Publications by authors named "Makito Tanaka"

27 Publications

  • Page 1 of 1

An aggressive systemic mastocytosis preceded by ovarian dysgerminoma.

BMC Cancer 2020 Nov 27;20(1):1162. Epub 2020 Nov 27.

Division of Molecular Genetics, Institute for Comprehensive Medical Science, Fujita Health University, 1-98 Dengakugakubo, Kutsukake-cho, Toyoake, Aichi, 470-1192, Japan.

Background: Aggressive systemic mastocytosis (ASM) is a rare malignant disease characterized by disordered mast cell accumulation in various organs. We here describe a female ASM patient with a previous history of ovarian dysgerminoma.

Methods: Molecular cytogenomic analyses were performed to elucidate an etiological link between the ASM and dysgerminoma of the patient.

Results: This patient was affected by ovarian dysgerminoma which was treated by chemotherapy and surgical resection. Having subsequently been in complete remission for 2 years, she developed symptoms of ASM. A somatic D816A mutation in the KIT gene was detected in her bone marrow, which facilitated the diagnosis of ASM. Unexpectedly, this KIT D816A variant was also detected in the prior ovarian dysgerminoma sample. Whole-exome sequencing allowed us to identify a somatic nonsense mutation of the TP53 gene in the bone marrow, but not in the dysgerminoma. Microarray analysis of the patient's bone marrow revealed a copy-number-neutral loss of heterozygosity at the TP53 locus, suggestive of the homozygous nonsense mutation in the TP53 gene. In addition, the loss of heterozygosity at the TP53 locus was also detected in the dysgerminoma.

Conclusions: These results indicated that either the mast cells causing the ASM in this case had originated from the preceding ovarian dysgerminoma as a clonal evolution of a residual tumor cell, which acquired the TP53 mutation, or that both tumors developed from a common cancer stem cell carrying the KIT D816A variation.
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http://dx.doi.org/10.1186/s12885-020-07653-zDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7693501PMC
November 2020

Frequency of subclinical herpes zoster in pediatric hematology-oncology patients receiving chemotherapy: A retrospective cohort analysis.

J Med Virol 2020 08 18;92(8):1260-1265. Epub 2019 Dec 18.

Department of Pediatrics, Fujita Health University School of Medicine, Toyoake, Aichi, Japan.

Varicella-zoster virus (VZV) reactivation from the enteric nervous system can cause ileus (Ogilvie's syndrome) in adult patients. Since no pediatric cases have been described, we sought to retrospectively analyze VZV reactivation in pediatric hematology-oncology patients to determine whether VZV infection including subclinical VZV reactivation can induce gastrointestinal complications such as Ogilvie's syndrome. Thirty-five patients who received chemotherapy at our institution between September 2013 and June 2018 were included. Serum samples were collected weekly during hospitalization and every 3 months during outpatient maintenance chemotherapy. A real-time polymerase chain reaction assay was used to measure VZV DNA load in serum. The clinical features of patients with VZV infection were retrospectively analyzed. Of 1165 serum samples, 7 (0.6%) were positive for VZV DNA. VZV DNA was detected in 3 of 35 patients. In patient A, VZV DNA was detected during two episodes. The first episode involved varicella-like eruptions caused by the Oka VZV vaccine strain. The second episode involved herpes zoster (HZ) caused by the same strain. Patients B and C had a clinical course that was typical for HZ caused by wild-type VZV. No gastrointestinal symptoms were observed at the time of VZV infection in these three patients. VZV DNA was not detected in any other samples. No pediatric cases with Ogilvie's syndrome caused by VZV reactivation were demonstrated in this cohort. Additionally, no subclinical VZV reactivation was found in this cohort. Further study is needed to elucidate the precise incidence of pediatric Ogilvie's syndrome caused by VZV reactivation.
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http://dx.doi.org/10.1002/jmv.25650DOI Listing
August 2020

Human herpesvirus-6B infection in pediatric allogenic hematopoietic stem cell transplant patients: Risk factors and encephalitis.

Transpl Infect Dis 2020 Feb 11;22(1):e13203. Epub 2019 Nov 11.

Department of Pediatrics, Fujita Health University School of Medicine, Toyoake, Japan.

Background: Human herpesvirus-6B (HHV-6B) infection after allogenic hematopoietic stem cell transplantation (allo-HSCT) is known to be associated with post-transplant limbic encephalitis in adults. Meanwhile, the association between HHV-6B infection and central nervous system complications remains unclear in pediatric allo-HSCT patients.

Methods: In this study, HHV-6B infection was monitored for more than 50 days after HSCT using virus isolation and real-time PCR. Clinical information such as patient background and encephalitis status was collected retrospectively from medical records. Risk factors for HHV-6B infection were determined by the Cox proportional hazards model, and the clinical features of HHV-6B encephalitis in pediatric allo-HSCT patients were elucidated.

Results: Human herpesvirus-6B infection was observed in 74 (33.8%) of 219 patients at 3-47 days (median 18, interquartile range 13-20). Risk factors identified in multivariable analysis were hematological malignancy (hazards ratio [HR], 5.0; 95% confidence interval [CI], 2.3/12.5; P < .0001), solid tumor (HR, 4.8; CI, 1.5/16.3; P = .0104), unrelated donor (HR, 2.1; CI, 1.0/4.6; P = .0378), and sex-mismatched donor (HR 1.8; CI, 1.1/3.0; P = .0257). HHV-6B encephalitis occurred in only one of the 219 patients (0.46%); this patient demonstrated the typical clinical course of posterior reversible encephalopathy syndrome.

