大嶺 謙

Adoptive transfer of T cells expressing a chimeric antigen receptor (CAR) is a promising cell-based anticancer therapy. Although clinical studies of this approach show therapeutic efficacy, additional genetic modification is necessary to enhance the efficacy and safety of CAR-T cells. For example, production of an antitumor cytokine from CAR-T cells can potentially enhance their tumor-killing activity, but there are concerns that constitutive expression of anticancer molecules will cause systemic side effects. Therefore, it is important that exogenous gene expression is confined to the tumor locality. Here, we aimed to develop an inducible promoter driven by activation signals from a CAR. Transgene expression in T cells transduced with the CD19-targeted CAR and an inducible promoter, including inducible reporter genes (CAR-T/iReporter), was only induced strongly by co-culture with CD19-positive target cells. CAR-T/iReporter cells also showed redirected cytolysis toward CD19-positive, but not CD19-negative, tumor cells. Overall, our study indicated that the inducible promoter was selectively driven by activation signals from the CAR, and transduction with the inducible promoter did not affect original effector activities including interleukin-2 and interferon-γ production and the antitumor activity of CAR-redirected cytotoxic T lymphocytes. Moreover, this inducible promoter permits visualization and quantification of the activation status in CAR-T cells.

GALLIUM is a global phase III study that demonstrated a statistically significant and clinically meaningful improvement in progression-free survival (PFS) with obinutuzumab plus chemotherapy (G-chemo) versus rituximab plus chemotherapy (R-chemo) in previously untreated patients with follicular lymphoma (FL). In this single-country subgroup analysis, we explored patterns of efficacy and safety in patients enrolled in the GALLIUM study in Japan (Japanese subgroup). Patients were randomized to open-label induction treatment with G-chemo or R-chemo. Responders received maintenance monotherapy with their randomized antibody for up to 2 years. The primary endpoint was investigator-assessed PFS. Overall, 123 patients with FL were randomized in the Japanese subgroup (G-chemo, n = 65; R-chemo, n = 58). The majority of patients received cyclophosphamide, doxorubicin, vincristine, and prednisone chemotherapy (82.9 vs 33.1% in the global GALLIUM FL population). PFS at 3 years was 89.9% (G-chemo) vs. 74.7% (R-chemo); hazard ratio 0.42; 95% confidence interval 0.15, 1.15; P = 0.08. Higher rates of grade 3-5 adverse events (96.9 vs. 89.7%) and serious adverse events (35.4 vs. 22.4%) were observed with G-chemo vs R-chemo, respectively. Neutropenia was frequent in the Japanese subgroup (92.3% G-chemo; 79.3% R-chemo). Overall, the results in the Japanese subgroup were consistent with those in the global GALLIUM population.

The 1,3-beta-D-Glucan (BDG) assay is widely used for the diagnosis of fungal infections, especially in patients with hematologic malignancies. Some antimicrobials have been reported to cause false-positive results for BDG, but there has been no report on the effect of penicillin G (PCG) on BDG levels. We experienced a patient who developed false-positive BDG elevation during the administration of PCG for osteomyelitis due to Streptococcus pneumoniae infection. The serum BDG level increased up to 81.0 pg/ml during the continuous administration of PCG at 24 million units per day. However, chest and paranasal CT scan showed no evidence of fungal infection. The BDG level decreased to 38.0 pg/ml at 14 hours after the discontinuation of PCG. The amount of BDG in one vial of PCG inferred from these serum BDG levels is very similar to the actual BDG concentration in a vial of PCG. Therefore, during the administration of PCG, elevated BDG levels should be interpreted with caution, as they may be false-positive results.

Tyrosine kinase inhibitors (TKIs) are standard therapy for chronic myeloid leukemia (CML). However, the effects of these agents on mature B cell lymphoma are not well known. We describe a 50-year-old man who was diagnosed with CML in the chronic phase and treated with imatinib. After 3 years of imatinib therapy that achieved a complete cytogenetic response of CML, he developed Philadelphia-negative follicular lymphoma (FL). Rituximab monotherapy induced a partial response of FL, and he subsequently achieved a major molecular response (MMR) of CML. Three years later, however, the MMR was lost, followed by the progression of FL. Imatinib was switched to nilotinib for the treatment of CML, while we chose watchful waiting for FL. He achieved MMR again under treatment with nilotinib for 8 months including one month of substitutional use of dasatinib due to adverse events, but thereafter nilotinib was switched to bosutinib due to hyperbilirubinemia. With the administration of second-generation TKIs (2G-TKIs) for a total of 18 months, he achieved a complete response to FL without antilymphoma treatment. This is the first report to suggest that 2G-TKIs may have direct or indirect effects on FL.

