Establishment and characterization of a MALT lymphoma cell line carrying an API2-MALT1 translocation.

MALT lymphomas with API2(BIRC3)-MALT1 translocation usually have an indolent clinical course and rarely transform into aggressive lymphoma, and there have been no lymphoma cell lines carrying API2-MALT1 translocation reported to date. We established a novel lymphoma cell line named BMA19, carrying the API2-MALT1 translocation from a patient with histologic transformation of intestinal MALT lymphoma. The cells were suggested to carry API2-MALT1 and MYC-IGH translocations by chromosomal analysis, and these translocations were confirmed by polymerase chain reaction analysis. The expression of MYC was shown to be enhanced as a result of the MYC-IGH translocation, and it is considered to have played a role in the histologic transformation of MALT lymphoma. Whole exome sequencing of BMA19 identified several nucleotide variations in genes reported to be mutated in previous studies of marginal zone lymphomas. The MALT1 inhibitor MI-2 specifically decreased cell growth, and the BMA19 cell line was suggested to be still dependent on the API2-MALT1 signal. Subtractive microarray analysis showed that one of the earliest events resulting from MALT1 inhibition is increased susceptibility to endoplasmic reticulum stress-induced apoptosis. The BMA19 cell line is considered to conserve the biological properties of MALT lymphoma and is expected to be a valuable tool for research into the pathogenesis of MALT lymphoma with an API2-MALT1 translocation.

Outcomes of allogeneic stem cell transplantation for DLBCL: a multi-center study from the Kyoto Stem Cell Transplantation Group.

Allogeneic hematopoietic stem cell transplantation (allo-SCT) has been considered as a potentially curative treatment option for refractory or relapsed diffuse large B cell lymphoma (DLBCL) patients. However, there is little information available, especially for Japanese patients and in cord blood transplantation (CBT). We aimed to determine treatment outcomes of allo-SCT for DLBCL in the Kyoto Stem Cell Transplantation Group, a multi-institutional joint research group. Sixty-eight DLBCL patients who underwent their first allo-SCT between 2003 and 2016 were included. The median time from diagnosis to transplantation was 13.5 months. Thirty-one patients were in CR/PR at transplantation. Twenty-seven patients underwent CBT. The median follow-up for survivors was 44.2 months. Four-year overall survival (OS) and relapse-free survival (RFS) rates were 23% (95% CI, 13-35%) and 20% (95% CI, 11-31%), respectively. Cumulative incidences of non-relapse mortality and relapse were 23% and 57%, respectively. Patients in CR/PR at allo-SCT had better OS (4-year, 46% vs 4%, P < 0.001) and RFS (4-year, 36% vs 7%, P = 0.005). The source of the stem cell did not significantly affect OS (4-year, bone marrow vs cord blood vs peripheral blood, 28.6% vs 27.2% vs 6.5%, P = 0.193). In multivariate analysis, non-remission status at SCT associated with inferior OS and RFS. Duration from diagnosis to transplantation of less than 1 year associated with inferior RFS. Allo-SCT, including CBT, may be a promising therapeutic modality for DLBCL patients who have good disease control at transplantation.

Direct Introduction of a Dimesitylboryl Group Using Base-Mediated Substitution of Aryl Halides with Silyldimesitylborane.

The first dimesitylboryl substitution of aryl halides with a silylborane bearing a dimesitylboryl group in the presence of alkali-metal alkoxides is described. The reactions of aryl bromides or iodides with Ph2 MeSi-BMes2 and Na(OtBu) afforded the desired aryl dimesitylboranes in good to high yields and with high borylation/silylation ratios. Selective reaction of the sterically less-hindered C-Br bond of dibromoarenes provided monoborylated products. This reaction was used to rapidly construct a D-π-A aryl dimesityl borane with a non-symmetrical biphenyl spacer.

Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia Is Separated into Two Subgroups Associated with Survival by BCR-ABL Fluorescence in situ Hybridization of Segmented Cell Nuclei: Report from a Single Institution.

