Toshihiko Imamura

Frequent co-expression of HoxA9 and Meis1 genes in infant acute lymphoblastic leukaemia with MLL rearrangement

Summary. We studied the expression of HoxA9 and Meis1 by reverse transcriptase‐polymerase chain reaction analysis in leukaemic cells from cases of infant acute lymphoblastic leukaemia (ALL, n = 27) and childhood ALL (n = 29). These two genes were co‐expressed significantly more frequently in infant ALL than in childhood ALL (19/27 vs0/29 cases, P < 0·001) and were highly associated with MLL gene rearrangement in infant ALL cases (P < 0·001). These findings indicate that the HoxA9 and Meis1 genes are closely associated with MLL gene rearrangement in the development of infant ALL, which represents a distinct entity of childhood ALL.

Expression of the Ikaros gene family in childhood acute lymphoblastic leukaemia

Summary. The Ikaros (Ik) gene family, which includes Ik, Aiolos (Ai), and Helios (He), is a primary regulator of lymphocyte differentiation, and is involved in the development of acute lymphoblastic leukaemia (ALL). We analysed the expression of the Ik gene family isoforms in 97 ALL cases, consisting of 64 childhood and 33 infant ALL cases, using reverse transcription‐polymerase chain reaction (RT‐PCR). Expression of Ik was detected in all cases, 87 of which expressed either Ik1 or Ik2, or both, five of which expressed Ik1/Ik2 and Ik6, and another five of which expressed only Ik6. Therefore, the dominant negative isoform of Ik6 was expressed in 10 of the 38 cases of childhood precursor B ALL, but was absent in other types of childhood ALL (26·3%, χ2‐test, P = 0·0001). In terms of Aiolos and Helios expression, 49 (65·3%) out of the 75 and 40 (50%) out of the 80 ALL cases tested showed non‐spliced Ai1 and He1 respectively. Only one case of T lineage ALL expressed a small‐sized isoform of Helios (designated as He6). It was also found that the expression of Ai1 and He1 was low in Ik6‐positive patients (Fishers exact test; Ai1 P = 0·005, Hel P = 0·035).

Rearrangement of the MOZ gene in pediatric therapy‐related myelodysplastic syndrome with a novel chromosomal translocation t(2;8)(p23;p11)

In this study, we examined a pediatric case of therapy‐related myelodysplastic syndrome (tMDS). The symptoms developed 17 months after treatment for acute myeloblastic leukemia (AML, M2 subtype according to the French–American–British [FAB] classification) involving a chromosome abnormality at t(8;21)(q22;q22). Upon diagnosis of tMDS, spectral karyotyping analysis detected a new chromosomal translocation at t(2;8)(p23;p11.2). In addition, fluorescence in situ hybridization analysis suggested a rearrangement in the monocytic leukemia zinc finger (MOZ) gene, located in the 8p11 region of chromosome 8. However, no partner gene on 2p23 could be identified. To our knowledge, this is the first report of tMDS associated with a rearrangement of the MOZ gene. MOZ‐linked fusion proteins such as MOZ‐CBP (CREB binding protein), MOZ‐TIF2 (transcriptional intermediary factor 2), and MOZ‐p300 (adenoviral E1A‐associated protein) are associated with AML chromosomal abnormalities at t(8;16)(p11;p13), inv(8)(p11q13), and t(8;22)(p11;q13), respectively, and are thought to account for leukemogenesis occurring through the aberrant regulation of histone acetylation. Through a similar mechanism, we believe that MOZ, fused to an unidentified partner gene at 2p23, may have caused an alteration in histone acetylation, resulting in the development of tMDS in this patient.

IKZF1 deletion is associated with a poor outcome in pediatric B-cell precursor acute lymphoblastic leukemia in Japan

Genetic alterations of Ikaros family zinc finger protein 1 (IKZF1), point mutations in Janus kinase 2 (JAK2), and overexpression of cytokine receptor‐like factor 2 (CRLF2) were recently reported to be associated with poor outcomes in pediatric B‐cell precursor (BCP)‐ALL. Herein, we conducted genetic analyses of IKZF1 deletion, point mutation of JAK2 exon 16, 17, and 21, CRLF2 expression, the presence of P2RY8‐CRLF2 fusion and F232C mutation in CRLF2 in 202 pediatric BCP‐ALL patients newly diagnosed and registered in Japan Childhood Leukemia Study ALL02 protocol to find out if alterations in these genes are determinants of poor outcome. All patients showed good response to initial prednisolone (PSL) treatment. Ph+, infantile, and Down syndrome–associated ALL were excluded. Deletion of IKZF1 occurred in 19/202 patients (9.4%) and CRLF2 overexpression occurred in 16/107 (15.0%), which are similar to previous reports. Patients with IKZF1 deletion had reduced event‐free survival (EFS) and overall survival (OS) compared to those in patients without IKZF1 deletion (5‐year EFS, 62.7% vs. 88.8%, 5‐year OS, 71.8% vs. 90.2%). Our data also showed significantly inferior 5‐year EFS (48.6% vs. 84.7%, log rank P = 0.0003) and 5‐year OS (62.3% vs. 85.4%, log rank P = 0.009) in NCI‐HR patients (n = 97). JAK2 mutations and P2RY8‐CRLF2 fusion were rarely detected. IKZF1 deletion was identified as adverse prognostic factor even in pediatric BCP‐ALL in NCI‐HR showing good response to PSL.

