Language：English  

DOI： 10.1080/15592294.2023.2268814

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1182/blood.2022017813

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1038/s41598-023-33852-2

Language：English  

DOI： 10.1124/molpharm.122.000546

Language：English   Publishing type：Research paper (scientific journal)  

Philadelphia chromosome-like acute lymphoblastic leukemia (Ph-like ALL) is intractable and mostly harbors genetic alterations that activate JAK or ABL signaling. The commonest subtype of Ph-like ALL exhibits the CRLF2 gene arrangement that brings about JAK1/2-STAT5 pathway activation. However, JAK1/2 inhibition alone is insufficient for treatment, necessitating combinatorial therapies targeting multiple signals. To better understand the mechanisms underlying the insufficient efficacy of JAK inhibition, this study explored gene expression changes upon treatment with a JAK1/2 inhibitor (ruxolitinib) and revealed BCL6 elevation as one such mechanism. Upregulated BCL6 suppressed TP53 expression along with its downstream cell cycle inhibitor p21 (CDKN2A) and proapoptotic molecules, such as FAS, TNFRSF10B, BID, BAX, BAK, PUMA, and NOXA, conferring cells in part with therapy resistance. The BCL6 inhibition (FX1) alone was able to upregulate TP53 and restore TP53 expression that ruxolitinib had diminished. In addition, ruxolitinib and FX1 concertedly downregulated MYC. As a result, FX1 treatment alone had growth-inhibitory and apoptosis-sensitizing effects, but the combination of ruxolitinib and FX1 more potently inhibited leukemia cell growth, enhanced apoptosis sensitivity, and prolonged xenografted mice survival. These findings provide one mechanism for the insufficiency of JAK inhibition for CRLF2-rearranged ALL treatment and BCL6 inhibition as a potentially helpful adjunctive therapy combined with JAK inhibition.

DOI： 10.3324/haematol.2022.280879

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1007/s12185-022-03521-7

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Poly (ADP-ribose) polymerase (PARP) inhibitors (PARPi) function by inhibiting base excision repair and inducing synthetic lethality in homologous recombination repair-deficient cells, such as BRCA1/2-mutated cancer cells. The BCR/ABL1 fusion protein causes dysregulated cell proliferation and is responsible for chronic myelogenous leukemia and Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ALL). BCR/ABL1 also induces genomic instability by downregulating BRCA1. We investigated the effect of the PARPi, olaparib, against Ph+ALL cell lines and found that they show variable sensitivity, presumably due to cancer-associated genetic alterations other than BCR/ABL1. To investigate the reasons for the variable responses of Ph+ALL cells to PARPi treatment, we analyzed the transcriptomes of olaparib-sensitive and -resistant Ph+ALL cell lines, which revealed that activation of the phosphatidylinositol 3-kinase (PI3K) pathway was a hallmark of PARPi resistance. Based on these findings, we examined the effects of adding a PI3K inhibitor (PI3Ki) to PARPi treatment to overcome PARPi insensitivity in Ph+ALL cell lines. Combination with PI3Ki increased PARPi cytotoxicity in PARPi-resistant Ph+ALL cell lines. Tyrosine kinase inhibitor (TKI) therapy is the gold standard for Ph+ALL, and, based on our findings, we propose that PARPi combined with TKI and PI3K inhibition could be a novel therapeutic strategy for Ph+ALL.

DOI： 10.1007/s12185-022-03520-8

PubMed

Authorship：Lead author,　Last author,　Corresponding author   Language：Japanese  

DOI： 10.11406/rinketsu.63.1566

PubMed

Language：English   Publishing type：Research paper (scientific journal)   Publisher：Springer Science and Business Media LLC  

Abstract

The Philadelphia (Ph) chromosome was the first translocation identified in leukemia. It is supposed to be generated by aberrant ligation between two DNA double-strand breaks (DSBs) at the BCR gene located on chromosome 9q34 and the ABL1 gene located on chromosome 22q11. Thus, mimicking the initiation process of translocation, we induced CRISPR/Cas9-mediated DSBs simultaneously at the breakpoints of the BCR and ABL1 genes in a granulocyte-macrophage colony-stimulating factor (GM-CSF) dependent human leukemia cell line. After transfection of two single guide RNAs (sgRNAs) targeting intron 13 of the BCR gene and intron 1 of the ABL1 gene, a factor-independent subline was obtained. In the subline, p210 BCR::ABL1 and its reciprocal ABL1::BCR fusions were generated as a result of balanced translocation corresponding to the Ph chromosome. Another set of sgRNAs targeting intron 1 of the BCR gene and intron 1 of the ABL1 gene induced a factor-independent subline expressing p190 BCR::ABL1. Both p210 and p190 BCR::ABL1 induced factor-independent growth by constitutively activating intracellular signaling pathways for transcriptional regulation of cell cycle progression and cell survival that are usually regulated by GM-CSF. These observations suggested that simultaneous DSBs at the BCR and ABL1 gene breakpoints are initiation events for oncogenesis in Ph+ leukemia. (200/200 words).

DOI： 10.1038/s41417-022-00522-w

PubMed

Other Link： https://www.nature.com/articles/s41417-022-00522-w

Language：English   Publishing type：Research paper (scientific journal)   Publisher：Wiley  

DOI： 10.1111/bjh.18405

PubMed

Other Link： https://onlinelibrary.wiley.com/doi/full-xml/10.1111/bjh.18405

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1111/ped.15233

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Imatinib and second-generation tyrosine kinase inhibitors (TKIs) have dramatically improved the prognosis of Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL). However, overcoming TKI resistance due to the T315I gatekeeper mutation of BCR/ABL1 is crucial for further improving the prognosis. The clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is appropriate for establishing a human model of Ph+ ALL with the T315I mutation, because it can induce specific mutations via homologous recombination (HR) repair in cells with intact endogenous HR pathway. Here we used CRISPR/Cas9 to introduce the T315I mutation into the Ph+ lymphoid leukemia cell line KOPN55bi, which appeared to have an active HR pathway based on its resistance to a poly (ADP-Ribose) polymerase-1 inhibitor. Single-guide RNA targeting at codon 315 and single-strand oligodeoxynucleotide containing ACT to ATT nucleotide transition at codon 315 were electroporated with recombinant Cas9 protein. Dasatinib-resistant sublines were obtained after one-month selection with the therapeutic concentration of dasatinib, leading to T315I mutation acquisition through HR. T315I-acquired sublines were highly resistant to imatinib and second-generation TKIs but moderately sensitive to the therapeutic concentration of ponatinib. This authentic human model is helpful for developing new therapeutic strategies overcoming TKI resistance in Ph+ ALL due to T315I mutation.

DOI： 10.1007/s12185-022-03369-x

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Glucocorticoid (GC) is a key drug in the treatment of B-cell precursor acute lymphoblastic leukemia (BCP-ALL), and the initial GC response is an important prognostic factor. GC receptors play an essential role in GC sensitivity, and somatic mutations of the GC receptor gene, NR3C1, are reportedly identified in some BCP-ALL cases, particularly at relapse. Moreover, associations of somatic mutations of the CREB-binding protein (CREBBP) and Wolf-Hirschhorn syndrome candidate 1 (WHSC1) genes with the GC-resistance of ALL have been suggested. However, the significance of these mutations in the GC sensitivity of BCP-ALL remains to be clarified in the intrinsic genes. In the present study, we sequenced NR3C1, WHSC1, and CREBBP genes in 99 BCP-ALL and 22 T-ALL cell lines (32 and 67 cell lines were known to be established at diagnosis and at relapse, respectively), and detected their mutations in 19 (2 cell lines at diagnosis and 15 cell lines at relapse), 26 (6 and 15), and 38 (11 and 15) cell lines, respectively. Of note, 14 BCP-ALL cell lines with the NR3C1 mutations were significantly more resistant to GC than those without mutations. In contrast, WHSC1 and CREBBP mutations were not associated with GC resistance. However, among the NR3C1 unmutated BCP-ALL cell lines, WHSC1 mutations tended to be associated with GC resistance and lower NR3C1 gene expression. Finally, we successfully established GC-resistant sublines of the GC-sensitive BCP-ALL cell line (697) by disrupting ligand binding and DNA binding domains of the NR3C1 gene using the CRISPR/Cas9 system. These observations demonstrated that somatic mutations of the NR3C1 gene, and possibly the WHSC1 gene, confer GC resistance in BCP-ALL.