Conclusion: Hematological malignancy, solid tumor, unrelated donor, and sex-mismatched donor were significant risk factors for HHV-6B infection after pediatric allo-HSCT. In pediatric allo-HSCT patients, the incidence of HHV-6B encephalitis was low and the clinical features differed from those in adult patients.
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http://dx.doi.org/10.1111/tid.13203DOI Listing
February 2020

Clonality investigation of clinical Escherichia coli isolates by polymerase chain reaction-based open-reading frame typing method.

J Infect Chemother 2020 Jan 27;26(1):38-42. Epub 2019 Jul 27.

Division of Laboratory Medicine, Sapporo Medical University Hospital, Japan; Division of Infection Control, Sapporo Medical University Hospital, Japan; Department of Infection Control and Laboratory Medicine, Sapporo Medical University School of Medicine, Japan. Electronic address:

Escherichia coli (E. coli) causes urinary tract infections, pneumonia, surgical site infections, and bloodstream infections and is the important pathogen for both community-acquired and healthcare-associated infections. To investigate the clonality of E. coli is important for infection control and prevention. We aimed to investigate the clonality of clinical E. coli isolates using Cica Geneus E. coli polymerase chain reaction (PCR)-based open-reading frame typing (POT) KIT and clarify the clinical usefulness of this kit. About 124 E. coli isolates obtained from inpatients at Sapporo Medical University Hospital were used. The POT method was used to classify 124 clinical isolates into 87 POT numbers. In addition to the clonality, it was possible to obtain additional information that 20 of the 124 isolates were extended-spectrum β-lactamase (ESBL) producing E. coli (5 isolates of CTX-M-1 group and 15 isolates of CTX-M-9 group) and 13 were sequence type (ST) 131 clone. Furthermore, when these ESBL-producing 20 isolates were compared with pulsed-field gel electrophoresis (PFGE) or multilocus sequence typing (MLST), Simpson's index of diversity was 0.968 in POT method, 0.979 in PFGE, and 0.584 in MLST. POT method had an analytical power similar to that of PFGE. In conclusion, attention should be paid to the difference in the interpretation of the results between the POT method and the PFGE, but POT method may be useful to timely monitor the spread of E. coli in medical facilities.
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http://dx.doi.org/10.1016/j.jiac.2019.06.014DOI Listing
January 2020

Late-phase human herpesvirus 6B reactivation in hematopoietic stem cell transplant recipients.

Transpl Infect Dis 2018 Aug 13;20(4):e12916. Epub 2018 Jun 13.

Department of Pediatrics, Fujita Health University School of Medicine, Toyoake, Japan.

Background: We sought to determine whether late-phase human herpesvirus 6B (HHV-6B) infection in hematopoietic stem cell transplant (HSCT) recipients was associated with serious outcomes and mortality.

Methods: The occurrence and course of HHV-6B infection was monitored for at least 60 days after transplant using virus isolation and real-time polymerase chain reaction. Risk factors for late-phase HHV-6B infection were examined, and the propensity score was calculated with significant risk factors. The inverse probability-weighted multivariable logistic regression analysis was performed to estimate odds ratios (ORs) and the 95% confidence intervals (95% CI) for mortality.

Results: Late-phase HHV-6B infection was observed in 12/89 (13.5%) of the HSCT recipients. Older age (OR: 10.3, 95% CI: 2.1/72.9, P = .0027), hematologic malignancy (OR: 10.3, 95% CI: 1.8/97.1, P = .0063), unrelated donor transplantation (OR: 5.3, 95% CI: 1.1/36.0, P = .0345), and sex-mismatched donor transplantation (OR: 6.3, 95% CI: 1.4/39.5, P = .0149) were identified as risk factors for late-phase HHV-6B infection. Fifteen subjects died (17%). Inverse probability-weighted multivariable logistic model analysis revealed that late-phase HHV-6B infection was an independent risk factor for mortality (OR: 4.2, 95% CI: 1.7/11.0, P = .0012). Among 5 of the fatal cases of late-phase HHV-6B infection, viral infection might be associated with severe clinical manifestations.

Conclusion: Late-phase HHV-6B infection in HSCT recipients was associated with worse outcomes. The full spectrum of clinical features of the infection has not been fully elucidated, and therefore, recipients with high-risk factors for late-phase HHV-6B infection should be carefully monitored.
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http://dx.doi.org/10.1111/tid.12916DOI Listing
August 2018

HLA-DP constitutively presents endogenous peptides generated by the class I antigen processing pathway.

Nat Commun 2017 05 10;8:15244. Epub 2017 May 10.

Tumor Immunotherapy Program, Campbell Family Institute for Breast Cancer Research, Campbell Family Cancer Research Institute, Princess Margaret Cancer Centre, University Health Network, Toronto, Ontario, Canada M5G 2M9.