Philadelphia chromosome-positive (Ph+) acute lymphoblastic leukemia (ALL) may include the lymphoid blast crisis of chronic myeloid leukemia (CML-BC). We applied fluorescence in situ hybridization (FISH) of the BCR-ABL fusion gene to peripheral blood and/or bone marrow smears to determine whether the fusion was restricted to mononuclear cell nuclei or if segmented cell nuclei representing mature neutrophils also carried the fusion (Seg-FISH). Among 20 patients with Ph+ ALL without a prior diagnosis of CML, 9 were Seg-FISH+ and 11 were Seg-FISH-. Seg-FISH+ cases were characterized by a higher rate of p210-type BCR-ABL transcripts, higher white cell and blast counts, and a higher rate of myeloid and T-lymphoid antigen expression than Seg-FISH- cases, in addition to 'major route' cytogenetic abnormalities associated with CML-BC. Eighteen patients were treated with tyrosine kinase inhibitors (TKIs) either alone or in combination with multiagent chemotherapy, and 7 underwent allogeneic hematopoietic stem cell transplantation. Progression-free and overall survivals were greater in the Seg-FISH+ group than in the Seg-FISH- group. These results suggest that the Seg-FISH+ group represents lymphoid CML-BC that occurs de novo, while the Seg-FISH- represents Ph+ ALL in the strict sense, and the two groups are associated with survival when treated with TKIs or TKI-combined therapy.

Outcomes of very elderly patients with aggressive B-cell non-Hodgkin lymphoma treated with reduced-dose chemotherapy.

BACKGROUND: Although the number of patients aged 80 years or older with aggressive B-cell non-Hodgkin lymphoma (B-NHL) has increased in the clinical setting, management has been challenging due to lower tolerability. The aim of the present study was to evaluate the efficacy and safety of reduced-dose chemotherapy for very elderly patients. METHODS: We retrospectively analyzed the clinical outcomes of patients aged ≥80 years old with diffuse large B-cell lymphoma (n = 58) or grade 3 follicular lymphoma (n = 3). RESULTS: Patient ages ranged from 80 to 93 years old, with a median of 83 years old. Twenty-four patients were treated with CHOP or THP-COP, the dosages of which were variably reduced, in combination with rituximab (R-vCHOP), while another 24 patients were treated with R-miniCHOP. Twelve R-vCHOP and 16 R-miniCHOP patients completed the chemotherapy cycles. The estimated 2-year progression-free and overall survival rates in the R-vCHOP and R-miniCHOP groups were 53 and 39 % (P = 0.92) and 53 and 48 % (P = 0.95), respectively. Performance status ≥2, lactate dehydrogenase level >normal, serum albumin ≤3.5 g/dL, C-reactive protein ≥2.0 mg/dL, age-adjusted International Prognostic Score 2/3, and withdrawal from the chemotherapy cycle were associated with poor survival. The frequency of chemotherapy-related hospitalization during the second or later cycles was significantly less in the R-miniCHOP group. CONCLUSIONS: The efficacies of R-vCHOP and R-miniCHOP were similar in patients aged ≥80 years old with aggressive B-NHL. The reduced frequency of hospitalization observed for R-miniCHOP treatment is beneficial for very elderly patients.

Mild Acute Graft-Versus-Host Disease Improves Outcomes After HLA-Haploidentical-Related Donor Transplantation Using Posttransplant Cyclophosphamide and Cord Blood Transplantation.

Haploidentical-related donor transplantation using posttransplant cyclophosphamide (PTCy-haplo) and cord blood transplantation (CBT) are valid alternatives for patients with hematological malignancies when HLA-matched donor transplantation (MDT) is unavailable. However, the effects of graft-versus-host disease (GVHD) on outcomes after these transplants have not been fully elucidated. Therefore, we evaluated the effects of acute and chronic GVHD on transplant outcomes after PTCy-haplo transplants and compared them with CBT and MDT. We included a total of 914 adult patients with hematological malignancies in the Kyoto Stem Cell Transplantation Group registry who received PTCy-haplo (N = 120), CBT (N = 402), and MDT (N = 392), and achieved neutrophil engraftment. A multivariate analysis revealed that grade I-II acute GVHD improved of overall survival (OS) after PTCy-haplo [hazard ratio (HR) = 0.39, P = 0.018] and CBT (HR = 0.48, P < 0.001), but not after MDT (HR = 0.80, P = 0.267) compared with patients without acute GVHD. Grade I-II acute GVHD had a trend toward reducing the risk of nonrelapse mortality (NRM) after PTCy-haplo (HR = 0.13, P = 0.060) and this positive effect was significant after CBT (HR = 0.39, P = 0.003). A negative impact of grade III-IV acute GVHD on NRM was observed after CBT and MDT, but not after PTCy-haplo. Limited chronic GVHD had a positive impact on OS after CBT and MDT, but not after PTCy-haplo. In conclusion, mild acute GVHD improved outcomes after PTCy-haplo and CBT, and limited chronic GVHD improved outcomes after CBT and MDT. These data indicated that the effects of GVHD on transplant outcomes depended on transplant platforms.

