Tomoko Yokosuka

Influence of CYP3A5 and ABCB1 gene polymorphisms on calcineurin inhibitor-related neurotoxicity after hematopoietic stem cell transplantation

Yanagimachi M, Naruto T, Tanoshima R, Kato H, Yokosuka T, Kajiwara R, Fujii H, Tanaka F, Goto H, Yagihashi T, Kosaki K, Yokota S. Influence of CYP3A5 and ABCB1 gene polymorphisms on calcineurin inhibitor‐related neurotoxicity after hematopoietic stem cell transplantation. 
Clin Transplant 2010: 24: 855–861.

RAG1 Deficiency May Present Clinically as Selective IgA Deficiency

BackgroundRecombination-activating gene (RAG) 1 and 2 deficiency is seen in patients with severe combined immunodeficiency (SCID) and Omenn syndrome. However, the spectrum of the disease has recently expanded to include a milder phenotype.ObjectiveWe analyzed a 4-year-old boy who was initially given the diagnosis of selective immunoglobulin A deficiency (SIgAD) based on immunoglobulin serum levels without any opportunistic infections, rashes, hepatosplenomegaly, autoimmunity or granulomas. The patient was found to be infected with varicella zoster; however, the clinical course was not serious. He produced antiviral antibodies.MethodsWe performed lymphocyte phenotyping, quantification of T cell receptor excision circles (TRECs) and kappa deleting recombination excision circles (KRECs), an analysis of target sequences of RAG1 and 2, a whole-genome SNP array, an in vitro V(D)J recombination assay, a spectratype analysis of the CDR3 region and a flow cytometric analysis of the bone marrow.ResultsLymphocyte phenotyping demonstrated that the ratio of CD4+ to CD8+ T cells was inverted and the majority of CD4+T cells expressed CD45RO antigens in addition to the almost complete lack of B cells. Furthermore, both TRECs and KRECs were absent. Targeted DNA sequencing and SNP array revealed that the patient carried a deletion of RAG1 and RAG2 genes on the paternally-derived chromosome 11, and two maternally-derived novel RAG1 missense mutations (E455K, R764H). In vitro analysis of recombination activity showed that both RAG1 mutant proteins had low, but residual function.ConclusionsThe current case further expands the phenotypic spectrum of mild presentations of RAG deficiency, and suggests that TRECs and KRECs are useful markers for detecting hidden severe, as well as mild, cases.

Chemo-sensitivity in a panel of B-cell precursor acute lymphoblastic leukemia cell lines, YCUB series, derived from children

Sensitivity to 10 anticancer drugs was evaluated in 6 childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL) cell lines. Authenticity of newly established cell lines was confirmed by genomic fingerprinting. The line YCUB-5R established at relapse was more resistant to 4-hydroperoxy-cyclophosphamide, cytarabine, L-asparaginase, topotecan, fludarabine, and etoposide than YCUB-5 from the same patient at diagnosis. Of the drugs tested, etoposide and SN-38 (irinotecan) showed highest efficacy in the panel, with 50% growth inhibition at 0.22-1.8 microg/ml and 0.57-3.6 ng/ml, respectively. This cell line panel offers an in vitro model for the development of new therapies for childhood BCP-ALL.

Suppressed neutrophil function in children with acute lymphoblastic leukemia

Infection is a major obstacle in cancer chemotherapy. Neutropenia has been considered to be the most important risk factor for severe infection; however, other factors, such as impaired neutrophil function, may be involved in susceptibility to infection in patients undergoing chemotherapy. In this study, we analyzed neutrophil function in children with acute lymphoblastic leukemia (ALL). Whole blood samples were obtained from 16 children with ALL at diagnosis, after induction chemotherapy, and after consolidation chemotherapy. Oxidative burst and phagocytic activity of neutrophils were analyzed by flow cytometry. Oxidative burst of neutrophils was impaired in ALL patients. The percentage of neutrophils with normal oxidative burst after PMA stimulation was 59.0 ± 13.2 or 70.0 ± 21.0% at diagnosis or after induction chemotherapy, respectively, which was significantly lower compared with 93.8 ± 6.1% in healthy control subjects (P = 0.00004, or 0.002, respectively); however, this value was normal after consolidation chemotherapy. No significant differences were noted in phagocytic activity in children with ALL compared with healthy control subjects. Impaired oxidative burst of neutrophils may be one risk factor for infections in children with ALL, especially in the initial periods of treatment.