Classical antigen processing leads to the presentation of antigenic peptides derived from endogenous and exogenous sources for MHC class I and class II molecules, respectively. Here we show that, unlike other class II molecules, prevalent HLA-DP molecules with β-chains encoding Gly84 (DP) constitutively present endogenous peptides. DP does not bind invariant chain (Ii) via the class II-associated invariant chain peptide (CLIP) region, nor does it present CLIP. However, Ii does facilitate the transport of DP from the endoplasmic reticulum (ER) to the endosomal/lysosomal pathway by transiently binding DP via a non-CLIP region(s) in a pH-sensitive manner. Accordingly, like class I, DP constitutively presents endogenous peptides processed by the proteasome and transported to the ER by the transporter associated with antigen processing (TAP). Therefore, DP, found only in common chimpanzees and humans, uniquely uses both class I and II antigen-processing pathways to present peptides derived from intracellular and extracellular sources.
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http://dx.doi.org/10.1038/ncomms15244DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5436232PMC
May 2017

[The Study of a New Formula for Correcting Serum Calcium Levels, Considering the Difference in Albumin Analysis Methods].

Rinsho Byori 2016 06;64(6):619-624

In Japan, Payne's formula [corrected Ca=total Ca+ (4-ALB)] has been used to correct serum calcium concentration levels. However, the current methods for measuring calcium and albumin differ from those that were used to establish the Payne's formula. For albumin measurement, particularly, values differ be- tween BCG and improved BCP method. In 2014, Ohba et al. reported a modified formula [corrected Ca= total Ca+0.7X (4-ALB)], which is more suitable for correcting calcium with the improved BCP method than with the Payne's method. In the same year, the recommendation for converting albumin concentration with the improved BCP method to that with the BCG method was presented by the Japanese Society of La- boratory Medicine. Thus, we conceived a new modified formula [corrected Ca=total Ca+ {4- (BCP+ 0.3) }], which is included in the contents of this recommendation, and examined the effects of this formula in comparison with the Payne and Ohba methods. The patients recruited for the calcium adjustment were as follows: (i) all patients; (ii) patients with albumin concentrations <4.0 g/dL; and (iii) patients with albumin concentrations ≤3.5 g/dL. Payne's formula overcorrected calcium with the improved BCP method. Ohba's method was suitable for (i), while the new for- mula was specifically more suitable for (iii) than the Payne and Ohba formulas. The present study showed that our new formula is more suitable for calcium adjustment in patients with albumin concentrations ≤3.5 g/dL. [Original].
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June 2016

CDR3β sequence motifs regulate autoreactivity of human invariant NKT cell receptors.

J Autoimmun 2016 Apr 31;68:39-51. Epub 2015 Dec 31.

Tumor Immunotherapy Program, Campbell Family Institute for Breast Cancer Research, Campbell Family Cancer Research Institute, Princess Margaret Cancer Centre, University Health Network, Toronto, ON, M5G 2M9, Canada; Department of Immunology, University of Toronto, Toronto, ON, M5S 1A8, Canada. Electronic address:

Invariant natural killer T (iNKT) cells are a subset of T lymphocytes that recognize lipid ligands presented by monomorphic CD1d. Human iNKT T cell receptor (TCR) is largely composed of invariant Vα24 (Vα24i) TCRα chain and semi-variant Vβ11 TCRβ chain, where complementarity-determining region (CDR)3β is the sole variable region. One of the characteristic features of iNKT cells is that they retain autoreactivity even after the thymic selection. However, the molecular features of human iNKT TCR CDR3β sequences that regulate autoreactivity remain unknown. Since the numbers of iNKT cells with detectable autoreactivity in peripheral blood is limited, we introduced the Vα24i gene into peripheral T cells and generated a de novo human iNKT TCR repertoire. By stimulating the transfected T cells with artificial antigen presenting cells (aAPCs) presenting self-ligands, we enriched strongly autoreactive iNKT TCRs and isolated a large panel of human iNKT TCRs with a broad range autoreactivity. From this panel of unique iNKT TCRs, we deciphered three CDR3β sequence motifs frequently encoded by strongly-autoreactive iNKT TCRs: a VD region with 2 or more acidic amino acids, usage of the Jβ2-5 allele, and a CDR3β region of 13 amino acids in length. iNKT TCRs encoding 2 or 3 sequence motifs also exhibit higher autoreactivity than those encoding 0 or 1 motifs. These data facilitate our understanding of the molecular basis for human iNKT cell autoreactivity involved in immune responses associated with human disease.
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http://dx.doi.org/10.1016/j.jaut.2015.12.005DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4792736PMC
April 2016

Correlation of CYP2C19 phenotype with voriconazole plasma concentration in children.

J Pediatr Hematol Oncol 2013 Jul;35(5):e219-23

Department of Pediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Background: Voriconazole is a triazole antifungal agent with potent activity against a broad spectrum of pathogens, including Aspergillus and Candida species. In human adults, allelic polymorphisms of CYP2C19 are known to correlate with significant variation in voriconazole plasma concentration. Here, we report an analysis of CYP2C19 phenotype and voriconazole plasma concentrations in children.

Methods: This retrospective study included 37 children who had voriconazole plasma concentrations measured from May 2006 to June 2011. All had single-nucleotide polymorphisms that define the 3 major CYP2C19 alleles. Patients were classified as follows: normal metabolizers, intermediate metabolizers, poor metabolizers, or hypermetabolizers.

Results: The frequencies of the 3 CYP2C19 genetic polymorphisms were similar to those previously reported for Japanese adults. Trough plasma concentrations of voriconazole were significantly higher in the poor metabolizer and intermediate metabolizer groups compared with the normal metabolizer and hypermetabolizer groups (P=0.004). Two patients with high plasma concentrations experienced voriconazole-related severe adverse events (syndrome of inappropriate antidiuretic hormone secretion and cardiac toxicities).