In the ongoing, international, phase 3 study Evaluating Nilotinib Efficacy and Safety in Clinical Trials–Newly Diagnosed Patients (ENESTnd), nilotinib 300 and nilotinib 400 mg, both twice daily, are compared with imatinib 400 mg once daily for the treatment of newly diagnosed chronic myeloid leukemia in the chronic phase (CML-CP). Results for the overall population in ENESTnd (n = 846) showed that nilotinib resulted in higher response rates vs. imatinib and was well tolerated. Outcomes among Japanese patients in ENESTnd were specifically analyzed after 1 year of follow-up, and showed similar trends to the overall population we present updated analysis of the Japanese subgroup based on 5 years of follow-up. Among Japanese patients in the nilotinib 300-mg (n = 29), nilotinib 400-mg (n = 23), and imatinib (n = 25) arms, 86.2, 78.3, and 60.0%, respectively, achieved major molecular response [BCR-ABL1 ≤ 0.1% on the International Scale (BCR-ABL1IS)] by 5 years, and 65.5, 69.6, and 40.0%, respectively, achieved MR4.5 (BCR-ABL1IS ≤ 0.0032%). Safety results were consistent with prior reports. In this subgroup, one death occurred during treatment in the nilotinib 400-mg twice-daily arm (unknown cause), and one patient in each arm progressed to accelerated phase/blast crisis by the data cutoff.

Double-hit lymphoma (DHL) is defined as lymphoma with concurrent BCL2 and MYC translocations. While the most common histological subtype of DHL is diffuse large B-cell lymphoma, the present patient had leukemic follicular lymphoma (FL). A 52-year-old man was admitted to our hospital due to general fatigue and cervical and inguinal lymph node swelling. The patient was leukemic and the pathological diagnosis of the inguinal lymph node was FL grade 1. Chromosomal analysis revealed a complex karyotype including a rare three-way translocation t(8 14 18)(q24 q32 q21) involving the BCL2, MYC, and IGH genes. Based on a combination of fluorescence in situ hybridization (FISH), using BCL2, MYC and IGH, and spectral karyotyping (SKY), the karyotype was interpreted as being the result of a multistep mechanism in which the precursor B-cell gained t(14 18) in the bone marrow and acquired a translocation between der(14)t(14 18) and chromosome 8 in the germinal center, resulting in t(8 14 18). The pathological diagnosis was consistently FL, not only at presentation but even after a second relapse. The patient responded well to standard chemotherapies but relapsed after a short remission. This patient is a unique case of leukemic DH-FL with t(8 14 18) that remained in FL even at a second relapse.

Xenogeneic graft-versus-host disease (GVHD) models in highly immunodeficient mice are currently being used worldwide to investigate human immune responses against foreign antigens in vivo. However, the individual roles of CD4+ and CD8+ T cells, and donor/host hematopoietic and nonhematopoietic antigen-presenting cells (APCs) in the induction and development of GVHD have not been fully investigated. In the present study, we comprehensively investigated the immune responses of human T cells and the antigen presentation capacity of donor/host hematopoietic and nonhematopoietic APCs in xenogeneic GVHD models using nonobese diabetic/Shi-scid-IL2rgnul l mice. CD4+ T cells and, to a lesser extent, CD8+ T cells individually mediated potentially lethal GVHD. In addition to inflammatory cytokine production, CD4+ T cells also supported the activation and proliferation of CD8+ T cells. Using bone marrow chimeras, we demonstrated that host hematopoietic, but not nonhematopoietic, APCs play a critical role in the development of CD4+ T cell-mediated GVHD. During early GVHD, we detected 2 distinct populations in memory CD4+ T cells. One population was highly activated and proliferated in major histocompatibility complex antigen (MHC)+/+ mice but not in MHC−/− mice, indicating alloreactive T cells. The other population showed a less activated and slowly proliferative status regardless of host MHC expression, and was associated with higher susceptibility to apoptosis, indicating nonalloreactive T cells in homeostasis-driven proliferation. These observations are clinically relevant to donor T cell response after allogeneic hematopoietic stem cell transplantation. Our findings provide a better understanding of the immunobiology of humanized mice and support the development of novel options for the prevention and treatment for GVHD.

The early clearance of blast cells in peripheral blood (PB) during induction chemotherapy can predict the clinical outcome in acute leukemia. We retrospectively analyzed the kinetics of white blood cell (WBC) count, blast cell percentage (BCP), and blast cell count (BCC) in PB in 78 patients with de novo acute myeloid leukemia who underwent a uniform induction chemotherapy between December 2001 and December 2015 at Jichi Medical University. By a repeated-measures analysis of variance, the interaction of the decline in BCP with the achievement of complete remission (CR) was stronger than those of the decline in WBC or BCC. A receiver operating characteristic curve analysis for the achievement of CR showed that the areas under the curve for the decline in WBC, BCP, and BCC were 0.592, 0.703, and 0.634, respectively, and a decline in BCP of 9.25%/day within 4 or 5days from induction chemotherapy was the optimal cutoff value. A multivariate analysis showed that a rapid decline in BCP (9.25%/day) was a significant predictive factor for CR, independent of the cytogenetic risk (p=0.0096). A rapid decline in BCP during the first 5days of induction chemotherapy may be a good predictor of CR. Copyright (c) 2015 John Wiley & Sons, Ltd.