Anomalous reactivity of silylborane: transition-metal-free boryl substitution of aryl, alkenyl, and alkyl halides with silylborane/alkoxy base systems.

An unexpected borylation of organic halides with a silyborane in the presence of an alkoxy base has been observed. This formal nucleophilic boryl substitution can be applied to a broad range of substrates with high functional group compatibility. Even sterically hindered aryl bromides afforded the corresponding boryl compounds in high yields. Preliminary mechanistic studies indicated that this boryl substitution is promoted by neither transition-metal contamination nor a radical-mediated process.

Development of a novel Indium-111 radiolabeled mogamulizumab targeting CCR4 for imaging adult T-cell leukemia/lymphoma in vivo.

OBJECTIVE: Adult T-cell leukemia/lymphoma (ATL), caused by human T-cell lymphotropic virus type I (HTLV-1) infection, is among the most aggressive categories and has the worst prognosis among T-cell lymphomas. Mogamulizumab, an anti-CC chemokine receptor 4 (CCR 4), has been shown to be effective in the treatment of ATL; however, some ATL cases are often resistant, particularly the lymphoma-type ATL. To evaluate drug delivery in vivo and identify the distribution of CCR4-positive cells in the body, we developed a novel mogamulizumab tracer labeled with Indium-111 (111In) via diethylenetriaminepentaacetic acid (DTPA) for single-photon emission computerized tomography (SPECT), named [111In]In-DTPA-mogamulizumab, and evaluated its potential for visualizing CCR4 expression in vivo. METHODS: [111In]In-DTPA-mogamulizumab was added to HCT116/CCR4 or HCT116/empty vector (EV) cells, and their radioactivity was measured 1 h after administration. A blocking study was additionally performed by treating HCT116/CCR4 cells with excess mogamulizumab in addition to [111In]In-DTPA-mogamulizumab. The biodistribution and SPECT imaging of [111In]In-DTPA-mogamulizumab in HCT116/CCR4 and HCT116/EV dual-xenografted BALB/c-nu mice were evaluated for 72 h after intravenous injection. RESULTS: [111In]In-DTPA-mogamulizumab was acquired with a radiochemical purity > 95%. The cellular uptake level of [111In]In-DTPA-mogamulizumab by HCT116/CCR4 cells was significantly higher than that by HCT116/EV cells (HCT116/CCR4: 0.951 ± 0.069, HCT116/EV: 0.006 ± 0.001%dose/mg protein, p < 0.01), and the uptake was significantly suppressed by co-incubation with excess mogamulizumab (0.013 ± 0.003%dose/mg protein, p < 0.01). In the in vivo study, the radioactivity of the HCT116/CCR4 tumor tissue was significantly higher than that of the HCT116/EV tumor tissue at 72 h after the administration of [111In]In-DTPA-mogamulizumab (HCT116/CCR4: 20.5 ± 5.4, HCT116/EV: 5.7 ± 1.0%ID/g), and HCT116/CCR4 tumors were clearly and specifically visualized on SPECT imaging. CONCLUSIONS: We have successfully developed a novel SPECT imaging tracer targeting CCR4, [111In]In-DTPA-mogamulizumab, which showed good specificity and pharmacokinetics, indicating potential in visualizing CCR4 expression in vivo.

KIR3DL1 Allotype-Dependent Modulation of NK Cell Immunity against Chronic Myeloid Leukemia.