Conclusions: The current study suggests that a significant association exists in children between the voriconazole plasma concentration and the CYP2C19 phenotype. Dose adjustment based on CYP2C19 phenotype may be useful during voriconazole therapy, especially for Japanese children, who as a group have a higher incidence of the poor metabolizer and intermediate metabolizer phenotypes.
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http://dx.doi.org/10.1097/MPH.0b013e3182880eaaDOI Listing
July 2013

Somatic mosaicism for oncogenic NRAS mutations in juvenile myelomonocytic leukemia.

Blood 2012 Aug 2;120(7):1485-8. Epub 2012 Jul 2.

Department of Pediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Juvenile myelomonocytic leukemia (JMML) is a rare pediatric myeloid neoplasm characterized by excessive proliferation of myelomonocytic cells. Somatic mutations in genes involved in GM-CSF signal transduction, such as NRAS, KRAS, PTPN11, NF1, and CBL, have been identified in more than 70% of children with JMML. In the present study, we report 2 patients with somatic mosaicism for oncogenic NRAS mutations (G12D and G12S) associated with the development of JMML. The mutated allele frequencies quantified by pyrosequencing were various and ranged from 3%-50% in BM and other somatic cells (ie, buccal smear cells, hair bulbs, or nails). Both patients experienced spontaneous improvement of clinical symptoms and leukocytosis due to JMML without hematopoietic stem cell transplantation. These patients are the first reported to have somatic mosaicism for oncogenic NRAS mutations. The clinical course of these patients suggests that NRAS mosaicism may be associated with a mild disease phenotype in JMML.
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http://dx.doi.org/10.1182/blood-2012-02-406090DOI Listing
August 2012

De novo childhood myelodysplastic/myeloproliferative disease with unique molecular characteristics.

Br J Haematol 2012 Jul 10;158(1):129-37. Epub 2012 May 10.

Department of Paediatrics, Nagoya University Graduate School of Medicine, Nagoya, Aichi, Japan.

Myelodysplastic/myeloproliferative uclassifiable (MDS/MPN-U) is a rare myeloid neoplasm characterized by myelodysplasia and myeloproliferation at the time of initial presentation, which is usually a diagnosis of exclusion. The molecular pathogenesis of MDS/MPN-U patients remains to be elucidated. Among five patients diagnosed with MDS/MPN-U, three patients harboured RUNX1 (AML1) mutations; one carried somatic mosaicism of RUNX1 mutation with JAK2(V617F) mutation and one had dual RUNX1 and FLT3-internal tandem duplication mutations with progression to acute myeloid leukaemia (AML). Germline mutation of TP53 was detected as a sole genetic lesion in one patient. JAK2(V617F) and somatic mosaicism of KRAS and TET2 mutations co-existed in one patient. Otherwise, no alterations were detected in PTPN11, NRAS, CBL and ASXL1 genes. ETV6-PDGFRB fusion transcript was not detected in all patients. Four patients recieved haematopoietic stem cell transplantation (HSCT); three patients relapsed and one achieved complete remission after three donor lymphocyte infusions. Our findings suggest that the mutational spectrum observed in childhood MDS/MPN-U is quite different from that seen in juvenile myelomonocytic leukaemia and, to some extent, resemble chronic myelomonocytic leukaemia. Moreover, two patients had constitutional alterations of genes frequently found in AML. Further investigations are required to define the roles of these genetic alterations in the pathogenesis of childhood MDS/MPN-U.
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http://dx.doi.org/10.1111/j.1365-2141.2012.09140.xDOI Listing
July 2012

Excellent outcome of allogeneic bone marrow transplantation for Fanconi anemia using fludarabine-based reduced-intensity conditioning regimen.

Int J Hematol 2012 Jun 22;95(6):675-9. Epub 2012 Apr 22.

Department of Pediatrics, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, Aichi 466-8550, Japan.

Fanconi anemia (FA) is a disorder characterized by developmental anomalies, bone marrow failure and a predisposition to malignancy. It has recently been shown that hematopoietic stem cell transplantation using fludarabine (FLU)-based reduced-intensity conditioning is an efficient and quite safe therapeutic modality. We retrospectively analyzed the outcome of bone marrow transplantation (BMT) in eight patients with FA performed in two institutes between 2001 and 2011. There were seven females and one male with a median age at diagnosis = 4.5 years (range 2-12 years). The constitutional characteristics associated with FA, such as developmental anomalies, short stature and skin pigmentation, were absent in three of the patients. One patient showed myelodysplastic features at the time of BMT. All patients received BMT using FLU, cyclophosphamide (CY) and rabbit anti-thymocyte globulin (ATG) either from a related donor (n = 4) or an unrelated donor (n = 4). Acute graft-versus-host disease (GVHD) of grade I developed in one patient, while chronic GVHD was not observed in any patient. All patients are alive and achieved hematopoietic recovery at a median follow-up of 72 months (range 4-117 months). BMT using FLU/low-dose CY/ATG -based regimens regardless to the donor is a beneficial therapeutic approach for FA patients.
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http://dx.doi.org/10.1007/s12185-012-1079-9DOI Listing
June 2012

Ex vivo expansion of human CD8+ T cells using autologous CD4+ T cell help.