DOI： 10.1016/j.jsbmb.2022.106068

PubMed

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1097/MD.0000000000029054

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1182/blood.2021012976.

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Acute lymphoblastic leukemia with chromosomal rearrangements involving the mixed-lineage leukemia (MLL) gene (MLL-r ALL) remains an incurable disease. Thus, development of a safe and effective therapeutic agent to treat this disease is crucial to address this unmet medical need. BRD4, a member of the bromodomain and extra-terminal domain (BET) protein family, and cyclic AMP response element binding protein binding protein (CBP) and p300, two paralogous histone acetyltransferases, are all considered cancer drug targets and simultaneous targeting of these proteins may have therapeutic advantages. Here, we demonstrate that a BET/CBP/p300 multi-bromodomain inhibitor, CN470, has anti-tumor activity against MLL-r ALL in vitro and in vivo. CN470, potently inhibited ligand binding to the bromodomains of BRD4, CBP, and p300 and suppressed the growth of MLL-r ALL cell lines and patient-derived cells with MLL rearrangements. CN470 suppressed mRNA and protein expression of MYC and induced apoptosis in MLL-r ALL cells, following a cell cycle arrest in the G1 phase. Moreover, CN470 reduced BRD4 binding to acetylated histone H3. The in vivo effects of CN470 were investigated using SEMLuc/GFP cells expressing luminescent markers in an orthotopic mouse model. Mice administered CN470 daily had prolonged survival compared to the vehicle group. Further, CN470 also showed anti-tumor effects against an MLL-r ALL patient-derived xenograft model. These findings suggest that inhibition of BET/CBP/p300 by the multi-bromodomain inhibitor, CN470, represents a promising therapeutic approach against MLL-r ALL.

DOI： 10.1016/j.bbrc.2021.12.078

PubMed

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

Asparaginase therapy is a key component of chemotherapy for T-cell acute lymphoblastic leukemia (T-ALL) patients. Asparaginase depletes serum asparagine by deamination into aspartic acid. Normal hematopoietic cells can survive due to asparagine synthetase (ASNS) activity, while leukemia cells are supposed to undergo apoptosis due to silencing of the ASNS gene. Since the ASNS gene has a typical CpG island in its promoter, its methylation status in T-ALL cells may be associated with asparaginase sensitivity. Thus, we investigated the significance of ASNS methylation status in asparaginase sensitivity of T-ALL cell lines and prognosis of childhood T-ALL. Sequencing of bisulfite PCR products using next-generation sequencing technology in 22 T-ALL cell lines revealed a stepwise allele-specific methylation of the ASNS gene, in association with an aberrant methylation of a 7q21 imprinted gene cluster. T-ALL cell lines with ASNS hypermethylation status showed significantly higher in vitro l-asparaginase sensitivity in association with insufficient asparaginase-induced upregulation of ASNS gene expression and lower basal ASNS protein expression. A comprehensive analysis of diagnostic samples from childhood T-ALL patients in Japanese cohorts (n = 77) revealed that methylation of the ASNS gene was associated with an aberrant methylation of the 7q21 imprinted gene cluster. In childhood T-ALL patients in Japanese cohorts (n = 75), ASNS hypomethylation status was significantly associated with poor therapeutic outcome, and all cases with poor prognostic SPI1 fusion exclusively showed ASNS hypomethylation status. These observations demonstrate that ASNS hypomethylation status is associated with asparaginase resistance and is a poor prognostic biomarker in childhood T-ALL.

DOI： 10.1182/bloodadvances.2021004271

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

CRLF2-rearranged (CRLF2r) acute lymphoblastic leukemia (ALL) accounts for more than half of Philadelphia chromosome-like (Ph-like) ALL and is associated with a poor outcome in children and adults. Overexpression of CRLF2 results in activation of Janus kinase (JAK)-STAT and parallel signaling pathways in experimental models, but existing small molecule inhibitors of JAKs show variable and limited efficacy. Here, we evaluated the efficacy of proteolysis-targeting chimeras (PROTACs) directed against JAKs. Solving the structure of type I JAK inhibitors ruxolitinib and baricitinib bound to the JAK2 tyrosine kinase domain enabled the rational design and optimization of a series of cereblon (CRBN)-directed JAK PROTACs utilizing derivatives of JAK inhibitors, linkers, and CRBN-specific molecular glues. The resulting JAK PROTACs were evaluated for target degradation, and activity was tested in a panel of leukemia/lymphoma cell lines and xenograft models of kinase-driven ALL. Multiple PROTACs were developed that degraded JAKs and potently killed CRLF2r cell lines, the most active of which also degraded the known CRBN neosubstrate GSPT1 and suppressed proliferation of CRLF2r ALL in vivo, e.g. compound 7 (SJ988497). Although dual JAK/GSPT1-degrading PROTACs were the most potent, the development and evaluation of multiple PROTACs in an extended panel of xenografts identified a potent JAK2-degrading, GSPT1-sparing PROTAC that demonstrated efficacy in the majority of kinase-driven xenografts that were otherwise unresponsive to type I JAK inhibitors, e.g. compound 8 (SJ1008030). Together, these data show the potential of JAK-directed protein degradation as a therapeutic approach in JAK-STAT-driven ALL and highlight the interplay of JAK and GSPT1 degradation activity in this context.

DOI： 10.1182/blood.2020006846

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

In chemotherapy for childhood acute lymphoblastic leukaemia (ALL), maintenance therapy consisting of oral daily mercaptopurine and weekly methotrexate is important. NUDT15 variant genotype is reportedly highly associated with severe myelosuppression during maintenance therapy, particularly in Asian and Hispanic populations. It has also been demonstrated that acquired somatic mutations of the NT5C2 and PRPS1 genes, which are involved in thiopurine metabolism, are detectable in a portion of relapsed childhood ALL. To directly confirm the significance of the NUDT15 variant genotype and NT5C2 and PRPS1 mutations in thiopurine sensitivity of leukaemia cells in the intrinsic genes, we investigated 84 B-cell precursor-ALL (BCP-ALL) cell lines. Three and 14 cell lines had homozygous and heterozygous variant diplotypes of the NUDT15 gene, respectively, while 4 and 2 cell lines that were exclusively established from the samples at relapse had the NT5C2 and PRPS1 mutations, respectively. Both NUDT15 variant genotype and NT5C2 and PRPS1 mutations were significantly associated with DNA-incorporated thioguanine levels after exposure to thioguanine at therapeutic concentration. Considering the continuous exposure during the maintenance therapy, we evaluated in vitro mercaptopurine sensitivity after 7-day exposure. Mercaptopurine concentrations lethal to 50% of the leukaemia cells were comparable to therapeutic serum concentration of mercaptopurine. Both NUDT15 variant genotype and NT5C2 and PRPS1 mutations were significantly associated with mercaptopurine sensitivity in 83 BCP-ALL and 23 T-ALL cell lines. The present study provides direct evidence to support the general principle showing that both inherited genotype and somatically acquired mutation are crucially implicated in the drug sensitivity of leukaemia cells.