Tyrosine kinase inhibitor (TKI)-treated chronic myeloid leukemia (CML) patients with increased NK cell number have a better prognosis, and thus, NK cells may suppress CML. However, the efficacy of TKIs varies for reasons yet to be fully elucidated. As NK cell activity is modulated by interactions between their killer cell Ig-like receptors (KIRs) and HLAs of target cells, the combination of their polymorphisms may have functional significance. We previously showed that allelic polymorphisms of KIR3DL1 and HLAs were associated with the prognosis of TKI-treated CML patients. In this study, we focus on differential NK cell activity modulation through KIR3DL1 allotypes. KIR3DL1 expression levels varied according to their alleles. The combination of KIR3DL1 expression level and HLA-Bw4 motifs defined NK cell activity in response to the CML-derived K562 cell line, and Ab-mediated KIR3DL1 blocking reversed this activity. The TKI dasatinib enhanced NK cell activation and cytotoxicity in a KIR3DL1 allotype-dependent manner but did not significantly decrease effector regulatory T cells, suggesting that it directly activated NK cells. Dasatinib also enhanced NK cell cytotoxicity against K562 bearing the BCR-ABL1 T315I TKI resistance-conferring mutation, depending on KIR3DL1/HLA-Bw4 allotypes. Transduction of KIR3DL1*01502 into the NK cell line NK-92 resulted in KIR3DL1 expression and suppression of NK-92 activity by HLA-B ligation, which was reversed by anti-KIR3DL1 Ab. Finally, KIR3DL1 expression levels also defined activation patterns in CML patient-derived NK cells. Our findings raise the possibility of a novel strategy to enhance antitumor NK cell immunity against CML in a KIR3DL1 allotype-dependent manner.

EZH2 inhibitors restore epigenetically silenced CD58 expression in B-cell lymphomas.

Loss of CD58 is a common mechanism for tumor immune evasion in lymphoid malignancies. CD58 loss is known to occur due to both genetic and non-genetic causes; therefore, we hypothesized that restoring CD58 expression in lymphoma cells may be an effective treatment approach. To explore the potential for restoring CD58 expression, we first screened 11 B-cell lymphoma lines and found that 3 had decreased CD58 expression. Among these, CD58 was genetically damaged in two lines but not in the third line. Using the cell line with downregulated CD58 without a genetic abnormality, we performed epigenetic library screening and found that two EZH2 inhibitors, EPZ6438 and GSK126, specifically enhanced CD58 expression. By examining the effect of three EZH2 inhibitors with different selectivity profiles in different B-cell lines, EZH2 inhibition was shown to have a common activity in upregulating CD58 expression. Restoring the expression of CD58 in lymphoma cells using an EZH2 inhibitor was shown to enhance interferon-γ production of T and NK cells against lymphoma cells. H3K27 was shown to be highly trimethylated in the CD58 promoter region, and EZH2 inhibition induced its demethylation and activated transcription of the CD58 gene. These results indicated that EZH2 is involved in the epigenetic silencing of CD58 in lymphoma cells as a mechanism for tumor immune escape, and EZH2 inhibitors are able to restore epigenetically suppressed CD58 expression. Our findings provide a molecular basis for the combination of an EZH2 inhibitor and immunotherapy for lymphoma treatment.

B cells with aberrant activation of Notch1 signaling promote Treg and Th2 cell-dominant T-cell responses via IL-33.

The Notch-signaling pathway in a variety of mature B-cell neoplasms is often activated by gene alterations, but its role remains unclear. Here, we show that B cells harboring dysregulated activation of Notch1 signaling have an immunomodulatory effect on T cells by amplifying regulatory T (Treg) and T helper 2 (Th2) cell responses in an interleukin-33 (IL-33)-dependent manner. A conditional mouse model, in which constitutive expression of an active form of Notch1 is induced in B cells by Aicda gene promoter-driven Cre recombinase, revealed no obvious phenotypic changes in B cells; however, mice demonstrated an expansion of Treg and Th2 cell subsets and a decrease in cytokine production by Th1 and CD8+ T cells. The mice were susceptible to soft tissue sarcoma and defective production of CD8+ T cells specific for inoculated tumor cells, suggesting impaired antitumor T-cell activity. Gene-expression microarray revealed that altered T-cell responses were due to increased IL-33 production by Notch1-activated B cells. Knockout of IL33 or blockade of IL-33 by a receptor-blocking antibody abrogated the Treg and Th2 cell-dominant T-cell response triggered by B cells. Gene-expression data derived from human diffuse large B-cell lymphoma (DLBCL) samples showed that an activated Notch-signaling signature correlates positively with IL33 expression and Treg cell-rich gene-expression signatures. These findings indicate that B cells harboring dysregulated Notch signaling alter T-cell responses via IL-33, and suggest that aberrant activation of Notch signaling plays a role in fostering immune privilege in mature B-cell neoplasms.