PLoS One 2012 12;7(1):e30229. Epub 2012 Jan 12.

Department of Medical Oncology, Dana-Farber Cancer Institute, Massachusetts, United States of America.

Background: Using in vivo mouse models, the mechanisms of CD4+ T cell help have been intensively investigated. However, a mechanistic analysis of human CD4+ T cell help is largely lacking. Our goal was to elucidate the mechanisms of human CD4+ T cell help of CD8+ T cell proliferation using a novel in vitro model.

Methods/principal Findings: We developed a genetically engineered novel human cell-based artificial APC, aAPC/mOKT3, which expresses a membranous form of the anti-CD3 monoclonal antibody OKT3 as well as other immune accessory molecules. Without requiring the addition of allogeneic feeder cells, aAPC/mOKT3 enabled the expansion of both peripheral and tumor-infiltrating T cells, regardless of HLA-restriction. Stimulation with aAPC/mOKT3 did not expand Foxp3+ regulatory T cells, and expanded tumor infiltrating lymphocytes predominantly secreted Th1-type cytokines, interferon-γ and IL-2. In this aAPC-based system, the presence of autologous CD4+ T cells was associated with significantly improved CD8+ T cell expansion in vitro. The CD4+ T cell derived cytokines IL-2 and IL-21 were necessary but not sufficient for this effect. However, CD4+ T cell help of CD8+ T cell proliferation was partially recapitulated by both adding IL-2/IL-21 and by upregulation of IL-21 receptor on CD8+ T cells.

Conclusions: We have developed an in vitro model that advances our understanding of the immunobiology of human CD4+ T cell help of CD8+ T cells. Our data suggests that human CD4+ T cell help can be leveraged to expand CD8+ T cells in vitro.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0030229PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3257268PMC
July 2012

IL-21 can supplement suboptimal Lck-independent MAPK activation in a STAT-3-dependent manner in human CD8(+) T cells.

J Immunol 2012 Feb 11;188(4):1609-19. Epub 2012 Jan 11.

Department of Medical Oncology, Dana-Farber Cancer Institute, Boston, MA 02215, USA.

Although both MHC class II/CD8α double-knockout and CD8β null mice show a defect in the development of MHC class I-restricted CD8(+) T cells in the thymus, they possess low numbers of high-avidity peripheral CTL with limited clonality and are able to contain acute and chronic infections. These in vivo data suggest that the CD8 coreceptor is not absolutely necessary for the generation of Ag-specific CTL. Lack of CD8 association causes partial TCR signaling because of the absence of CD8/Lck recruitment to the proximity of the MHC/TCR complex, resulting in suboptimal MAPK activation. Therefore, there should exist a signaling mechanism that can supplement partial TCR activation caused by the lack of CD8 association. In this human study, we have shown that CD8-independent stimulation of Ag-specific CTL previously primed in the presence of CD8 coligation, either in vivo or in vitro, induced severely impaired in vitro proliferation. When naive CD8(+) T cells were primed in the absence of CD8 binding and subsequently restimulated in the presence of CD8 coligation, the proliferation of Ag-specific CTL was also severely hampered. However, when CD8-independent T cell priming and restimulation were supplemented with IL-21, Ag-specific CD8(+) CTL expanded in two of six individuals tested. We found that IL-21 rescued partial MAPK activation in a STAT3- but not STAT1-dependent manner. These results suggest that CD8 coligation is critical for the expansion of postthymic peripheral Ag-specific CTL in humans. However, STAT3-mediated IL-21 signaling can supplement partial TCR signaling caused by the lack of CD8 association.
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http://dx.doi.org/10.4049/jimmunol.1003446DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3273545PMC
February 2012

Induction of HLA-DP4-restricted anti-survivin Th1 and Th2 responses using an artificial antigen-presenting cell.

Clin Cancer Res 2011 Aug 24;17(16):5392-401. Epub 2011 Jun 24.

Department of Medical Oncology, Dana-Farber Cancer Institute, USA.

Purpose: In previous cancer vaccine clinical trials targeting survivin, induction of specific CD8(+) T-cell responses did not consistently lead to clinical responses. Considering the critical role of CD4(+) T-cell help in generating antitumor immunity, integration of anti-survivin CD4(+) T-cell responses may enhance the efficacy of anti-survivin cancer immunotherapy. Human leukocyte antigen (HLA)-DP4 is emerging as an attractive MHC target allele of CD4(+) T cell-mediated immunotherapy, because it is one of the most frequent HLA alleles in many ethnic groups. In this article, we aimed to elucidate DP4-restricted CD4(+) T-cell responses against survivin in cancer patients.

Experimental Design: We generated a human cell-based artificial antigen-presenting cell (aAPC) expressing HLA-DP4, CD80, and CD83 and induced DP4-restricted antigen-specific CD4(+) T cells. The number, phenotype, effector function, and in vitro longevity of generated CD4(+) T cells were determined.

Results: We first determined previously unknown DP4-restricted CD4(+) T-cell epitopes derived from cytomegalovirus pp65, to which sustained Th1-biased recall responses were induced in vitro by using DP4-aAPC. In contrast, DP4-aAPC induced in vitro both Th1 and Th2 long-lived anti-survivin CD4(+) T cells from cancer patients. Both survivin-specific Th1 and Th2 cells were able to recognize survivin-expressing tumors in a DP4-restricted manner. Neither survivin-specific interleukin 10 secreting Tr1 cells nor Th17 cells were induced by DP4-aAPC.