Sustained elevation of serum interleukin-18 and its association with hemophagocytic lymphohistiocytosis in XIAP deficiency

X-linked lymphoproliferative syndrome (XLP) is a rare primary immunodeficiency characterized by increased vulnerability to Epstein-Barr virus infection. XLP type 1 is caused by mutations in SH2D1A, whereas X-linked inhibitor of apoptosis (XIAP) encoded by XIAP/BIRC4 is mutated in XLP type 2. In XIAP deficiency, hemophagocytic lymphohistiocytosis (HLH) occurs more frequently and recurrence is common. However, the underlying mechanisms remain mostly unknown. We describe the characteristics of the cytokine profiles of serum samples from 10 XIAP-deficient patients. The concentration of interleukin (IL)-18 was strikingly elevated in the patients presented with HLH, and remained high after the recovery from HLH although levels of other pro-inflammatory cytokines approached the normal range. Longitudinal examination of two patients demonstrated marked exacerbation of IL-18 levels during every occasion of HLH. These findings may suggest the association between HLH susceptibility and high serum IL-18 levels in XIAP deficiency.

Circulating Methylated-DCR2 Gene in Serum as an Indicator of Prognosis and Therapeutic Efficacy in Patients with MYCN Nonamplified Neuroblastoma

Background:MYCN amplification (MNA) in neuroblastoma is a strong indicator of poor prognosis. However, some MYCN nonamplified (non-MNA) cases show poor outcomes, and examining the status of the gene requires an operation, which may have surgical complications. Therefore, a new marker is needed to identify cases of non-MNA neuroblastomas with poor prognoses using less risky procedures. Aberrant hypermethylation of the DCR2 promoter has recently been associated with rapidly progressing neuroblastoma. We aimed to develop a noninvasive DCR2 methylation assay for patients with neuroblastoma using serum DNA, which predominantly originates from tumor-released DNA. Methods: Using DNA-based real-time PCR, we simultaneously quantified a methylated-DCR2 specific sequence (M) and a reference sequence (R) located in the promoter region in serum DNA, and evaluated DCR2 methylation status as M/R ratios in 86 patients with neuroblastoma. Results: Serum DCR2 M/R ratios were strongly correlated with those in the tumor (r = 0.67; P = 0.002). DCR2 methylation was associated with stage both in the whole neuroblastoma group and in the non-MNA group (P < 0.001), and DCR2-methylated patients showed significantly poorer 5-year event-free survival in the whole neuroblastoma group (43% versus 84%; P < 0.001), especially in the non-MNA group (12% versus 96%;P < 0.001). Among five DCR2-methylated patients whose clinical courses were followed, serum M/R ratios were close to 0 in the patients in remission, whereas the ratios increased in patients who relapsed. Conclusions: Detection of methylated-DCR2 in serum DNA has promise as a noninvasive assay for predicting prognosis and therapeutic efficacy in neuroblastoma, especially in non-MNA cases. Furthermore, it might be a sensitive marker of tumor recurrence in DCR2-methylated cases.

Potential use of procalcitonin concentrations as a diagnostic marker of the PFAPA syndrome