A unique subtype of diffuse large B-cell lymphoma primarily involving the bone marrow, spleen, and liver, defined by fluorodeoxyglucose-positron emission tomography combined with computed tomography.

We describe 10 cases of diffuse large B-cell lymphoma (DLBCL) confined to the bone marrow (BM), spleen, and liver, as evidenced by the uniformly increased uptake of fluorodeoxyglucose (FDG) on positron emission tomography combined with computed tomography (PET/CT). Ages ranged from 56 to 87. All, but one patient presented with 'B' symptoms, a poor performance status, and hepatosplenomegaly. All patients showed cytopenia and elevated lactate dehydrogenase levels and were classified into the high-risk category of the International Prognostic Index scoring. BM infiltration was diffuse, interstitial/intrasinusoidal, or mixed, and all showed the nongerminal center B immunophenotype. Five patients had a rearrangement involving 3q27/BCL6, while six had increased copies of MYC, BCL2, or BCL6. All patients were initially treated with rituximab, cyclophosphamide, doxorubicin, vincristine, and prednisolone, leading to complete responses in six out of eight evaluable patients. We propose BM, spleen, and liver-type DLBCL, which is defined by the findings of FDG-PET/CT.

Modular fuzzy-neuro controller driven by spoken language commands.

We present a methodology of controlling machines using spoken language commands. The two major problems relating to the speech interfaces for machines, namely, the interpretation of words with fuzzy implications and the out-of-vocabulary (OOV) words in natural conversation, are investigated. The system proposed in this paper is designed to overcome the above two problems in controlling machines using spoken language commands. The present system consists of a hidden Markov model (HMM) based automatic speech recognizer (ASR), with a keyword spotting system to capture the machine sensitive words from the running utterances and a fuzzy-neural network (FNN) based controller to represent the words with fuzzy implications in spoken language commands. Significance of the words, i.e., the contextual meaning of the words according to the machine's current state, is introduced to the system to obtain more realistic output equivalent to users' desire. Modularity of the system is also considered to provide a generalization of the methodology for systems having heterogeneous functions without diminishing the performance of the system. The proposed system is experimentally tested by navigating a mobile robot in real time using spoken language commands.

inv(2)(p23q13)/RAN-binding protein 2 (RANBP2)-ALK fusion gene in myeloid leukemia that developed in an elderly woman.

A 75-year-old woman presented with marked leukocytosis; the white cell count was 143.6 × 10³/µL with 38.6 % monocytes and 13.6 % immature granulocytes, including blasts. Bone marrow (BM) aspirate smears showed >90 % cellularity with hyperplasia of myeloid-lineage cells, 14.6 % monocytes, and 32.1 % blasts. The granulocyte series showed a range of dysplastic morphologies. The rate of peroxidase positivity was 51.5 %. CD36+ cells with monocytic differentiation comprised 64.6 % mononuclear cells. Metaphase spreads obtained from the BM revealed an aneuploid karyotype with -7 and a submetacentric marker chromosome derived from chromosome 2, which was determined to be inv(2)(p23q13) by fluorescence in situ hybridization using the Vysis ALK probe. RAN-binding protein 2 (RANBP2)-ALK fusion mRNA was confirmed by reverse transcriptase-mediated polymerase chain reaction and nucleotide sequencing. High-sensitivity anti-ALK immunohistochemistry of a BM biopsy specimen demonstrated nuclear membrane staining of leukemia cells. As the leukemia showed features of chronic myelomonocytic leukemia, the patient was treated with standard daunorubicin-cytarabine followed by azacitidine, leading to the durable suppression of leukemia progression. These findings suggest that inv(2)(p23q13)/RABBP2-ALK defines a small subset of myeloid leukemia characterized by differentiation to monocytes and sharing features of myelodysplastic syndrome/myeloproliferative neoplasm.