DOI： 10.1111/jcmm.16981

PubMed

Language：English   Publishing type：Research paper (scientific journal)   Publisher：Springer Science and Business Media LLC  

<title>Abstract</title>Tyrosine kinase inhibitor (TKI) treatment has dramatically improved the survival of chronic myeloid leukemia (CML) patients, but measurable residual disease typically persists. To more effectively eradicate leukemia cells, simultaneous targeting of BCR-ABL1 and additional CML-related survival proteins has been proposed. Notably, several highly specific myeloid cell leukemia 1 (MCL1) inhibitors have recently entered clinical trials for various hematologic malignancies, although not for CML, reflecting the insensitivity of CML cell lines to single MCL1 inhibition. Here, we show that combining TKI (imatinib, nilotinib, dasatinib, or asciminib) treatment with the small-molecule MCL1 inhibitor S63845 exerted strong synergistic antiviability and proapoptotic effects on CML lines and CD34+ stem/progenitor cells isolated from untreated CML patients in chronic phase. Using wild-type BCR-ABL1-harboring CML lines and their T315I-mutated sublines (generated by CRISPR/Cas9-mediated homologous recombination), we prove that the synergistic proapoptotic effect of the drug combination depended on TKI-mediated BCR-ABL1 inhibition, but not on TKI-related off-target mechanisms. Moreover, we demonstrate that colony formation of CML but not normal hematopoietic stem/progenitor cells became markedly reduced upon combination treatment compared to imatinib monotherapy. Our results suggest that dual targeting of MCL1 and BCR-ABL1 activity may efficiently eradicate residual CML cells without affecting normal hematopoietic stem/progenitors.

DOI： 10.1038/s41419-021-04154-0

PubMed

Other Link： https://www.nature.com/articles/s41419-021-04154-0

Language：English   Publishing type：Research paper (scientific journal)   Publisher：The American Association of Immunologists  

DOI： 10.4049/immunohorizons.2100054

PubMed

Language：English   Publishing type：Research paper (scientific journal)   Publisher：BMC  

Background The genetic variants of theARID5Bgene have recently been reported to be associated with disease susceptibility and treatment outcome in childhood acute lymphoblastic leukemia (ALL). However, few studies have explored the association of ARID5B with sensitivities to chemotherapeutic agents. Methods We genotyped susceptibility-linked rs7923074 and rs10821936 as well as relapse-linked rs4948488, rs2893881, and rs6479778 ofARDI5Bby direct sequencing of polymerase chain reaction (PCR) products in 72 B-cell precursor-ALL (BCP-ALL) cell lines established from Japanese patients. We also quantified theirARID5Bexpression levels by real-time reverse transcription PCR, and determined their 50% inhibitory concentration (IC50) values by alamarBlue assays in nine representative chemotherapeutic agents used for ALL treatment. Results No significant associations were observed in genotypes of the susceptibility-linked single nucleotide polymorphisms (SNPs) and the relapsed-linked SNPs withARID5Bgene expression levels. Of note, IC50 values of vincristine (VCR) (median IC50: 39.6 ng/ml) in 12 cell lines with homozygous genotype of risk allele (C) in the relapse-linked rs4948488 were significantly higher (p = 0.031 in Mann-Whitney U test) than those (1.04 ng/ml) in 60 cell lines with heterozygous or homozygous genotypes of the non-risk allele (T). Furthermore, the IC50 values of mafosfamide [Maf; active metabolite of cyclophosphamide (CY)] and cytarabine (AraC) tended to be associated with the genotype of rs4948488. Similar associations were observed in genotypes of the relapse-linked rs2893881 and rs6479778, but not in those of the susceptibility-linked rs7923074 and rs10821936. In addition, the IC50 values of methotrexate (MTX) were significantly higher (p = 0.023) in 36 cell lines with lowerARID5Bgene expression (median IC50: 37.1 ng/ml) than those in the other 36 cell lines with higher expression (16.9 ng/ml). Conclusion These observations in 72 BCP-ALL cell lines suggested that the risk allele of the relapse-linked SNPs ofARID5Bmay be involved in a higher relapse rate because of resistance to chemotherapeutic agents such as VCR, CY, and AraC. In addition, lowerARID5Bgene expression may be associated with MTX resistance.

DOI： 10.1186/s12935-020-01524-0

Web of Science

Language：English   Publishing type：Research paper (scientific journal)  

The long-term prognosis of Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph + ALL) is still unsatisfactory even after the emergence of tyrosine kinase inhibitors (TKIs) against chimeric BCR-ABL, and this is associated with the high incidence of genetic alterations of Ikaros family zinc finger 1 (IKZF1), most frequently the hemi-allelic loss of exons 4-7 expressing a dominant-negative isoform Ik6. We found that lenalidomide (LEN), a representative of immunomodulatory drugs (IMiDs), which have been long used for the treatment of multiple myeloma, specifically induced accumulation of Ik6 with the disappearance of functional isoforms within 24 h (i.e., abrupt and complete shut-down of the IKZF1 activity) in Ik6-positive Ph+ALL cells in a neddylation-dependent manner. The functional IKZF3 isoforms expression was also abruptly and markedly downregulated. The LEN treatment specifically suppressed proliferation of Ik6-positive-Ph+ALL cells by inducing cell cycle arrest via downregulation of cyclins D3 and E and CDK2, and of importance, markedly upregulated their apoptosis in synergy with the TKI imatinib (IM). Apoptosis of IM-resistant Ph+ALL cells with T315I mutation of BCR-ABL was also upregulated by LEN in the presence of the newly developed TKI ponatinib. Analyses of flow cytometry, western blot, and oligonucleotide array revealed that apoptosis was caspase-/p53-dependent and associated with upregulation of pro-apoptotic Bax/Bim, enhanced dephosphorylation of BCR-ABL/Akt, and downregulation of oncogenic helicase genes HILLS, CDC6, and MCMs4 and 8. Further, the synergism of LEN with IM was clearly documented as a significant prolongation of survival in the xenograft mice model. Because this synergism was further potentiated in vitro by dexamethasone, a key drug for ALL treatment, the strategy of repositioning IMiDs for the treatment of Ik6-positive Ph+ALL patients certainly shed new light on an outpatient-based treatment option for achieving their long-term durable remission and higher QOL, particularly for those who are not tolerable to intensified therapeutic approaches.

DOI： 10.1038/s41420-021-00523-y

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Immunotherapies specific for B-cell precursor acute lymphoblastic leukemia (BCP-ALL), such as anti-CD19 chimeric antigen receptor (CAR) T-cells and blinatumomab, have dramatically improved the therapeutic outcome in refractory cases. In the anti-leukemic activity of those immunotherapies, TNF-related apoptosis-inducing ligand (TRAIL) on cytotoxic T-cells plays an essential role by inducing apoptosis of the target leukemia cells through its death receptors (DR4 and DR5). Since there are CpG islands in the promoter regions, hypermethylation of the DR4 and DR5 genes may be involved in resistance of leukemia cells to immunotherapies due to TRAIL-resistance. We analyzed the DR4 and DR5 methylation status in 32 BCP-ALL cell lines by sequencing their bisulfite PCR products with a next-generation sequencer. The DR4 and DR5 methylation status was significantly associated with the gene and cell-surface expression levels and the TRAIL-sensitivities. In the clinical samples at diagnosis (459 cases in the NOPHO study), both DR4 and DR5 genes were unmethylated in the majority of cases, whereas methylated in several cases with dic(9;20), MLL-rearrangement, and hypodiploidy, suggesting that evaluation of methylation status of the DR4 and DR5 genes might be clinically informative to predict efficacy of immunotherapy in certain cases with such unfavorable karyotypes. These observations provide an epigenetic rational for clinical efficacy of immunotherapy in the vast majority of BCP-ALL cases.