We retrospectively analyzed the relationship between white blood cell (WBC) count elevation after priming and clinical response in 115 patients with AML (61 untreated and 54 relapsed or refractory) treated with low-dose cytarabine, aclarubicin, and G-CSF priming. Receiver operating characteristic curve analysis showed that the ratio of maximum WBC count to pretreatment WBC count (WBCratio) was most strongly associated with complete remission (CR) in previously untreated patients among several parameters we analyzed in this study; however, the prediction accuracy was not clinically significant considering the area under the curve of 0.694. Based on the cutoff value of the WBCratio, CR rate and event-free survival in the high WBCratio group were significantly better than those in the low WBCratio group in untreated patients. Regarding the WBC differential counts, a high ratio of the maximum to pretreatment value of neutrophils rather than that of peripheral blasts was associated with a superior CR rate. In addition, an increase in blasts after G-CSF priming had a significant negative impact on CR rate in untreated patients. In conclusion, an increase in blast counts after G-CSF priming was not predictive of achieving CR.

The development of acute myeloid leukemia (AML) in patients with untreated chronic lymphocytic leukemia (CLL) is rare. We experienced a 65-year-old man who developed AML with aberrant CD7 expression and monoallelic CEBPA mutation during watchful waiting for CLL. He failed to achieve complete response (CR) by standard induction therapy for AML. We retrospectively reviewed 27 patients who developed AML with untreated CLL published between 1973 and 2016. The median age at diagnosis of AML was 68 years, and the median duration between the diagnoses of AML and CLL was 4.2 years. Diagnosis of AML and CLL was made simultaneously in 16 patients. The CR rate of AML was 42.9%, and the median survival was only 1.5 months after the diagnosis of AML. Patients who achieved CR tended to survive longer than those who did not. Our results demonstrated that the development of AML in patients with untreated CLL was associated with a poor response to chemotherapy and an extremely poor prognosis.

We evaluated 121 patients with follicular lymphoma (FL) and analyzed the association between the soluble interleukin-2 receptor (sIL-2R) level at diagnosis and the cumulative incidence of transformation. By a receiver-operating characteristic analysis, we determined a cutoff value of sIL-2R for transformation at 4360U/mL to classify patients into two groups. Patients in the high sIL-2R group showed a shorter progression-free survival (PFS) and shorter disease-specific survival (DSS) (p<0.001 and p=0.018). Furthermore, the cumulative incidence of transformation in the high sIL-2R group was higher than that in the low sIL-2R group (40.9% vs. 7.3% at 5 years, p<0.001). In a multivariate analysis, high sIL-2R was an independent predictive risk factor for transformation (HR 7.42, 95% CI: 2.75-20.0, p<0.001). This study showed that the sIL-2R level at diagnosis may be a prognostic factor for transformation, PFS, and DSS in patients with FL.

Objectives: CD25 has been reported to be highly expressed in leukemia stem cells and correlated with adverse outcomes in young patients with acute myeloid leukemia (AML). However, the significance of CD25 expression in elderly patients with AML has not yet been investigated. Methods: We retrospectively analyzed 154 newly diagnosed AML patients aged 60 years or over by flow cytometry. Results: CD25-positive AML was characterized by high white blood cell counts, secondary AML, rare favorable karyotypes, and positivity for CD34 and CD7 antigens, compared with CD25-negative AML. CD25 positivity was significantly correlated with an inferior complete remission (CR), event-free survival (EFS), and overall survival. Multivariate analysis showed CD25 positivity to be a significant prognostic predictor of CR and EFS. A regimen of low-dose cytarabine and aclarubicin combined with granulocyte-colony-stimulating factor (CAG) led to higher CR rates in the CD25-positive AML patients than intensive chemotherapies. CD25 expression was increased at relapse and in the development of leukemic status from myelodysplastic syndrome or myeloproliferative neoplasm. Discussion: An effective treatment strategy for elderly patients with CD25-positive AML has not been established. Further studies are needed to evaluate the effect of a CAG regimen and allogenic stem cell transplantation in patients. Conclusion: CD25 is an independent prognostic factor in elderly AML patients. Alternative therapies for CD25-positive elderly AML patients are needed.

