately 50 m/s), markedly reduced M-wave amplitude and sensory disturbance. He was diagnosed as CMT type 2 based on his symptoms and electrophysiologic findings. His symptoms gradually worsened, and even after VCR was discontinued, he could not walk alone for 7 months. VCR has previously been considered to be relatively safe in CMT type 2, however, some patients with CMT type 2 might show severe neurologic toxicities, as seen in patients with CMT type 1.

Mass screening for neuroblastoma: quo vadis? A 9-year experience from the Pediatric Oncology Study Group of the Kyushu area in Japan.

Since 1985, a nationwide program of mass screening for neuroblastoma has been available for 6-month-old infants throughout Japan. From 1985 to 1993, the authors studied 285 patients with neuroblastoma among their regional population of 15 million. There was an increase in the total number of patients per year in comparison to the previous 6-year period (1979 to 1984). However, no significant difference was noted in the number of patients older than 1 year or in the incidence of advanced-stage (stages III and IV) unscreened cases. The majority of neuroblastomas in the screened group showed favorable biological factors, even in the advanced stages. However, there was a small group with histologically and/or biologically unfavorable factors; five of 115 had amplified N-myc oncogene, four of 74 showed unfavorable Shimada histological findings, and three of 33 had an unfavorable DNA ploidy pattern. One case from this group with unfavorable factors died of the tumor. 3) Thirty-eight cases were negative at the time of mass screening, but later presented with neuroblastoma. Most of them were diagnosed between 1 and 3 years of age, and 30 of the 38 cases (78.9%) were advanced stage with unfavorable prognostic factors. Thus, the authors conclude that mass screening at 6 months can detect a selected population of infants with neuroblastoma; some of the tumors may represent subclinical masses destined for spontaneous regression. However, some tumors with unfavorable factors have been detected by mass screening before progression and/or dissemination. Infants in this group are considered to benefit most from early diagnosis and treatment.

Improved survival rates in children over 1 year of age with stage III or IV neuroblastoma following an intensive chemotherapeutic regimen

Combined chemotherapeutic regimens of (1) cyclophosphamide (40 mg/kg x two days), (2) cisplatinum (20 mg/m2 x five days) plus VM-26 (100 mg/m2), and (3) Adriamycin (60 mg/m2) plus DTIC (250 mg/m2 x five days) were prescribed for 42 Japanese children greater than 1 year of age with stage III or IV neuroblastoma. The protocol was separated into three forms (A, B, and C) from the combination pattern of three such high-dose courses. The cumulative survival rates for those with stages III and IV 48 months after initiation of therapy were 76.2% and 20.1%, respectively, and the overall rate was 36.7%. The tumor disappeared during the course of treatment in 25 of 42 children (59.5%). The 48-month survival rate was superior in patients greater than 5 years of age than younger patients (P less than .01). Even in patients with a tumor originating in the suprarenal region, the 48-month survival rate was 30.5%. Among 12 patients in whom the N-myc oncogene was measured, one of five with one to ten copies of amplification died, whereas all seven with greater than ten copies died or had a recurrence. Thus, the present chemotherapeutic regimens, in particular alternate administration of each high-dose course, considerably improved the survival of patients with stage III neuroblastoma. More aggressive protocols are needed for those with stage IV neuroblastoma who are greater than 1 year of age, particularly in those with an amplified N-myc oncogene.

Treatment responses of childhood aplastic anaemia with chromosomal aberrations at diagnosis

Summary.  The clinical outcome of childhood aplastic anaemia (AA) with aberrant cytogenetic clones at diagnosis was surveyed. Among 198 children with newly diagnosed AA registered with the AA Committee of the Japanese Society of Paediatric Hematology between 1994 and 1998, cytogenetic studies of bone marrow (BM) cells were completed in 159 patients. Apart from one Robertsonian translocation, seven patients (4·4%) showed clonal chromosomal abnormalities in hypoplastic BM without myelodysplastic features. The patients included six girls and one boy with a median age of 11 years (range 5–14 years). Sixpatients had del(6), del(5), del(13), del(20), or −7, and one showed add(9). Four patients responded to the first immunosuppressive therapy (IST: cyclosporin A plus anti‐thymocyte globulin) and one obtained a spontaneous remission. Cytogenetic abnormalities remained in two patients with an IST response. On the other hand, two patients showed no IST response. One did not respond to repeat IST and died of acute graft‐versus‐host disease after an unrelated‐BM transplant. Another obtained a complete response after a successful BM transplant. No haematological findings at diagnosis predicted the treatment response. No significant morphological changes developed during the course of the illness. A literature review revealed that half of 24 AA patients with chromosomal abnormalities responded to the first IST, and that +6 was the sole predictable marker for IST unresponsiveness. These results suggest that IST can be applied as the initial therapy for AA with cytogenetic abnormalities in the absence of completely matched donors.