DOI： 10.3390/genes12060864

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Identification of genetic variants associated with glucocorticoids (GC) sensitivity of leukaemia cells may provide insight into potential drug targets and tailored therapy. In the present study, within 72 leukaemic cell lines derived from Japanese patients with B-cell precursor acute lymphoblastic leukaemia (ALL), we conducted genome-wide genotyping of single nucleotide polymorphisms (SNP) and attempted to identify genetic variants associated with GC sensitivity and NR3C1 (GC receptor) gene expression. IC50 measures for prednisolone (Pred) and dexamethasone (Dex) were available using an alamarBlue cell viability assay. IC50 values of Pred showed the strongest association with rs904419 (P = 4.34 × 10-8 ), located between the FRMD4B and MITF genes. The median IC50 values of prednisolone for cell lines with rs904419 AA (n = 13), AG (n = 31) and GG (n = 28) genotypes were 0.089, 0.139 and 297 µmol/L, respectively. For dexamethasone sensitivity, suggestive association was observed for SNP rs2306888 (P = 1.43 × 10-6 ), a synonymous SNP of the TGFBR3 gene. For NR3C1 gene expression, suggestive association was observed for SNP rs11982167 (P = 6.44 × 10-8 ), located in the PLEKHA8 gene. These genetic variants may affect GC sensitivity of ALL cells and may give rise to opportunities in personalized medicine for effective and safe chemotherapy in ALL patients.

DOI： 10.1111/jcmm.15882

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Blinatumomab, a bispecific antibody that directs CD3+ T cells to CD19+ tumor cells, shows variable efficacy in B-progenitor acute lymphoblastic leukemia (B-ALL). To determine tumor-intrinsic and -extrinsic determinants of response, we studied 44 adults with relapsed or refractory B-ALL (including 2 minimal residual disease positive) treated with blinatumomab using bulk tumor and single-cell sequencing. The overall response rate in patients with hematological disease was 55%, with a high response rate in those with CRLF2-rearranged Philadelphia chromosome-like ALL (12 [75%] of 16). Pretreatment samples of responders exhibited a tumor-intrinsic transcriptomic signature of heightened immune response. Multiple mechanisms resulted in loss of CD19 expression, including CD19 mutations, CD19-mutant allele-specific expression, low CD19 RNA expression, and mutations in CD19 signaling complex member CD81. Patients with low hypodiploid ALL were prone to CD19- relapse resulting from aneuploidy-mediated loss of the nonmutated CD19 allele. Increased expression of a CD19 isoform with intraexonic splicing of exon 2, CD19 ex2part, at baseline or during therapy was associated with treatment failure. These analyses demonstrate both tumor-intrinsic and -extrinsic factors influence blinatumomab response. We show that CD19 mutations are commonly detected in CD19- relapse during blinatumomab treatment. Identification of the CD19 ex2part splice variant represents a new biomarker predictive of blinatumomab therapy failure.

DOI： 10.1182/blood.2020006287

PubMed

Language：English   Publishing type：Research paper (scientific journal)   Publisher：American Society of Hematology  

Karyotype is an important prognostic factor in childhood B-cell precursor acute lymphoblastic leukemia (BCP-ALL), but the underlying pharmacogenomics remains unknown. Asparaginase is an integral component in current chemotherapy for childhood BCP-ALL. Asparaginase therapy depletes serum asparagine. Normal hematopoietic cells can produce asparagine by asparagine synthetase (ASNS) activity, while ALL cells are unable to synthesize adequate amounts of asparagine. The ASNS gene has a typical CpG island in its promoter. Thus, methylation of the ASNS CpG island could be one of the epigenetic mechanisms for ASNS gene silencing in BCP-ALL. To gain deep insights into the pharmacogenomics of asparaginase therapy, we investigated the association of ASNS methylation status with asparaginase sensitivity. ASNS CpG island is largely unmethylated in normal hematopoietic cells but is allele-specifically methylated in BCP-ALL cells. The ASNS gene is located at 7q21, an evolutionally conserved imprinted gene cluster. ASNS methylation in childhood BCP-ALL is associated with an aberrant methylation of the imprinted gene cluster at 7q21. Aberrant methylation of mouse Asns and a syntenic imprinted gene cluster is also confirmed in leukemic spleen samples from ETV6-RUNX1 knock-in mice. In three childhood BCP-ALL cohorts, ASNS is highly methylated in BCP-ALL with favorable karyotypes but is mostly unmethylated in BCP-ALL with poor prognostic karyotypes. Higher ASNS methylation is associated with higher l-asparaginase sensitivity in BCP-ALL through lower ASNS gene and protein expression levels. These observations demonstrate that silencing of the ASNS gene due to aberrant imprinting is a pharmacogenetic mechanism for the leukemia-specific activity of asparaginase therapy in BCP-ALL.

DOI： 10.1182/blood.2019004090

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1002/hon.2762

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

Therapy resistance in leukemia may be due to cancer cell-intrinsic and/or -extrinsic mechanisms. Mutations within BCR-ABL1, the oncogene giving rise to chronic myeloid leukemia (CML), lead to resistance to tyrosine kinase inhibitors (TKI), and some are associated with clinically more aggressive disease and worse outcome. Using the retroviral transduction/transplantation model of CML and human cell lines we faithfully recapitulate accelerated disease course in TKI resistance. We show in various models, that murine and human imatinib-resistant leukemia cells positive for the oncogene BCR-ABL1T315I differ from BCR-ABL1 native (BCR-ABL1) cells with regards to niche location and specific niche interactions. We implicate a pathway via integrin β3, integrin-linked kinase (ILK) and its role in deposition of the extracellular matrix (ECM) protein fibronectin as causative of these differences. We demonstrate a trend towards a reduced BCR-ABL1T315I+ tumor burden and significantly prolonged survival of mice with BCR-ABL1T315I+ CML treated with fibronectin or an ILK inhibitor in xenogeneic and syngeneic murine transplantation models, respectively. These data suggest that interactions with ECM proteins via the integrin β3/ILK-mediated signaling pathway in BCR-ABL1T315I+ cells differentially and specifically influence leukemia progression. Niche targeting via modulation of the ECM may be a feasible therapeutic approach to consider in this setting.

DOI： 10.1038/s41375-020-0866-1

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

A heritable polymorphism within regulatory sequences of the LMO1 gene is associated with its elevated expression and increased susceptibility to develop neuroblastoma, but the oncogenic pathways downstream of the LMO1 transcriptional co-regulatory protein are unknown. Our ChIP-seq and RNA-seq analyses reveal that a key gene directly regulated by LMO1 and MYCN is ASCL1, which encodes a basic helix-loop-helix transcription factor. Regulatory elements controlling ASCL1 expression are bound by LMO1, MYCN and the transcription factors GATA3, HAND2, PHOX2B, TBX2 and ISL1-all members of the adrenergic (ADRN) neuroblastoma core regulatory circuitry (CRC). ASCL1 is required for neuroblastoma cell growth and arrest of differentiation. ASCL1 and LMO1 directly regulate the expression of CRC genes, indicating that ASCL1 is a member and LMO1 is a coregulator of the ADRN neuroblastoma CRC.