The prognosis of patients with systemic lymphoma with central nervous system (CNS) involvement is very poor and there is no established standard therapy. We retrospectively analyzed 18 patients (4 untreated and 14 relapsed) with systemic lymphoma with CNS involvement who received methotrexate and cytarabine-based multiagent chemotherapy (modified Bonn protocol). Complete and partial responses were achieved in 56 and 22% of the patients, respectively. The 1-year overall survival (OS) and progression-free survival (PFS) was 81.0 and 39.2%, respectively. Patients with parenchymal involvement showed a better 1-year PFS than those with either leptomeningeal involvement or both. In a multivariate analysis, poor performance status (PS) was the only independent prognostic factor for the 1-year OS and PFS (HR 10.8, 95% CI 1.09-108, p = 0.042; HR 20.8, 95% CI 2.39-181, p = 0.006, respectively). Grade 4 neutropenia and thrombocytopenia occurred in 17 patients each (94%), but there were no grade 4 nonhematopoietic adverse events. The modified Bonn pro-tocol resulted in relatively favorable response and survival, and provided clinical benefits to patients with good PS, in particular. This study demonstrated that the modified Bonn protocol could be a feasible and encouraging treatment approach for lymphoma with CNS and systemic involvement. (C) 2017 S. Karger AG, Basel

Objectives: Follicular lymphoma (FL) is a clinically and biologically heterogeneous disease. Therefore, it is important to identify factors that can predict its clinical outcome. Methods: We retrospectively evaluated the usefulness of soluble interleukin-2 receptor (sIL-2R) levels after R-CHOP (posttreatment sIL-2R) in 72 patients with newly diagnosed FL who had either a complete response (CR) or partial response. With the use of a recursive partitioning analysis, we determined the cut-off values of post- and pretreatment sIL-2R levels that were associated with disease progression, which corresponded to 486.5 and 5405U/mL, respectively. Results: The high posttreatment sIL-2R group showed a significantly inferior progression-free survival (PFS) compared to the low posttreatment sIL-2R group in all patients (3-year PFS 52.6% vs. 77.4%, P=0.003), and in patients with CR (3-year PFS 57.1% vs. 82.1%, P=0.034). Although a multivariate analysis showed that pretreatment sIL-2R, but not posttreatment sIL-2R, was an independently significant predictive factor for disease progression, among patients with low pretreatment sIL-2R levels, those with high posttreatment sIL-2R levels tended to have inferior PFS. There was a significant trend in PFS among the high pretreatment sIL-2R group, the low pre- and high posttreatment sIL-2R group, and the low pre- and low posttreatment sIL-2R group (P<0.001). Conclusion: Among patients with a low pretreatment sIL-2R level who exhibited a positive response to R-CHOP, the posttreatment sIL-2R level may help to identify those with a poor prognosis.

The combination of mitoxantrone (MIT), etoposide (ETP), and cytarabine (Ara-C) (MEC) is a frequently used salvage therapy for acute leukemia, but has been associated with severe myelosuppression. Therefore, we investigated the miniMEC regimen with reduced doses of AraC and MIT. Thirteen ALL and 44 AML patients, all relapsed or refractory, received miniMEC, which consisted of MIT at 8 mg/m(2) for 3 d, ETP at 100 mg/m(2) for 5 d, and Ara-C at 100 mg/m(2) infused over 24 h for 7 d. CR + CRi was achieved in eight ALL patients (61.5%) and 16 AML patients (36.4%). Median duration of neutropenia was 30 d (range, 1-50). Thirty-one patients (54.4%) subsequently received allogeneic stem cell transplantation (SCT), and overall survival was significantly improved in this group (median OS 161 versus 481 d, p=0.006). We concluded that miniMEC is a safe and effective bridging therapy to SCT.

Anti-thymocyte globulin (ATG) is a key drug in immunosuppressive therapy for patients with aplastic anemia. The mainstay of ATG therapy had been a horse ATG (hATG) formulation, Lymphoglobulin or ATGAM, but Lymphoglobulin was recently discontinued, and Thymoglobulin, a rabbit ATG (rATG) formulation, is currently used as the first-line drug in many countries, including Japan. However, a recent randomized clinical trial reported significantly unfavorable outcomes associated with the use of rATG regimens. We retrospectively analyzed clinical outcomes of adult patients with moderate to severe aplastic anemia who were treated with 3.5 mg/kg of Thymoglobulin (n = 22) or 15 mg/kg of Lymphoglobulin (n = 25) in our facility. The estimated overall response rates in the rATG and hATG groups were 64.6 versus 56.0 % at 6 months, and 76.4 versus 69.2 % at 12 months, respectively; and there was no statistical difference between the two groups (P = 0.32). Overall survival at 24 months was not significantly different: rATG 89.8 % versus hATG 96.0 % (P = 0.39). Early phase infection was observed in 37.5 % of cases in the rATG and 14.8 % in the hATG group, but the frequency was not statistically different (P = 0.107). Our data indicate that Thymoglobulin at a dose of 3.5 mg/kg is a viable alternative when hATG is not available.