Methylated chrysin reduced cell proliferation, but antagonized cytotoxicity of other anticancer drugs in acute lymphoblastic leukemia.

The efficacy of 5,7-dimethoxyflavone (DMF), a methylated analog of chrysin, as a therapeutic agent to treat acute lymphoblastic leukemia (ALL) was investigated. Using a panel of ALL cell lines, the IC50 (half-maximal inhibitory concentration) of DMF varied between 2.8 and 7.0 &mgr;g/ml. DMF induced G0/G1 cell cycle arrest, concomitant with a decreased expression of phosphorylated retinoblastoma-associated protein 1. DMF increased the rate of apoptosis, although it was apparent only after a long period of exposure (96 h). The accumulation of oxidative stress was not involved in the growth-inhibitory effects of DMF. As DMF reduced the intracellular levels of glutathione, the combination effects of DMF with other anticancer drugs were evaluated using the improved Isobologram and the combination index method. In the simultaneous drug combination assay, DMF antagonized the cytotoxicity of 4-hydroperoxy-cyclophosphamide, cytarabine, vincristine, and L-asparaginase in all tested ALL cells. This study demonstrated that DMF, a methylated flavone, was an effective chemotherapy agent that could inhibit cell cycle arrest and induce apoptosis in ALL cell lines. However, combination therapy with DMF and other anticancer drugs is not recommended.

Mandibular Ewing sarcoma with chromosomal translocation t(21;22)(q22;q12).

Ewing sarcoma (ES) is a primary bone malignant neoplasm and is the second most common primary malignancy of the bone found in childhood and adolescence after osteosarcoma. ES has an annual frequency in the population younger than 20 years of approximately 2.9 per million. ES occurs most frequently in the long bones of the extremities and pelvis and very rarely in the jaw. Recently, it was revealed that chromosomal translocation t(11;22)(q24;q12), which fuses the EWS gene on chromosome 22 and the FLI-1 gene on chromosome 11, occurs in most cases of ES. We report here a rare case of mandibular ES in a 10-year-old child with chromosomal translocation t(21;22)(q22;q12) in which the EWS gene is fused with the ERG gene on chromosome 21.

Kaposiform hemangioendothelioma infiltrates the gut wall: a rare case report.

yotype: 46,XY,t(5;15)(p15.3;q11.2)[3]/ 46,XY[17] (Fig. 1). The multiplex RT-PCR for recurrent genetic abnormalities showed negative results. The cerebrospinal fluid analysis showed no evidence of leukemic infiltration. The patient was treated again with CCG 106B protocol. Recently, he achieved remission after reinduction chemotherapy and was prepared for cord blood transplantation. Although we could not perform the cytogenetic analysis at initial diagnosis in this case, considering the similar immunophenotyping profiles and morphologies between the initial and the relapsed clones, the initial clone was expected to have the same cytogenetic abnormality with t(5;15)(p15.3;q11.2) of the relapsed one. Therefore, we could make a diagnosis in this patient with t(5;15) (p15.3;q11.2) at his age of 21 months. The previously described 7 infant patients in the literature with t(5;15) (p15;q11-13)1–3 were treated on intense protocols and they achieved complete remission (100%) and event-free survival (71%), with a relatively good prognosis, clearly less severe than the 11q23 rearrangement cases.5,6 Our case and Kwon and colleagues’4 case presented leukemia at the age of 21 months and 45 months, respectively, and showed the relatively late onset of the disease compared with the previously reported 7 infant cases.1–3 The case reported by Kwon and colleagues showed leukocytosis and a good response to steroid and continuous complete remission at 20 months, whereas our case revealed cytopenia and a failure to induction chemotherapy, and a recurrence of the disease, which means unfavorable treatment outcome compared with the previously reported ones.1–4 To our knowledge, this case with t(5;15)(p15;q11-q13) is the first report of refractoriness and recurrence of the disease, which means unfavorable treatment outcome and the second case of childhood onset after the one reported by Kwon and colleagues. Although this cytogenetic abnormality in infant ALL has been reported as a favorable prognostic indicator, this case suggests that the prognostic implication of t(5;15)(p15;q11-q13) in childhood ALL should be considered carefully. Larger studies would be required to confirm the prognostic significance of t(5;15)(p15;q11-q13), not only in infant ALL but also in childhood ALL. Seung Hee Lee, MD* In-Suk Kim, MD, PhD* So Eun Jun, MDw Young Tak Lim, MDw Eun Yup Lee, MD* Departments of *Laboratory Medicine wPediatrics, Pusan National University Busan, Korea