DOI： 10.1038/s41467-019-13515-5

PubMed

Authorship：Lead author,　Corresponding author   Language：English   Publishing type：Research paper (scientific journal)  

DOI： 10.1002/hon.2654

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

t(17;19)(q21-q22;p13), responsible for TCF3-HLF fusion, is a rare translocation in childhood B-cell precursor acute lymphoblastic leukemia(BCP-ALL). t(1;19)(q23;p13), producing TCF3-PBX1 fusion, is a common translocation in childhood BCP-ALL. Prognosis of t(17;19)-ALL is extremely poor, while that of t(1;19)-ALL has recently improved dramatically in intensified chemotherapy. In this study, TCF3-HLF mRNA was detectable at a high level during induction therapy in a newly diagnosed t(17;19)-ALL case, while TCF3-PBX1 mRNA was undetectable at the end of induction therapy in most newly diagnosed t(1;19)-ALL cases. Using 4 t(17;19)-ALL and 16 t(1;19)-ALL cell lines, drug response profiling was analyzed. t(17;19)-ALL cell lines were found to be significantly more resistant to vincristine (VCR), daunorubicin (DNR), and prednisolone (Pred) than t(1;19)-ALL cell lines. Sensitivities to three (Pred, VCR, and l-asparaginase [l-Asp]), four (Pred, VCR, l-Asp, and DNR) and five (Pred, VCR, l-Asp, DNR, and cyclophosphamide) agents, widely used in induction therapy, were significantly poorer for t(17;19)-ALL cell lines than for t(1;19)-ALL cell lines. Consistent with poor responses to VCR and DNR, gene and protein expression levels of P-glycoprotein (P-gp) were higher in t(17;19)-ALL cell lines than in t(1;19)-ALL cell lines. Inhibitors for P-gp sensitized P-gp-positive t(17;19)-ALL cell lines to VCR and DNR. Knockout of P-gp by CRISPRCas9 overcame resistance to VCR and DNR in the P-gp-positive t(17;19)-ALL cell line. A combination of cyclosporine A with DNR prolonged survival of NSG mice inoculated with P-gp-positive t(17;19)-ALL cell line. These findings indicate involvement of P-gp in resistance to VCR and DNR in Pgp positive t(17;19)-ALL cell lines. In all four t(17;19)-ALL cell lines, RAS pathway mutation was detected. Furthermore, among 16 t(1;19)-ALL cell lines, multiagent resistance was usually observed in the cell lines with RAS pathway mutation in comparison to those without it, suggesting at least a partial involvement of RAS pathway mutation in multiagent resistance of t(17;19)-ALL.

DOI： 10.1002/cam4.2356

PubMed

Authorship：Lead author,　Corresponding author   Language：English   Publishing type：Research paper (scientific journal)  

JMML is an aggressive hematopoietic malignancy of early childhood, and allogeneic HSCT is the only curative treatment for this disease. Umbilical cord blood is one of donor sources for HSCT in JMML patients who do not have an HLA-compatible relative, but engraftment failure remains a major problem. Here, we report two cases of JMML who were successfully rescued by HSCT from an HLA-mismatched parent after development of primary engraftment failure following unrelated CBT. Both patients had severe splenomegaly and underwent unrelated CBT from an HLA-mismatched donor. Immediately after diagnosis of engraftment failure, both patients underwent HSCT from their parent. For the second HSCT, we used RIC regimens consisting of FLU, CY, and a low dose of rabbit ATG with or without TBI and additionally administered ETP considering their persistent severe splenomegaly. Both patients achieved engraftment without severe treatment-related adverse effects. After engraftment of second HSCT, their splenomegaly was rapidly regressed, and both patients showed no sign of relapse for over 4 years. These observations demonstrate that HSCT from an HLA-mismatched parent could be a feasible salvage treatment for primary engraftment failure in JMML patients.

DOI： 10.1111/petr.13378

PubMed

Language：English   Publishing type：Research paper (scientific journal)   Publisher：American Association for Cancer Research (AACR)  

DOI： 10.1158/1078-0432.CCR-18-0919

PubMed

Language：English   Publishing type：Research paper (scientific journal)  

In many cancers, somatic mutations confer tumorigenesis and drug-resistance. The recently established clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 system is a potentially elegant approach to functionally evaluate mutations in cancers. To reproduce mutations by homologous recombination (HR), the HR pathway must be functional, but DNA damage repair is frequently impaired in cancers. Imatinib is a tyrosine kinase inhibitor for BCR-ABL1 in Philadelphia chromosome-positive (Ph+) leukemia, and development of resistance due to kinase domain mutation is an important issue. We attempted to introduce the T315I gatekeeper mutation into three Ph+ myeloid leukemia cell lines with a seemingly functional HR pathway due to resistance to the inhibitor for poly (ADP) ribose polymerase1. Imatinib-resistant sublines were efficiently developed by the CRISPR/Cas9 system after short-term selection with imatinib; resulting sublines acquired the T315I mutation after HR. Thus, the usefulness of CRISPR/Cas9 system for functional analysis of somatic mutations in cancers was demonstrated.

DOI： 10.1038/s41598-018-27767-6

PubMed

Language：English   Publishing type：Research paper (scientific journal)   Publisher：Blackwell Publishing Ltd  

Cytosine arabinoside (Ara-C) is one of the key drugs for the treatment of acute myeloid leukemia. It is also used for consolidation therapy of acute lymphoblastic leukemia (ALL). Ara-C is a deoxyadenosine analog and is phosphorylated to form cytosine arabinoside triphosphate (Ara-CTP) as an active form. In the first step of the metabolic pathway, Ara-C is phosphorylated to Ara-CMP by deoxycytidine kinase (DCK). However, the current cumulative evidence in the association of the Ara-C sensitivity in ALL appears inconclusive. We analyzed various cell lines for the possible involvement of DCK in the sensitivities of B-cell precursor ALL (BCP-ALL) to Ara-C. Higher DCK expression was associated with higher Ara-C sensitivity. DCK knockout by genome editing with a CRISPR-Cas9 system in an Ara-C-sensitive-ALL cell line induced marked resistance to Ara-C, but not to vincristine and daunorubicin, indicating the involvement of DCK expression in the Ara-C sensitivity of BCP-ALL. DCK gene silencing due to the hypermethylation of a CpG island and reduced DCK activity due to a nonsynonymous variant allele were not associated with Ara-C sensitivity. Clofarabine is a second-generation deoxyadenosine analog rationally synthesized to improve stability and reduce toxicity. The IC50 of clofarabine in 79 BCP-ALL cell lines was approximately 20 times lower than that of Ara-C. In contrast to Ara-C, although the knockout of DCK induced marked resistance to clofarabine, sensitivity to clofarabine was only marginally associated with DCK gene expression level, suggesting a possible efficacy of clofarabine for BCP-ALL that shows relative Ara-C resistance due to low DCK expression.

DOI： 10.1002/cam4.1323

Scopus

Language：English   Publishing type：Research paper (scientific journal)   Publisher：PUBLIC LIBRARY SCIENCE  

Prognosis of childhood acute lymphoblastic leukemia (ALL) has been dramatically improved. However, prognosis of the cases refractory to primary therapy is still poor. Recent phase 2 study on the efficacy of combination chemotherapy with bortezomib (BTZ), a proteasome inhibitor, for refractory childhood ALL demonstrated favorable clinical outcomes. However, septic death was observed in over 10% of patients, indicating the necessity of biomarkers that could predict BTZ sensitivity. We investigated in vitro BTZ sensitivity in a large panel of ALL cell lines that acted as a model system for refractory ALL, and found that Philadelphia chromosome-positive (Ph+) ALL, IKZF1 deletion, and biallelic loss of CDKN2A were associated with favorable response. Even in Ph-negative ALL cell lines, IKZF1 deletion and bilallelic loss of CDKN2A were independently associated with higher BTZ sensitivity. BTZ showed only marginal cross-resistance to four representative chemotherapeutic agents (vincristine, dexamethasone, l-asparaginase, and daunorubicin) in B-cell precursor-ALL cell lines. To improve the efficacy and safety of proteasome inhibitor combination chemotherapy, we also analyzed the anti-leukemic activity of carfilzomib (CFZ), a second-generation proteasome inhibitor, as a substitute for BTZ. CFZ showed significantly higher activity than BTZ in the majority of ALL cell lines except for the P-glycoprotein-positive t(17; 19) ALL cell lines, and IKZF1 deletion was also associated with a favorable response to CFZ treatment. P-glycoprotein inhibitors effectively restored the sensitivity to CFZ, but not BTZ, in P-glycoprotein-positive t(17; 19) ALL cell lines. P-glycoprotein overexpressing ALL cell line showed a CFZ-specific resistance, while knockout of P-glycoprotein by genome editing with a CRISPR/Cas9 system sensitized P-glycoprotein-positive t(17; 19) ALL cell line to CFZ. These observations suggested that IKZF1 deletion could be a useful biomarker to predict good sensitivity to CFZ and BTZ, and that CFZ combination chemotherapy may be a new therapeutic option with higher anti-leukemic activity for refractory ALL that contain P-glycoprotein-negative leukemia cells.