PFAPA syndrome is a clinical entity of unknown etiology characterized by periodic episodes of high fever accompanied by aphthous stomatitis, pharyngitis/tonsillitis, and cervical adenitis [3, 5]. Since specific laboratory abnormalities for the PFAPA syndrome are inexistent, it is usually diagnosed clinically after excluding other probable causes of the fever, such as infection [1]. In PFAPA patients, discriminating between a fever attack due to bacterial infection and a fever attack due to noninfectious inflammation constitutes a major difficulty. Because procalcitonin, a propeptide of calcitonin, is reported to be a sensitive marker of systemic bacterial infection [2, 4], we followed peripheral leukocyte counts, CRP values and procalcitonin concentrations during the fever attacks associated with PFAPA syndrome in the hope of defining reliable criteria for its diagnosis. We determined serum procalcitonin concentrations in six PFAPA syndrome patients (two males and four females) with a median age of 7.5 (range 3–10) years and in 32 controls (bacterial, n=10 and non-bacterial, n=22). Sampling was performed on the third to fifth day of fever. In the PFAPA syndrome patients, febrile episodes started at the median age of 2.5 (range 1–7) years with each episode lasting 5–7 days and recurring every 3–4 weeks. The ethical committees of our institutes approved the study protocol and the guardians of all the patients gave their informed consent. Serum procalcitonin concentrations were measured by using the fully automated enzyme immunoluminescent assay (Wako Pure Chemical Industries, Ltd.), which employs katacalcin monoclonal antibody and calcitonin polyclonal antibody labeled with peroxidase for SphereLight 180 (Olympus Corporation). The detection limit was 0.1 ng/ml and the normal reference was set at <0.5 ng/ml. In PFAPA patients, the correlations between procalcitonin, CRP values and leukocyte counts were examined over 13 febrile episodes. Serum procalcitonin values ranged from 0.20 to 11.36 (median value 1.05) ng/ml in positive control subjects (Table 1), while all the negative controls had undetectable levels. During febrile episodes in PFAPA patients, which were confirmed not to be due to adenoviral or group A streptococcal infections, leukocyte counts and serum concentrations of CRP were invariably and significantly Eur J Pediatr (2007) 166:621–622 DOI 10.1007/s00431-006-0281-2

Comprehensive analyses and characterization of haemophagocytic lymphohistiocytosis in Vietnamese children

Haemophagocytic lymphohistiocytosis (HLH) is a fatal haematological disorder with diverse aetiology. This prospective study was undertaken to characterize HLH cases in Vietnamese children. Clinical and laboratory data, genetic analyses and outcome of the HLH patients were analysed. A total of 33 patients were enrolled from March 2007 to December 2008, with a median age of 3 years. Mutations of the SH2D1A (SAP) and PRF1 genes were detected in one patient, respectively. The virus association was high, up to 63·6% (21/33), including Epstein–Barr virus (19/33), cytomegalovirus (2/33) and dengue virus (2/33). Five patients had malignant lymphoma and two had autoimmune diseases. Twenty‐eight patients were treated according to the HLH‐2004 protocol. The first response rate was 64·3% (18/28), with an early death rate of 35·7% (10/28). High levels of interferon‐γ, interleukin‐10, MIG and interferon‐inducible protein‐10 (IP‐10) were associated with early mortality (P < 0·05). Reactivation among the responders was high (9/18) and the uneventful resolution was low (3/18) after a median follow‐up of 35 weeks. In conclusion, the majority of HLH cases are associated with virus infections in Vietnamese children. Familial HLH is rare. The frequent reactivation and high mortality demands a more appropriate therapeutic regimen in tropical areas like Vietnam.

Recent advances in Langerhans cell histiocytosis

The purpose of this review is to provide an updated overview of the pathogenesis and treatment of Langerhans cell histiocytosis (LCH). The pathogenesis of LCH remains obscure and the optimal treatment for LCH has not been established, although incremental progress has been made. Proinflammatory cytokines and chemokines are known to play a role in LCH, which suggests that LCH is an immune disorder. However, the oncogenic BRAF mutation is also detected in more than half of LCH patients, which suggests that LCH is a neoplastic disorder. Remaining major issues in the treatment of LCH are how to rescue patients who have risk‐organ involvement but do not respond to first‐line therapy, the optimal treatment for the orphan disease of multifocal adult LCH, and how to reduce and treat central nervous system‐related consequences, such as central diabetes insipidus and neurodegeneration. More research is needed to better understand the pathogenesis of this disease and to resolve the treatment issues.

Langerhans cell histiocytosis with multifocal bone lesions: comparative clinical features between single and multi-systems

Langerhans cell histiocytosis (LCH) can be a single system or multi-system disease. Both disease types can be associated with multi-focal bone lesions, but their bone involvement patterns have not been compared systematically. Of the new pediatric LCH cases enrolled into the JLSG-02 study during 2002–2007, 67 cases of single system multifocal bone (SMFB) LCH and 97 cases of multi-system bone (MSB) LCH were analyzed to determine if the bone involvement patterns differ in these two types, and whether these differences correlate with outcome. Statistical analysis was performed with Mann–Whitney U test, Fisher’s exact test, and other measures. Onset ages were higher for SMFB (P < 0.001), but the two types did not differ in the number of bone lesions per patient. The skull was most frequently affected in both types, followed by the spine. Lesions in the temporal bone (P = 0.002), ear-petrous bone (P < 0.001), orbita (P = 0.003), and zygomatic bone (P = 0.016) were significantly more common in MSB. The two types did not differ in response to treatment, but MSB was associated with a significantly higher incidence of diabetes insipidus (DI) (P < 0.001). Novel measures are required in preventing the development of DI in MSB-type LCH patients with “risk” bone lesions.