Conclusions: DP4-restricted anti-survivin Th1 and Th2 immunity with sufficient functional avidity can be induced from cancer patients. The development of strategies to concurrently induce both CD4(+) and CD8(+) T-cell responses against survivin is warranted for optimal anti-survivin cancer immunotherapy.
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http://dx.doi.org/10.1158/1078-0432.CCR-10-3083DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3156899PMC
August 2011

Aberrant phosphorylation of STAT5 by granulocyte-macrophage colony-stimulating factor in infant cytomegalovirus infection mimicking juvenile myelomonocytic leukemia.

Leuk Res 2011 Sep 14;35(9):1261-4. Epub 2011 May 14.

Department of Pediatrics, Nagoya University Graduate School of Medicine, Shouwa-ku, Nagoya, Japan.

Juvenile myelomonocytic leukemia (JMML) progenitor cells exhibit in vitro hypersensitivity to granulocyte-macrophage colony-stimulating factor (GM-CSF). Phospho-specific flow cytometry using anti-phosphorylated STAT5 antibody is a new method recently reported to detect GM-CSF hypersensitivity of cells. However, colony assays using methylcellulose medium to measure GM-CSF-hypersensitivity remain as the current gold standard. Interestingly, cytomegalovirus (CMV) infection in infancy often presents with a variety of clinical symptoms that mimic JMML, with CMV giving a positive result by colony assay. We wanted to determine whether aberrant STAT5 activation occurs in CMV infection by using phospho-specific flow cytometry, and to ascertain whether this method is effective at discriminating CMV infection from JMML. Peripheral blood mononuclear cells from patients with JMML and CMV infection displayed an elevated proportion of p-STAT5 cells after low-dose GM-CSF stimulation when compared with cells from normal individuals. However, we found no significant differences in the percentage of p-STAT5 positive cells from patients with CMV infection and JMML at any doses of the GM-CSF doses used. We conclude that patients with CMV infection cannot be discriminated from patients with JMML by this new diagnostic method.
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http://dx.doi.org/10.1016/j.leukres.2011.04.014DOI Listing
September 2011

Establishment of antitumor memory in humans using in vitro-educated CD8+ T cells.

Sci Transl Med 2011 Apr;3(80):80ra34

Department of Medical Oncology, Dana-Farber Cancer Institute, 450 Brookline Avenue, Boston, MA 02215, USA.

Although advanced-stage melanoma patients have a median survival of less than a year, adoptive T cell therapy can induce durable clinical responses in some patients. Successful adoptive T cell therapy to treat cancer requires engraftment of antitumor T lymphocytes that not only retain specificity and function in vivo but also display an intrinsic capacity to survive. To date, adoptively transferred antitumor CD8(+) T lymphocytes (CTLs) have had limited life spans unless the host has been manipulated. To generate CTLs that have an intrinsic capacity to persist in vivo, we developed a human artificial antigen-presenting cell system that can educate antitumor CTLs to acquire both a central memory and an effector memory phenotype as well as the capacity to survive in culture for prolonged periods of time. We examined whether antitumor CTLs generated using this system could function and persist in patients. We showed that MART1-specific CTLs, educated and expanded using our artificial antigen-presenting cell system, could survive for prolonged periods in advanced-stage melanoma patients without previous conditioning or cytokine treatment. Moreover, these CTLs trafficked to the tumor, mediated biological and clinical responses, and established antitumor immunologic memory. Therefore, this approach may broaden the availability of adoptive cell therapy to patients both alone and in combination with other therapeutic modalities.
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http://dx.doi.org/10.1126/scitranslmed.3002207DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3861895PMC
April 2011

A panel of human cell-based artificial APC enables the expansion of long-lived antigen-specific CD4+ T cells restricted by prevalent HLA-DR alleles.

Int Immunol 2010 Nov 8;22(11):863-73. Epub 2010 Nov 8.

Department of Medical Oncology, Dana-Farber Cancer Institute, 44 Binney Street, Boston, MA 02115, USA.

Many preclinical experiments have attested to the critical role of CD4(+) T cell help in CD8(+) cytotoxic T lymphocyte (CTL)-mediated immunity. Recent clinical trials have demonstrated that reinfusion of CD4(+) T cells can induce responses in infectious diseases and cancer. However, few standardized and versatile systems exist to expand antigen-specific CD4(+) T(h) for clinical use. K562 is a human erythroleukemic cell line, which lacks expression of HLA class I and class II, invariant chain and HLA-DM but expresses adhesion molecules such as intercellular adhesion molecule-1 and leukocyte function-associated antigen-3. With this unique immunologic phenotype, K562 has been tested in clinical trials of cancer immunotherapy. Previously, we created a K562-based artificial antigen-presenting cell (aAPC) that generates ex vivo long-lived HLA-A2-restricted CD8(+) CTL with a central/effector memory phenotype armed with potent effector function. We successfully generated a clinical version of this aAPC and conducted a clinical trial where large numbers of anti-tumor CTL are reinfused to cancer patients. In this article, we shifted focus to CD4(+) T cells and developed a panel of novel K562-derived aAPC, where each expresses a different single HLA-DR allele, invariant chain, HLA-DM, CD80, CD83 and CD64; takes up soluble protein by endocytosis and processes and presents CD4(+) T-cell peptides. Using this aAPC, we were able to determine novel DR-restricted CD4(+) T-cell epitopes and expand long-lived CD4(+) T-cells specific for multiple antigens without growing bystander Foxp3(+) regulatory T cells. Our results suggest that K562-based aAPC may serve as a translatable platform to generate both antigen-specific CD8(+) CTL and CD4(+) T(h).
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http://dx.doi.org/10.1093/intimm/dxq440DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2994545PMC
November 2010

Downregulation of GATA-2 and overexpression of adipogenic gene-PPARgamma in mesenchymal stem cells from patients with aplastic anemia.