DOI： 10.1371/journal.pone.0188680

Web of Science

Language：English   Publishing type：Research paper (scientific journal)  

Here, we describe a case of primary graft failure with severe sepsis in a boy who experienced frequent relapses of osteosarcoma. The patient had undergone haploidentical bone marrow transplant after engraftment of unrelated cord blood transplant performed 10 months earlier. Considering his severe condition, we transfused autologous peripheral stem cells along with a single dose of etoposide (50 mg/m2). Granulocyte engraftment was confirmed on human leukocyte antigen-microsatellite analysis of bone marrow on day 14. Although the patient died due to respiratory failure, transfusion of autologous hematopoietic stem cells is a reasonable rescue option for graft failure even in patients whose background hematopoiesis is reconstituted by a first donor.

DOI： 10.6002/ect.2016.0315

PubMed

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY  

Activation of tyrosine kinase 2 (TYK2) contributes to the aberrant survival of T-cell acute lymphoblastic leukaemia (T-ALL) cells. Here we demonstrate the anti-leukaemic activity of a novel TYK2 inhibitor, NDI-031301. NDI-031301 is a potent and selective inhibitor of TYK2 that induced robust growth inhibition of human T-ALL cell lines. NDI-031301 treatment of human T-ALL cell lines resulted in induction of apoptosis that was not observed with the JAK inhibitors tofacitinib and baricitinib. Further investigation revealed that NDI-031301 treatment uniquely leads to activation of three mitogen-activated protein kinases (MAPKs), resulting in phosphorylation of ERK, SAPK/JNK and p38 MAPK coincident with PARP cleavage. Activation of p38 MAPK occurred within 1h of NDI-031301 treatment and was responsible for NDI-031301-induced T-ALL cell death, as pharmacological inhibition of p38 MAPK partially rescued apoptosis induced by TYK2 inhibitor. Finally, daily oral administration of NDI-031301 at 100mg/kg bid to immunodeficient mice engrafted with KOPT-K1 T-ALL cells was well tolerated, and led to decreased tumour burden and a significant survival benefit. These results support selective inhibition of TYK2 as a promising potential therapeutic strategy for T-ALL.

DOI： 10.1111/bjh.14563

Web of Science

Language：English   Publishing type：Research paper (scientific journal)   Publisher：BMJ PUBLISHING GROUP  

DOI： 10.1136/jclinpath-2016-203915

Web of Science

Language：English   Publishing type：Research paper (scientific journal)   Publisher：ELIFE SCIENCES PUBLICATIONS LTD  

Earlier reports showed that hyperplasia of sympathoadrenal cell precursors during embryogenesis in Nf1-deficient mice is independent of Nf1's role in down-modulating RAS-MAPK signaling. We demonstrate in zebrafish that nf1 loss leads to aberrant activation of RAS signaling in MYCN-induced neuroblastomas that arise in these precursors, and that the GTPase-activating protein (GAP)-related domain (GRD) is sufficient to suppress the acceleration of neuroblastoma in nf1-deficient fish, but not the hypertrophy of sympathoadrenal cells in nf1 mutant embryos. Thus, even though neuroblastoma is a classical "developmental tumor", NF1 relies on a very different mechanism to suppress malignant transformation than it does to modulate normal neural crest cell growth. We also show marked synergy in tumor cell killing between MEK inhibitors (trametinib) and retinoids (isotretinoin) in primary nf 1a-/- zebrafish neuroblastomas. Thus, our model system has considerable translational potential for investigating new strategies to improve the treatment of very high-risk neuroblastomas with aberrant RAS-MAPK activation.

DOI： 10.7554/eLife.14713

Web of Science

Language：Japanese   Publishing type：Research paper (scientific journal)  

Authorship：Lead author,　Last author,　Corresponding author   Language：Japanese   Publishing type：(MISC) Introduction and explanation (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)   Publisher：AMER ASSOC CANCER RESEARCH  

S-phase progression of the cell cycle is accelerated in tumors through various genetic abnormalities, and, thus, pharmacologic inhibition of altered cell-cycle progression would be an effective strategy to control tumors. In the current study, we analyzed the antileukemic activity of three available small molecules targeting CDK4/CDK6 against lymphoid crisis of chronic myeloid leukemia (CML-LC) and Philadelphia chromosome-positive acute lymphoblastic leukemia (Ph+ ALL), and found that all three molecules showed specific activities against leukemic cell lines derived from CML-LC and Ph+ ALL. In particular, PD0332991 exhibited extremely high antileukemic activity against CML-LC and Ph+ ALL cell lines in the nanomolar range by the induction of G(0)-G(1) arrest and partially cell death through dephosphorylation of pRb and downregulation of the genes that are involved in S-phase transition. As an underlying mechanism for favorable sensitivity to the small molecules targeting CDK4/CDK6, cell-cycle progression of Ph+ lymphoid leukemia cells was regulated by transcriptional and posttranscriptional modulation of CDK4 as well as Cyclin D2 gene expression under the control of BCR-ABL probably through the PI3K pathway. Consistently, the gene expression level of Cyclin D2 in Ph+ lymphoid leukemia cells was significantly higher than that in Ph+ lymphoid leukemia cells. Of note, three Ph+ ALL cell lines having the T315I mutation also showed sensitivity to PD0332991. In a xenograft model, PD0332991, but not imatinib, suppressed dissemination of Ph+ ALL having the T315I mutation and prolonged survival, demonstrating that this reagent would be a new therapeutic modality for relapsed CML-LC and Ph+ ALL patients after treatment with tyrosine kinase inhibitors. (C) 2015 AACR.

DOI： 10.1158/1535-7163.MCT-14-1065

Web of Science

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATURE PUBLISHING GROUP  

We previously found that tyrosine kinase 2 (TYK2) signaling through its downstream effector phospho-STAT1 acts to upregulate BCL2, which in turn mediates aberrant survival of T-cell acute lymphoblastic leukemia (T-ALL) cells. Here we show that pharmacologic inhibition of heat shock protein 90 (HSP90) with a small-molecule inhibitor, NVP-AUY922 (AUY922), leads to rapid degradation of TYK2 and apoptosis in T-ALL cells. STAT1 protein levels were not affected by AUY922 treatment, but phospho-STAT1 (Tyr-701) levels rapidly became undetectable, consistent with a block in signaling downstream of TYK2. BCL2 expression was downregulated after AUY922 treatment, and although this effect was necessary for AUY922-induced apoptosis, it was not sufficient because many T-ALL cell lines were resistant to ABT-199, a specific inhibitor of BCL2. Unlike ABT-199, AUY922 also upregulated the proapoptotic proteins BIM and BAD, whose increased expression was required for AUY922-induced apoptosis. Thus, the potent cytotoxicity of AUY922 involves the synergistic combination of BCL2 downregulation coupled with upregulation of the proapoptotic proteins BIM and BAD. This two-pronged assault on the mitochondrial apoptotic machinery identifies HSP90 inhibitors as promising drugs for targeting the TYK2-mediated prosurvival signaling axis in T-ALL cells.