The BEAM regimen consisting of carmustine (BCNU), etoposide, cytarabine, and melphalan (MEL) is widely used before autologous hematopoietic stem cell transplantation (auto-HSCT) for lymphoma. However, intravenous BCNU is not available in Japan, and therefore, ranimustine (MCNU) has been used instead of BCNU (the MEAM regimen). We retrospectively analyzed the outcome of 79 adult patients who underwent auto-HSCT for lymphoma using this regimen in two centers, with 1- and 2-day dosing of MEL, respectively. Three-year overall survival (OS) and progression-free survival (PFS) probabilities were 77.3 and 56.5 % in the entire population and 71.7 and 58.0 % in patients with diffuse large B cell lymphoma. These outcomes were at least equivalent to those with the BEAM regimen. There was no regimen-related pulmonary toxicity. In a multivariate analysis, older age was the only factor that was significantly associated with for OS. In a comparison of the two MEL dosing schedules, while there was no significant differences in either OS or PFS, diarrhea was observed more frequently with 1-day dosing of MEL. In conclusion, the MEAM regimen appeared to be a promising conditioning regimen in auto-HSCT for lymphoma. A large prospective study is warranted to confirm the current findings.

This retrospective analysis compared the efficacy of intensive induction therapy consisting of daunorubicin and cytarabine (DNR-AraC) to that of less-intensive therapy including low-dose cytarabine, aclarubicin and granulocyte colony-stimulating factor priming (CAG). Patients aged 60 years or older who were newly diagnosed as acute myeloid leukemia (AML) were analyzed. Sixty-four and 48 patients were treated with DNR-AraC and CAG, respectively. The complete remission rates, 3-year overall survival and event-free survival in the DNR-AraC group were significantly superior to those in the CAG group (65.6% vs. 29.2%, p < 0.001, 38.4% vs. 12.3%, p = 0.0033, and 20.3% vs. 7.8%, p = 0.0030, respectively), although these differences were not statistically significant in multivariate analyses. Next, we calculated a propensity score for selecting the CAG regimen from six factors. The DNR-AraC regimen was associated with better survival than the CAG regimen in a low propensity score group, but there was no difference in survival between regimens in a high propensity score group. Intensive therapy should be performed for patients with sufficient general and comorbid conditions, but less-intensive therapy may be sufficient for patients with higher age, myelodysplasia-related changes, and lower white blood cell counts, which were relevant factors in the propensity score calculation. (C) 2015 Elsevier Ltd. All rights reserved.

<p>CD19に対するキメラ抗原受容体を発現させた遺伝子改変T細胞が再発・難治性のB細胞性腫瘍に対して有効であることが注目を集めている。2011年にCLLをはじめとした進行の遅い腫瘍のおよそ半数に有効であることが報告された。2012年にはB-NHLのおよそ8割に対しても同様に有効であることが報告された。2013年には治療が最も難しいと考えられていたB-ALLのおよそ9割に劇的な効果が報告された。つい最近では，通常CD19を発現していない多発性骨髄種に対しても自家移植と併用すると分子生物学的寛解を期待できる治療であることが報告された。一方で，サイトカイン放出症候群と中枢神経毒性は重篤な副作用として認識されている。本邦での臨床試験は昨年始まったばかりであるが，この分野の大きなブレークスルーになると期待されている新規治療法である。</p>

Erythropoiesis-stimulating agents (ESAs) are used to ameliorate anemia in lower-risk myelodysplastic syndromes (MDS). Serum erythropoietin (EPO) level &lt; 500 IU/L is widely accepted as a major predictive factor for response to ESAs. However, few data about EPO levels in the Japanese population are available. We therefore evaluated distribution of serum EPO levels in Japanese patients with MDS. Forty-three cases were analyzed; 30 were classified as lower-risk MDS (low or intermediate-1 by the international prognostic scoring system). Twenty-two cases were transfusion dependent. The overall median hemoglobin level was 7.7 g/dL. The median value of serum EPO was 254 IU/L (range: 16.4-23,000). Serum EPO levels had a strong inverse correlation with hemoglobin levels, and a significantly larger proportion of patients showed high EPO levels (&gt; 500 IU/L) in the transfusion-dependent group. In the higher-risk group, no significant correlation between EPO and hemoglobin was observed. Regression analyses showed that serum EPO of 500 IU/L corresponds to 8.29 g/dL of hemoglobin in lower-risk MDS. The results indicate that patients with hemoglobin levels of 8.0 g/dL or more, who are still transfusion independent, may be good candidates for ESA treatment.