Clinical Courses of Two Pediatric Patients with Acute Megakaryoblastic Leukemia Harboring the CBFA2T3-GLIS2 Fusion Gene

Acute megakaryoblastic leukemia (AMKL) in children without Down syndrome (DS) has an extremely poor outcome with 3-year survival of less than 40%, whereas AMKL in children with DS has an excellent survival rate. Recently, a novel recurrent translocation involving CBFA2T3 and GLIS2 was identified in about 30% of children with non-DS AMKL, and the fusion gene was reported as a strong poor prognostic factor in pediatric AMKL. We report the difficult clinical courses of pediatric patients with AMKL harboring the CBFA2T3-GLIS2 fusion gene.

Philadelphia chromosome-positive acute lymphoblastic leukemia and Down syndrome.

Children with Down syndrome (DS) are at high risk of developing acute lymphoblastic leukemia (ALL), but Philadelphia chromosome‐positive (Ph+) ALL is rare in DS children. We report a case of Ph+ALL with DS complicated by chronic heart failure. Complete molecular remission was obtained after imatinib‐combined chemotherapy, although infectious episodes during the neutropenic period worsened the heart condition. After two courses of intensification chemotherapy, the patient underwent reduced intensity stem cell transplantation from an HLA‐identical sibling donor followed by post‐transplant imatinib. The patient maintained molecular complete remission for >2 years. This case report is the first description of the safe and effective use of imatinib for DS‐Ph+ALL. This case suggests the potential of molecular targeting therapy in DS‐ALL, which is often complicated by congenital diseases, and the importance of treating DS‐ALL while maintaining intensity and reducing treatment‐related toxicity.

Fulminant Epstein-Barr virus-driven CD8 positive T-cell lymphoproliferative disorder with chromosomal abnormality†

To the Editor: Epstein-Barr virus (EBV) is a member of the herpes virus family, with infections commonly occurring in early childhood. EBV mostly infects B cells, but it can also infect T or NK cells. T cells infected by EBV can cause EBV-associated T/ NK cell lymphoproliferative disorders (EBV T/NK LPD). Fulminant EBV T/NK LPD is one of the most aggressive types of EBV T/NK LPD, which often develops shortly after a primary EBV infection [1,2]. The pathogenesis of fulminant EBV-LPD remains unclear and the prognosis of this disorder is poor [1]. A 3-year-old female with no previous remarkable medical history and no apparent history of infectious mononucleosis was admitted with a 5-day history of fever, cervical lymphadenopathy, and hepatosplenomegaly. One week after admission, laboratory findings showed bicytopenia and hyperferritinemia. The EBV serology at the time revealed positive VCA-IgG and negative EBNA. EBV was proven to infect CD8 positive cells. Her condition did not improve despite the treatment with steroids, cyclosporine A, and VP-16. Two months after her admission, she was transferred for more intensive treatment. She had high-grade fever, hepatosplenomegaly, pancytopenia (WBC count 200/mm, hemoglobin 6.8 g/dl, and platelets 37,000/ mm), and high LDH with disseminated intravascular coagulation. Her bone marrow aspirate revealed a hypocellular marrow with abnormal lymphocytes and few hemophagocytosis. Quantitative PCR analysis of EBV showed 1.75 10 copies/mg DNA. Only eight cells were suitable for G-banding analysis of her bone marrow because of hypocellularity. 46XX, add (10)(p11.2), der(17;18)(q10;q10) was revealed in one cell (Fig. 1). Clonality could not be defintively analyzed because of the low number of cells available for analysis. Combination therapy with daunorubicin and cyclophosphamide was not effective. She died 2 weeks after being transferred to our institute. Ablinteractor-1 (Abi-1), located on 10p11.2, encodes an Ablbinding protein [3]. Abl-binding protein regulates the activity of ABL-1, a tyrosine kinase that is implicated in cell differentiation, cell division, cell adhesion, and stress response. The chromosomal abnormality, add (10)(p11.2), might involve the mechanism of abnormal proliferation of EBV infected CD8 positive cells. Translocation t(17;18)(q10;q10) has been seen in patients with chronic myelogenous leukemia, myelodysplastic syndrome, and acute myeloid leukemia [4], although its function has not been identified. More studies are necessary to clarify the pathogenesis of EBV T/NK LPD. It is possible that the chromosomal abnormality in this case brought about NK cell proliferation and might have resulted in fulminant EBV LPD.