Exp Hematol 2009 Dec 20;37(12):1393-9. Epub 2009 Sep 20.

Department of Pediatrics, Nagoya University Graduate School of Medicine, Nagoya 466-8550, Japan.

Aplastic anemia (AA) is characterized by a reduced number of hematopoietic stem cells and fatty replacement in the bone marrow. Transcriptional factor GATA-2 plays several important roles in both hematopoiesis and adipogenesis. Decreased levels of GATA-2 compromise the proliferation and survival of hematopoietic stem cells. GATA-2 suppresses adipocyte differentiation through direct inhibition of adipogenic factors, including peroxisome proliferator-activated receptor-gamma (PPARgamma). Previous studies have shown that expression of GATA-2 is decreased in marrow CD34-positive cells in AA. To elucidate the mechanisms of fatty marrow replacement, we evaluated the mRNA expression for GATA-2 and PPARgamma in mesenchymal stem cells (MSCs) from patients with AA by quantitative real-time polymerase chain reaction. GATA-2 expression by MSCs from AA patients was significantly lower than in normal subjects. Conversely, expression of PPARgamma was significantly higher in AA patients. Western blot analysis demonstrated that protein levels of GATA-2 were lower in AA patients than those in normal subjects. Moreover, incubation with interferon-gamma induced downregulation of GATA-2 levels in MSCs from normal subjects. These findings indicate that fatty marrow replacement in AA patients can be explained by downregulation of GATA-2 and overexpression of PPARgamma in MSCs. Decreased expression of GATA-2 might be responsible for the pathogenesis and development of the clinical features of the disease.
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http://dx.doi.org/10.1016/j.exphem.2009.09.005DOI Listing
December 2009

Immunosuppressive activity of mesenchymal stem cells is not decreased in children with aplastic anemia.

Int J Hematol 2009 Jan 6;89(1):126-127. Epub 2009 Jan 6.

Department of Pediatrics, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, 466-8550, Japan.

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http://dx.doi.org/10.1007/s12185-008-0237-6DOI Listing
January 2009

Engraftment syndrome following allogeneic hematopoietic stem cell transplantation in children.

Pediatr Transplant 2009 Nov 1;13(7):831-7. Epub 2008 Nov 1.

Department of Pediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan.

ES is a complication that occurs immediately before or at the timing of neutrophil engraftment following autologous or allogeneic SCT. It is characterized by fever, skin rash, and non-cardiac pulmonary infiltrates. We evaluated the incidence, risk factors, and outcomes of ES following allogeneic SCT in children. Of 100 pediatric patients, 20 (20%) developed ES occurring at a median of 14 days (range 8-27 days) post-transplant. Patients presented with fever (100%), skin rash (100%), diffuse pulmonary infiltration (25%), and body weight gain (85%). On multivariate analysis, significant risk factors for ES included younger age (<8 yr old) and human leukocyte antigen disparity between donors and recipients. Univariate analysis showed that patients with ES had a higher incidence of developing chronic graft-versus-host disease and ES was not associated with other complications. Event-free survival did not significantly differ between patients with and without ES regardless of the presence of malignant or non-malignant diseases.
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http://dx.doi.org/10.1111/j.1399-3046.2008.01068.xDOI Listing
November 2009

Clinical impact of HLA-DR15, a minor population of paroxysmal nocturnal haemoglobinuria-type cells, and an aplastic anaemia-associated autoantibody in children with acquired aplastic anaemia.

Br J Haematol 2008 Jul 3;142(3):427-35. Epub 2008 Jun 3.

Department of Paediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Aplastic anaemia (AA) is defined as a pancytopenia caused by bone marrow failure, and its pathogenesis is thought to involve autoimmune processes. Several predictive markers of the response to immunosuppressive therapy (IST) have been proposed, which appear to reflect the immune pathophysiology. We prospectively investigated the presence of human leucocyte antigen (HLA)-DR15, a minor population of paroxysmal nocturnal haemoglobinuria (PNH)-type cells, and antibodies to the recently identified autoantigen postmeiotic segregation increased 1 (PMS1) in 103 children with AA enrolled in a multicentre study. In contrast to adults, children with AA did not show an increased frequency of HLA-DR15. In addition, a sensitive flow cytometric assay revealed that children with AA have a much lower prevalence of PNH-type cells (21.4%) than reported for adults with this disease. An immunoblotting assay detected anti-PMS1 antibody in 15 of 103 (14.6%) of the children. Finally, the response rate to IST was not significantly different between patients with and without DR15 (45.5% vs. 54.0%), PNH-type cells (68.2% vs. 53.1%) or anti-PMS1 antibody (40.0% vs. 59.1%). The current study did not confirm a correlation between these markers and the response to IST, suggesting that there is a difference in the pathophysiologies of adult and paediatric AA.
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http://dx.doi.org/10.1111/j.1365-2141.2008.07182.xDOI Listing
July 2008

Acute megakaryoblastic leukaemia (AMKL) in children: a comparison of AMKL with and without Down syndrome.