DOI： 10.1038/leu.2015.222

Web of Science

Language：English   Publishing type：Research paper (scientific journal)   Publisher：AMER CHEMICAL SOC  

The Janus kinases (JAKs) and their downstream effectors, signal transducer and activator of transcription proteins (STATs), form a critical immune cell signaling circuit, which is of fundamental importance in innate immunity, inflammation, and hematopoiesis, and dysregulation is frequently observed in immune disease and cancer. The high degree of structural conservation of the JAK ATP binding pockets has posed a considerable challenge to medicinal chemists seeking to develop highly selective inhibitors as pharmacological probes and as clinical drugs. Here we report the discovery and optimization of 2,4-substituted pyrimidines as covalent JAK3 inhibitors that exploit a unique cysteine (Cys909) residue in JAK3. Investigation of structure activity relationship (SAR) utilizing biochemical and transformed Ba/F3 cellular assays resulted in identification of potent and selective inhibitors such as compounds 9 and 45. A 2.9 angstrom cocrystal structure of JAK3 in complex with 9 confirms the covalent interaction. Compound 9 exhibited decent pharmacokinetic properties and is suitable for use in vivo. These inhibitors provide a set of useful tools to pharmacologically interrogate JAK3-dependent biology.

DOI： 10.1021/acs.jmedchem.5b00710

Web of Science

Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY-BLACKWELL  

A 9-year-old boy with paroxysmal nocturnal hemoglobinuria/aplastic anemia syndrome (PNH/AA) developed hemolytic crisis after receiving immunosuppressive therapy. Eculizumab dramatically relieved the signs and symptoms and then he safely underwent unrelated bone marrow transplantation, suggesting the feasibility and effectiveness of eculizumab before stem cell transplantation in children with PNH/AA in hemolytic crisis.

DOI： 10.1111/ped.12522

Web of Science

Language：Japanese   Publishing type：Research paper (scientific journal)  

Authorship：Lead author   Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)   Publisher：PERGAMON-ELSEVIER SCIENCE LTD  

Since BCR-ABL plays an essential role in the growth factor-independent proliferation of Philadelphia chromosome (Ph)+leukemia cells, imatinib treatment of Ph+leukemia cells inactivates signaling pathways of BCR-ABL, and subsequent addition of growth factors (GFs) could restore the signaling pathways without reactivating BCR-ABL. Here we demonstrated that non-lymphoid Ph+leukemia cell lines responded to diverse GFs depending on their immunophenotype and gene expression of transcription factors and GF receptors, while lymphoid Ph+leukemia cell lines restrictively responded to flit3 ligand and interleukin-7, suggesting that GF sensitivity of imatinib-treated Ph+leukemia cells could be powerful for specifying their distinctive lineage. (C) 2012 Elsevier Ltd. All rights reserved.

DOI： 10.1016/j.leukres.2012.10.001

Web of Science

Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATURE PUBLISHING GROUP  

Allogeneic stem cell transplantation (allo-SCT) is a potentially curative therapy for chronic myeloid leukemia and Philadelphia chromosome-positive (Ph(+)) acute lymphoblastic leukemia, and the graft-vs-leukemia (GVL) effect can eradicate residual leukemia after allo-SCT. Ph(+) leukemia cells frequently express death-inducing receptors (DR4 and DR5) for tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), which is one of the cytotoxic ligands expressed on cytotoxic T cells and natural killer cells mediating the GVL effect. Here we demonstrate that imatinib specifically downregulated DR4 and DR5 expression in cell lines and clinical samples of Ph(+) leukemia. Second-generation tyrosine kinase inhibitors (dasatinib and nilotinib) and short hairpin RNA against bcr-abl also downregulated DR4 and DR5 expression in Ph(+) leukemia cells, and transfection of bcr-abl into a Ph(-) leukemia cell line induced DR4 and DR5 expression, which was abrogated by imatinib treatment. Accordingly, Ph(+) leukemia cells that had been pretreated with imatinib showed resistance to the pro-apoptotic activity of recombinant human soluble TRAIL. These observations demonstrate that BCR-ABL is critically involved in the leukemia-specific expression of DR4 and DR5 and in the susceptibility of Ph(+) leukemia to TRAIL-mediated anti-leukemic activity, providing new insight into the mechanisms of the tumor-specific cytotoxic activities of TRAIL. Oncogene (2013) 32, 1670-1681; doi:10.1038/onc.2012.186; published online 4 June 2012

DOI： 10.1038/onc.2012.186

Web of Science

Language：Japanese  

Language：Japanese  

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)   Publisher：WILEY-LISS  

DOI： 10.1002/ajh.21738

Web of Science

Language：English   Publishing type：Research paper (scientific journal)  

Language：Japanese  

Authorship：Lead author   Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATURE PUBLISHING GROUP  

DOI： 10.1038/leu.2010.8

Web of Science

Authorship：Lead author   Language：Japanese  

Language：English   Publishing type：Research paper (scientific journal)  

Language：English   Publishing type：Research paper (scientific journal)   Publisher：AMER ASSOC CANCER RESEARCH  

Fms-like tyrosine kinase 3 (FLT3) is highly expressed in acute lymphoblastic leukemia with the mixed-lineage leukemia (MLL) gene rearrangement refractory to chemotherapy. We examined the biological effect of FLT3-ligand (FL) on IS Bprecursor leukemic cell lines with variable karyotypic abnormalities, and found that nine of nine MLL-rearranged cell lines with wild-type FLT3, in contrast to other leukemic cell lines, are significantly inhibited in their proliferation in a dose-dependent manner by FL. This inhibition was due to induction of the GO-G, arrest. A marked up-regulation of p27 by suppression of its protein degradation and an abrogation of constitutive signal transducers and activators of transcription 5 phosphorylation were revealed in arrested leukemia cells after FL stimulation. Importantly, FL, treatment rendered not only cell lines but also primary leukemia cells with MLL rearrangement resistant to chemotherapeutic agents. WLLrearranged leukemia cells adhering to the bone marrow stromal cell line, which expresses FL, as the membrane-bound form, were induced to quiescent state resistant to chemotherapeutic agents, but their chemosensitivity was significantly restored in the presence of neutralizing anti-FL, antibody. The FL/FLT3 interaction between leukemia cells and bone marrow stromal cells expressing FL, at high levels should contribute, at least in part, to persistent minimal-residual disease of MLL-rearranged leukemia in bone marrow.

DOI： 10.1158/0008-5472.CAN-07-0105

Web of Science

Language：Japanese  

Language：English   Publishing type：Research paper (scientific journal)   Publisher：BLACKWELL PUBLISHING  

Neutrophil-specific antigen (NA) expression on neutrophils was analysed in 18 Japanese children before and after allogeneic stem cell transplantation (allo-SCT) with myeloablative regimen. Donor-recipient NA-incompatibility was present in one of eight NA1/NA2 heterozygous patients and eight of 10 NA1/NA1 or NA2/NA2 homozygous patients. After allo-SCTs from NA-incompatible donors, a neutrophil recipient-to-donor conversion was confirmed in all cases. Conversion to donor NA type was complete before the absolute neutrophil count reached 0.1 x 10(9)/l. These observations indicate that flow cytometric analysis of NA antigens is a simple and useful method for monitoring neutrophil engraftment in NA-incompatible allo-SCT.

DOI： 10.1111/j.1365-2141.2007.06778.x

Web of Science

Language：Japanese  

Language：English   Publishing type：Research paper (scientific journal)   Publisher：BLACKWELL PUBLISHING  

DOI： 10.1111/j.1365-2559.2007.02691.x

Web of Science

Authorship：Lead author   Language：Japanese  

Language：Japanese  

Language：Japanese  

Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATURE PUBLISHING GROUP  

Malignant cells generally acquire some immune escape mechanisms for clonal expansion. Immune escape mechanisms also contribute to the failure of graft-versus-leukemia (GVL) effect after allogeneic hematopoietic stem cell transplantation (allo-SCT). Infant leukemias with mixed-lineage leukemia (MLL) rearrangement have a remarkably short latency, and GVL effect after allo-SCT has not been clearly evidenced in these leukemias. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)- and FasL-mediated cytotoxic pathways play important roles in cytotoxic T-lymphocyte-and natural killer cell-mediated antitumor immunity and optimal GVL activity. We investigated the in vitro sensitivity of MLL-rearranged acute lymphoblastic leukemia (ALL) and acute myeloblastic leukemia (AML) cells to TRAIL- and FasL-mediated cytotoxicity. Most of cell lines and primary leukemia cells were highly resistant to TRAIL primarily owing to low cell-surface expression of death receptors in ALL and simultaneous expression of decoy receptors in AML. Nearly half of cell lines and majority of primary leukemia cells showed low sensitivity to FasL. These results suggest that resistance to death-inducing ligands, particularly to TRAIL, could be one of the mechanisms for a rapid clonal expansion and a poor sensitivity to the GVL effect in infant leukemias with MLL rearrangement.