The DNA methylation inhibitor azacitidine (5-azacytidine) is used against myelodysplastic syndrome and acute myeloid leukemia, but drug resistance is an ongoing, intractable problem. To investigate resistance mechanisms, we generated two azacitidine-resistant cell lines, THP-1/AR and HL60/AR, and studied genetic disparities between them and their corresponding parental lines. In cells treated with azacitidine, significant mitotic variations were noted in parental cells which were absent in resistant cells, suggesting that resistance arises from negating azacitidine-mediated activation of apoptosis signaling and reestablishing G2/M checkpoint. Importantly, both resistant cell lines have common point mutations in the uridine-cytidine kinase 2 (UCK2) gene, which encodes the rate-limiting enzyme of the azacitidine activation pathway. Forced expression of mutated UCK2 in parental THP-1 cells abrogated azacitidine-induced apoptosis, whereas overexpression of wild type UCK2 in resistant THP-1/AR cells restored sensitivity to azacitidine, implying that UCK2 gene mutations perturb azacitidine activation and advance azacitidine resistance. Our study provides new insights into azacitidine resistance and establishes models useful in developing effective strategies to overcome it. (C) 2014 ISEH - Society for Hematology and Stem Cells. Published by Elsevier Inc.

Cellular and gene therapies represent promising treatment strategies at the frontier of medicine. Hematopoietic stem cells, lymphocytes, and mesenchymal stem cells (MSCs) can all serve as sources of cells for use in such therapies. Strategies for gene therapy are often based on those of cell therapy, and it is anticipated that some examples will be put to practical use in the near future. Given their ability to support hematopoiesis, MSCs may be useful for the enhancement of stem cell engraftment, and the acceleration of hematopoietic reconstitution. Furthermore, MSCs may advance the treatment of severe graft-versus-host disease, based on their immunosuppressive ability. This application is also based on the homing behavior of MSCs to sites of injury and inflammation. Interestingly, MSCs possess tumor-homing ability, opening up the possibility of applications in the targeted delivery of anti-cancer genes to tumors. Many reports have indicated that MSCs can be utilized to target tumors and to deliver anti-cancer molecules locally, as tumors are recognized as non-healing wounds with inflammatory tissue. Here, we review both the potential of MSCs as cellular vehicles for targeted cancer therapy and the molecular mechanisms underlying MSC accumulation at tumor sites.

CD25 expression in follicular lymphoma (FL) has not yet been investigated. Eighty-five patients with newly diagnosed FL were retrospectively evaluated. On two-color flow cytometric analysis, CD25 was detected on CD19 + and CD20 + lymphoma cells. CD25 expression in FL tended to be higher than in reactive lymphadenopathy, but was lower than in diffuse large B-cell lymphoma. Patients with CD25 + FL (n = 12) showed clinical features of elevated soluble interleukin-2 receptor (IL-2R) levels, B symptoms and an advanced age compared with CD25 - FL (n = 73). The overall response rate (ORR), progression-free survival (PFS) and overall survival (OS) in patients with CD25 + FL were significantly inferior to those with CD25 - FL (ORR, 60 vs. 93%; 2-year PFS, 32 vs. 80.3%; 6-year OS, 47.4 vs. 85.9%, respectively). Multivariate analysis demonstrated that CD25 positivity is an independent prognostic factor for PFS and OS in FL. CD25 + FL may constitute a distinct subgroup associated with aggressiveness and an inferior prognosis.

Adoptive T-cell therapy with CD19-specific chimeric antigen receptors (CARs) is promising for treatment of advanced B-cell malignancies. Tumor targeting of CAR-modified T-cells is likely to contribute therapeutic potency; therefore we examined the relationship between the ability of CD19-specific CAR (CD19-CAR)-transduced T-cells to accumulate at CD19(+) tumor lesions, and their ability to provide antitumor effects in xenograft mouse models. Normal human peripheral blood lymphocytes, activated with immobilized RetroNectin and anti-CD3 antibodies, were transduced with retroviral vectors that encode CD19-CAR. Expanded CD19-CAR T-cells with a high transgene expression level of about 75% produced IL-2 and IFN-gamma in response to CD19, and lysed both Raji and Daudi CD19(+) human B-cell lymphoma cell lines. Furthermore, these cells efficiently accumulated at Raji tumor lesions where they suppressed tumor progression and prolonged survival in tumor-bearing Rag2(-/-)gamma c(-/-) immunodeficient mice compared to control cohorts. These results show that the ability of CD19-CAR T-cells to home in on tumor lesions is pivotal for their anti-tumor effects in our xenograft models, and therefore may enhance the efficacy of adoptive T-cell therapy for refractory B-cell lymphoma. (C) 2013 Elsevier Inc. All rights reserved.