Br J Haematol 2008 Mar;140(5):552-61

Department of Paediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan.

To characterize childhood acute megakaryoblastic leukaemia (AMKL), we reviewed 45 children with AMKL diagnosed between 1986 and 2005 at Nagoya University Hospital and Japanese Red Cross Nagoya First Hospital. Twenty-four patients (53%) had AMKL associated with Down syndrome (DS-AMKL) and 21 (47%) had non-DS-AMKL. The median age of the DS-AMKL patients was 21 months (range, 8-38 months) and that of non-DS-AMKL patients was 15 months (range, 2-185 months). The morphology of blast cells was categorized into three groups according to the stage of megakaryocyte maturation. The blast cells were more immature in DS-AMKL than in non-DS-AMKL in terms of morphology and immunophenotyping. Cytogenetic abnormalities of leukaemic cells were classified into seven categories: normal karyotype including constitutional trisomy 21 in DS-AMKL; numerical abnormalities only; t(1;22)(p13;q13); 3q21q26 abnormalities; t(16;21)(p11;q22); -5/del(5q) and/or -7/del(7q); and other structural changes. The outcome of children with either DS-AMKL or non-DS-AMKL is excellent. The 10-year overall survival estimate was 79% [95% confidence interval (CI): 54-90] for DS-AMKL and 76% (95% CI: 58-91) for non-DS-AMKL (P = 0.81) with a median follow-up of 78 months (range, 20-243 months). Our study shows the diverse heterogeneity of childhood AMKL and the need for subclassification according to cytogenetic and morphological features.
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http://dx.doi.org/10.1111/j.1365-2141.2007.06971.xDOI Listing
March 2008

Quantification of granulocyte-macrophage colony-stimulating factor hypersensitivity in juvenile myelomonocytic leukemia by 3H-thymidine assay.

Leuk Res 2008 Jul 4;32(7):1036-42. Epub 2008 Mar 4.

Department of Pediatrics, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya 466-8550, Japan.

Recent molecular studies have revealed that the GM-CSF/RAS signaling pathway plays a central role in the pathogenesis of juvenile myelomonocytic leukemia (JMML). CFU-GM colony assay is an important test for GM-CSF hypersensitivity in patients with JMML, but requires specific skills. We established a simple and easy quantification method to test GM-CSF hypersensitivity, using a (3)H-thymidine assay. With this quantification method, JMML patients with RAS mutations showed significantly higher GM-CSF sensitivity than JMML patients with PTPN11 mutations. This method will be useful not only in the diagnosis of JMML, but also to evaluate the difference of GM-CSF sensitivity among patients.
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http://dx.doi.org/10.1016/j.leukres.2007.11.016DOI Listing
July 2008

Feasibility and results of bone marrow transplantation from an HLA-mismatched unrelated donor for children and young adults with acquired severe aplastic anemia.

Int J Hematol 2007 Jun;85(5):437-42

Department of Pediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Treating patients with severe aplastic anemia (SAA) who fail to respond to immunosuppressive therapy (IST) and do not have an HLA-matched donor is challenging. We report favorable outcomes in 11 patients who underwent bone marrow transplantation (BMT) from an HLA-mismatched unrelated donor. The median age was 11 years (range, 3-20 years). The conditioning regimen consisted of cyclophosphamide (200 mg/kg), antithymocyte globulin (10 mg/kg), and total body irradiation (5 Gy). Patients received tacrolimus and methotrexate for prophylaxis against graft-versus-host disease (GVHD). Donorrecipient pairs were mismatched for the HLA-DR antigen in 8 patients by serologic typing. HLA-A and HLA-B antigens were mismatched in 1 and 2 patients, respectively. Ten patients achieved engraftment. One patient who failed to engraft was rescued by a second transplantation from her mother, who was mismatched at 2 HLA antigens. Acute GVHD of grades II to IV occurred in 2 patients. Three patients developed limited chronic GVHD, and 1 patient developed extensive chronic GVHD of the lung. All patients are alive at 9 to 56 months after transplantation (median, 33 months). Considering our encouraging results, HLA-mismatched unrelated-donor BMT for SAA is feasible as a salvage therapy for nonresponders to IST.
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http://dx.doi.org/10.1532/IJH97.06229DOI Listing
June 2007

Increased midkine gene expression in childhood B-precursor acute lymphoblastic leukemia.

Leuk Res 2007 Aug 30;31(8):1045-51. Epub 2007 Jan 30.

Department of Pediatrics, Nagoya University Graduate School of Medicine, Nagoya, Japan.

Midkine (MK) is a heparin-binding growth factor that is overexpressed in a number of solid cancers. However, expression in acute leukemia has not been clarified. We examined MK gene expression using real-time PCR in 94 children with acute leukemia. In 30 of the 41 patients with B-precursor ALL, MK gene expression was overexpressed than normal BM. MK gene was also overexpressed in more than half of patients with FAB M1 and M2 types of AML. Quantification of MK gene by real-time PCR offers particular promise as a prognostic marker and a marker for minimal residual disease in children with B-precursor ALL.
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http://dx.doi.org/10.1016/j.leukres.2006.12.008DOI Listing
August 2007