DOI： 10.1038/sj.leu.2404429

Web of Science

Language：English   Publishing type：Research paper (scientific journal)   Publisher：NATURE PUBLISHING GROUP  

DOI： 10.1038/sj.leu.2404077

Web of Science

Language：Japanese  

Language：English   Publishing type：Research paper (scientific journal)   Publisher：CARDEN JENNINGS PUBL CO LTD  

Senescent erythrocytes undergo a loss of phospholipid asymmetry in the plasma membrane and are removed from the circulation by phagocytosis. To examine the loss of phospholipid asymmetry in mature erythrocytes after chemotherapy, we monitored phosphatidylserine (PS)-exposing erythrocytes by using flow cytometry to detect annexin V-bound erythrocytes in the circulation of acute lymphoblastic leukemia patients after consolidation chemotherapy. Both the percentage and the absolute number of annexin V-positive erythrocytes gradually increased immediately after chemotherapy. This result paralleled the change in the serum levels of bilirubin, suggesting a direct induction of PS-externalization in mature erythrocytes and a subsequent increase in erythrocyte clearance by splenic macrophages. The PS-exposing erythrocyte counts showed negative correlations to platelet and reticulocyte counts, which reflect the hematopoietic potential in the bone marrow. This result suggests that bone marrow suppression could be responsible for the enlarged senescent population in the circulation. Moreover, PS-exposing erythrocytes in the circulation remained at relatively high levels even after the serum level of bilirubin normalized, indicating that the decreased clearance of senescent erythrocytes as a result of impaired phagocytosis following bone marrow suppression might also be involved in the increase in PS-exposing erythrocytes in the circulation late after chemotherapy.

DOI： 10.1532/IJH97.05009

Web of Science

Language：Japanese  

Language：Japanese  

Language：Japanese   Publishing type：Research paper (scientific journal)  

Language：Japanese  

Language：Japanese  

Language：Japanese  

Responsible for pages：1133-1136   Language：Japanese   Book type：General book, introductory book for general audience

Responsible for pages：482-487   Language：Japanese   Book type：General book, introductory book for general audience

Responsible for pages：482-487   Language：Japanese   Book type：General book, introductory book for general audience

Event date： 2021.11

Language：Japanese   Presentation type：Poster presentation  

Event date： 2021.11

Language：Japanese   Presentation type：Symposium workshop panel(nominated)  

Event date： 2021.3

Language：Japanese   Presentation type：Poster presentation  

Event date： 2020.11

Language：Japanese   Presentation type：Poster presentation  

Event date： 2020.10

Language：English   Presentation type：Oral presentation(general)  

Event date： 2019.11

Language：English   Presentation type：Oral presentation(general)  

Event date： 2019.10

Language：English   Presentation type：Oral presentation(general)  

Venue：Tokyo  

Event date： 2018.11

Language：Japanese   Presentation type：Oral presentation(general)  

Event date： 2018.2

Language：Japanese   Presentation type：Poster presentation  

Venue：札幌  

Event date： 2017.7

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：松本  

Language：English   Presentation type：Poster presentation  

Venue：San Diego  

Event date： 2016.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜  

Event date： 2015.12

Language：English   Presentation type：Oral presentation(general)  

Event date： 2013.12

Language：English   Presentation type：Poster presentation  

Venue：New Orleans  

Event date： 2012.7

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：群馬県  

Event date： 2010.2

Language：Japanese   Presentation type：Poster presentation  

Venue：浜松  

Event date： 2009.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：浦安  

Event date： 2009.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：浦安  

Event date： 2009.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：浦安  

Event date： 2009.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：浦安  

Event date： 2009.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：浦安  

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：浦安  

Event date： 2009.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2009.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2009.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2009.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2009.10

Language：Japanese   Presentation type：Oral presentation(invited, special)  

Venue：宇都宮  

Event date： 2008.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2008.9

Language：Japanese   Presentation type：Oral presentation(general)  

Event date： 2007.12

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：仙台  

Event date： 2007.12

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：仙台  

Event date： 2007.12

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：仙台  

Event date： 2007.12

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：仙台  

Event date： 2007.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：甲府  

Event date： 2007.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜市  

Event date： 2007.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜市  

Event date： 2007.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：中央市  

Event date： 2007.6

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：鹿児島  

Event date： 2007.6

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：佐久  

Event date： 2007.3

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：中央市  

Event date： 2007.2

Language：Japanese   Presentation type：Poster presentation  

Venue：福岡  

Event date： 2006.12

Language：English   Presentation type：Oral presentation(general)  

Venue：Orlando  

Event date： 2006.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪  

Event date： 2006.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪  

Event date： 2006.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪  

Event date： 2006.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪  

Event date： 2006.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：松本市  

Event date： 2006.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：松本市  

Event date： 2006.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：福岡  

Event date： 2006.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：福岡  

Event date： 2006.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：福岡  

Event date： 2006.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：福岡  

Event date： 2006.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：福岡  

Event date： 2006.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：浜松  

Event date： 2006.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：中央市  

Event date： 2005.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：宇都宮  

Event date： 2005.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：宇都宮  

Event date： 2005.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：宇都宮  

Event date： 2005.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：宇都宮  

Event date： 2005.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：宇都宮  

Event date： 2005.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：栃木  

Event date： 2005.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：甲府  

Event date： 2005.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜  

Event date： 2005.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜  

Event date： 2005.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜  

Event date： 2005.6

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：奈良  

Event date： 2005.4

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：東京  

Event date： 2005.3

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：玉穂  

Event date： 2005.3

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：玉穂  

Event date： 2004.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2004.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2004.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2004.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2004.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2004.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都市  

Event date： 2004.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2004.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：京都  

Event date： 2004.6

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：豊科  

Event date： 2004.3

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：甲府  

Event date： 2003.12

Language：English   Presentation type：Oral presentation(general)  

Venue：San Diego  

Event date： 2003.12

Language：English   Presentation type：Oral presentation(general)  

Venue：San Diego  

Event date： 2003.12

Language：Japanese   Presentation type：Poster presentation  

Venue：福岡  

Event date： 2003.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：東京都  

Event date： 2003.11

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：東京  

Event date： 2003.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：金沢市  

Event date： 2003.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：金沢市  

Event date： 2003.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：金沢市  

Event date： 2003.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：金沢市  

Event date： 2003.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：金沢市  

Event date： 2003.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：金沢市  

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：金沢市  

Event date： 2003.8

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪市  

Event date： 2003.8

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪市  

Event date： 2003.8

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪市  

Event date： 2003.8

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪市  

Event date： 2003.8

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪市  

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪市  

Event date： 2002.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：大阪  

Event date： 2002.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：東京  

Event date： 2002.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：東京  

Event date： 2002.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：東京  

Event date： 2002.10

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：東京  

Event date： 2002.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜  

Event date： 2002.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜  

Event date： 2002.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜  

Event date： 2002.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：横浜  

Event date： 2001.9

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：山梨医科大学  

Event date： 2001.5

Language：Japanese   Presentation type：Oral presentation(general)  

Venue：仙台