CD25 (interluekin-2 receptor) expression in diffuse large B-cell lymphoma (DLBCL) cells has been not examined. To characterize CD25(+) DLBCL, 123 patients, who were newly diagnosed with DLBCL, were analyzed by single-color flow cytometry (FCM). CD25-positivity was significantly higher in DLBCL patients (n = 123; mean +/- SD, 27.8 +/- 30.6%) than in those with reactive lymphadenopathy (n = 16; mean +/- SD, 8.6 +/- 4.3%) and follicular lymphoma (n = 60; mean +/- SD, 12.7 +/- 12.4%). By two-color FCM, CD25/CD19 or CD25/CD20 dual positivity in DLBCL patients was shown: mean +/- SD, 63.7 +/- 25.5% (n = 13) and 55.0 +/- 28.1% (n = 14), respectively. Eighty-two percent of the patients with DLBCL received rituximab combined with CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone) chemotherapy. A cut-off value of 60% with CD25-positivity clearly divided patients with DLBCL into two groups: CD25-high or CD25-low DLBCL. Although clinical and immunophenotypic features were not significantly different in both groups, the former showed a significantly poorer response and more inferior progression-free survival than the latter. CD25 may be a new prognostic marker and could be a therapeutic target in DLBCL.

Bone marrow mononuclear cells from 93 patients with hematological malignancies after allogeneic hematopoietic stem cell transplantation (AHSCT) were analyzed using flow cytometry (FCM). The disease was acute myeloblastic leukemia (50 patients), acute lymphoblastic leukemia, and others. Conditioning was myeloablative (80 patients) or reduced intensity. The stem cell source was bone marrow (75 patients), peripheral blood stem cells, or cord blood. After AHSCT, granulocyte colony-stimulating factor was given to all patients. All patients showed engraftment of the donor cells. FCM was conducted on a median of 22 days after AHSCT. The gate was set around a granulocytic region consisting of immature granulocytes. The positivity rates of CD13, CD14, CD15, CD33, CD34, CD56, and HLA-DR in the cells were 59.9 ± 27.4%, 5.8 ± 8.8%, 98.3 ± 1.9%, 92.3 ± 12.4%, 2.6 ± 5.8%, 24.3 ± 16.7%, and 9.1 ± 6.6%, respectively. The greatest value of CD56 positivity was 73.1%. On the basis of CD56 expression, cases of more than 24% CD56 positivity were assigned to the CD56-high group (39 patients), while the rest were assigned to the CD56-low/negative group. There were no significant differences between the two groups in terms of disease status, sex, donor, hematopoietic stem cells, days of FCM analysis, or peripheral blood cell counts around the days of performing FCM. These results indicate that CD56 can be expressed in normal immature granulocytes at a variety of expression levels in regenerative bone marrow. Attention should be paid when evaluating aberrant antigen expression of CD56 in granulocytes. [J Clin Exp Hematop 53(3) : 247-250, 2013]

In the rituximab era, several large studies have suggested that full-dose rituximab plus cyclophosphamide, doxorubicin, vincristine and prednisone (R-CHOP) might be the best treatment for patients with diffuse large B-cell lymphoma (DLBCL) aged 60 years and older. However, it remains unclear whether this is also the case for those aged 70 years and older. Previously untreated patients with DLBCL aged 70 years and older (elderly) were treated with R-70%CHOP, and patients younger than 70 years (younger) were treated with full-dose R-CHOP every 3 weeks, for a total of 6-8 cycles. Complete remission (CR) rates in elderly versus younger patients were 75 vs. 78% (p = 0.7), respectively. The 3-year overall survival, event-free survival and progression-free survival of elderly versus younger patients were 58 vs. 78% (p &lt; 0.05), 45 vs. 70% (p &lt; 0.05) and 64 vs. 72% (p = 0.43), respectively. Severe adverse events were more frequent in the elderly, even with the dose reduction in that age group. Three-year PFS with R-70% CHOP for patients aged 70 years and older was not significantly worse than that with full-dose R-CHOP for younger patients, suggesting that R-70% CHOP might be a reasonable choice for patients with DLBCL aged 70 years and older, especially for those with comorbidities.

Aurora kinases play an essential role in the regulation of mitosis. The kinases are overexpressed in a variety of cancer cells and are involved in tumorgenesis. Although aurora kinase inhibitors are potential agents for treatment of leukemia, the establishment of efficacious combination therapies is an attractive approach for making good use of these agents. In this study, we examined the effects of a specific aurora kinase inhibitor, VE-465, in combination with various conventional anti-leukemia agents, including doxorubicin, daunorubicin, idarubicin, mitoxantron, cytosine arabinoside, vincristine and etoposide, on acute myeloid leukemia cell lines (HL60, U937, THP-1 and KY821), chronic myeloid leukemia cell lines (KCL22, K562 and KU812) and primary leukemia cells. We found that a combination of VE-465 and vincristine had a synergistic/additive inhibitory effect on the growth of leukemia cells. VE-465 initially increased G2/M-phase cells, followed by induction of sub-G1 cells. Vincristine enhanced this effect of VE-465. The combination of VE-465 and vincristine increased the levels of cleaved caspase 3, cleaved caspase 7, cleaved caspase 9, cleaved PARP and Phospho-Chk2, suggesting that the combination caused Chk2-mediated activation of the G2/M checkpoint, resulting in sequential induction of apoptosis. Interestingly, the combination markedly decreased the level of Phospho-ERK1/2, suggesting that the combination alters a network of cellular signaling pathways. In contrast, combinations of VE-465 and other agents showed no synergistic inhibitory effect but rather had an antagonistic effect. In conclusion, our results indicate the utility of the combination of VE-465 and vincristine as a potential therapy for myeloid leukemia. (C) 2011 Elsevier Inc. All rights reserved.

Vector integration can lead to proto-oncogene activation and malignancies during hematopoietic stem cell gene therapy. We previously used foamy virus vectors to deliver the CD18 gene under the control of an internal murine stem cell virus promoter and successfully treated dogs with canine leukocyte adhesion deficiency. Here we have tracked the copy numbers of 11 specific proviruses found in these animals for 36-42 months after transplantation, including examples within or near proto-oncogenes, tumor suppressor genes, and genes unrelated to cancer. We found no evidence for clonal expansion of any of the clones, including those with proviruses in the MECOM gene (MDS1-EVI1 complex). These results suggest that although foamy virus vectors may integrate near proto-oncogenes, this does not necessarily lead to clonal expansion and malignancies. Additionally, we show that copy number estimates of these specific proviruses based on linker-mediated PCR results are different from those obtained by quantitative PCR, but can provide a qualitative assessment of provirus levels.

According to the international working group response criteria for malignant lymphoma revised in 2007, 18F-fluorodeoxyglucose positron emission tomography ((18)FDG-PET) combined with or without computed tomography (CT) is recommended for pre-treatment staging and response assessment among patients with diffuse large B-cell lymphoma and Hodgkin lymphoma. Recently, along with the widespread use of PET/CT, unexpected uptake and accumulation of (18)FDG has been reported. Discussed in the present report are patients with malignant lymphoma and second primary carcinomas that were incidentally found by PET/CT. A total of 497 consecutive PET/CT were performed on 290 patients with malignant lymphoma in our institution from April 2008 through March 2010. Eight patients (2.8%) had pathologically confirmed second primary carcinomas consisting of 4 colon cancers, 3 lung cancers, and 1 pancreatic cancer. Two cases were diagnosed at the initial staging, and the others were detected after treatment for lymphoma. It is noteworthy that PET revealed high accumulations of (18)FDG in 5 (62.5%) of the 8 patients without corresponding tumors in conventional CT. All of the 4 patients with colon carcinoma underwent curative surgery. The present study suggests that incidental findings by PET in malignant lymphoma can lead to early detection and successful treatment of second malignancies.

Small molecules are attractive agents for the treatment of leukemia. We found that a combination of a farnesyltransferase inhibitor, tipifarnib, and an mTOR inhibitor, rapamycin, synergistically inhibited the growth of myeloid leukemia cell lines and primary leukemia cells by inducing apoptosis and cell-cycle blockage. The combined agents reduced the level of phospho-ERK1/2, suggesting that they altered the network of signaling pathways. They also showed synergistic effects in tipifarnib-resistant K562/RR cells. The results support the utility of this combination as a potential therapy for leukemia. The combination might also be effective in overcoming resistance to tipifarnib. (C) 2010 Elsevier Ltd. All rights reserved.

Heme plays an important biomodulating role in various cell functions. In this study, we examined the effects of hemin on cellular sensitivity to imatinib and other anti-leukemia reagents. Hemin treatment of human BCR/ABL-positive KCL22 leukemia cells increased IC50 values of imatinib, that is, the drug resistance, in a dose-dependent manner without any change in the BCR/ABL kinase activity. Imatinib-induced apoptosis was also suppressed by hemin treatment in KCL22 cells. Hemin treatment increased the activity of gamma-glutamylcysteine synthetase (gamma-GCS) light subunit gene promoter, which contains a Maf recognition element (MARE). Protein levels of gamma-GCS and heme oxygenase-1 (HO-1), two MARE-containing genes, were also increased after hemin treatment. Knockdown of Nrf2 expression by RNA interference largely abolished the effect of hemin on imatinib-treated cells, suggesting that Nrf2 recognition of MARE is essential for the hemin-mediated protective effect. Similar to hemin, treatment of cells with delta-aminolevulinic acid (delta-ALA), the obligatory heme precursor, also increased IC50 values of imatinib. In contrast, inhibition of cellular heme synthesis by succinylacetone increased the sensitivity of cells to imatinib in two imatinib-resistant cell lines, KCL22/SR and KU812/SR. Hemin treatment also decreased the sensitivity of cells to four anthracyclins, daunorubicin, idarubicin, doxorubicin, and mitoxantrone, in BCR/ABL-negative leukemia U937 and THP-1 cells, as well as in KCL22 cells. These findings thus indicate that cellular heme level plays an important role in determining the sensitivity of cells to imatinib and certain other anti-leukemia drugs and that the effect of heme may be mediated via its ability to upregulate Nrf2 activity.