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  • American Society of Hematology  (35)
  • Medicine  (35)
  • 1
    In: Blood, American Society of Hematology, Vol. 122, No. 21 ( 2013-11-15), p. 1528-1528
    Abstract: The Wilms' tumor gene (WT1), originally discovered as a tumor suppressor has been proven to have an oncogenic role in leukemia and several other cancers. WT1 mRNA expression levels in peripheral blood (PBWT1) has been reported as a useful marker for the risk evaluation of myelodysplastic syndrome (MDS). In the era of hypomethylating agents, the significance of PBWT1 on MDS prognosis is still unknown. This study aimed to clarify the impact of pre-treatment PBWT1 levels on overall response (OR) and overall survival (OS) in MDS patients treated with azacitidine (AZA). Patients and Methods We retrospectively analyzed all patients from March 2011 to March 2013 with World Health Organization 2008 defined MDS, CMML or AML with 20–30% bone marrow blasts who received AZA treatment in our department for at least one cycle (37.5–75.0 mg/m2/day during 7 days, every 28 days). Patients' peripheral blood specimens were collected before AZA initiation, mRNA was extracted from leukocytes using the RNeasy Mini-Kit (Qiagen, Valencia, CA), and the amount containing WT1 mRNA was measured using a WT1 mRNA Assay Kit (Otsuka Pharmaceutical Co., Ltd., Tokyo, Japan). Hematologic response was evaluated according to International Working Group 2006. OR was defined as a best overall response of complete remission (CR), partial remission, marrow CR, or hematologic improvement. Univariate analyses for OR were carried out using Fisher's exact test. Factors associated with at least borderline significance (p 〈 0.10) were subjected to a multivariate analysis, using logistic regression model. OS was estimated according to the Kaplan-Meier method. Multivariate analysis was performed with proportional hazard Cox model, including all variables with p 〈 0.10 in univariate analyses. Results Of 55 patients enrolled, pre-treatment PBWT1 levels were available in 41 patients and the median level was 790 copies/µg RNA (range, less than 50–310000). Baseline characteristics according to PBWT1 levels (≤ 790 [lower group] [n = 21] and 〉 790 [higher group] [n = 20] ) are summarized in Table 1. Median number of AZA treatment cycles was 4 (range, 1–18). Four patients (2 in higher group, and 2 in lower group) received allogeneic stem cell transplantation (alloSCT) after AZA treatment. OR rates were significantly lower in PBWT1 higher group than lower group (30.0 vs 71.4%, p = 0.03). In univariate analysis, other significant risk factors or with borderline significance for OR were higher serum ferritin levels ( 〉 1000 ng/ml) and RBC transfusion dependency ≥ 4 units/8 weeks. In multivariate analysis, higher PBWT1 levels independently predicted reduced likelihood of OR (odds ratio = 0.212, 95% CI 0.01-0.95, p = 0.02). OS was significantly inferior in PBWT1 higher group as shown in Figure 1. In univariate analysis, other significant factor was Revised International Prognostic Scoring System (IPSS-R) risk groups (high risk defined as IPSS-R high or higher, and low risk defined as IPSS-R intermediate or lower). In multivariate analysis, higher PBWT1 levels (hazard ratio [HR] = 9.75, 95% CI 1.22-77.58, p = 0.03) and IPSS-R high risk (HR=7.04, 95% CI 1.43-34.48, p = 0.02) were independent predictors for OS. Conclusion Our results suggest that PBWT1 can predict both response and survival of MDS patients treated with AZA. Although salvage therapy including alloSCT can affect the survival, poor survival might result from inferior response rates in PBWT1 high patients. In MDS with high PBWT1, restoration of epigenetically silenced tumor suppressor genes with AZA might not induce apoptosis. We propose that alternative therapeutic strategies should be sought in MDS patients with high PBWT1 levels. Disclosures: No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Publisher: American Society of Hematology
    Publication Date: 2013
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  • 2
    In: Blood, American Society of Hematology, Vol. 122, No. 21 ( 2013-11-15), p. 4551-4551
    Abstract: The prognosis of elderly patients with acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS) is poor, even in the patients who achieved good response after induction therapy. In order to undertake allogeneic stem cell transplantation (allo-SCT) with reduced toxicity and without total body irradiation (TBI), we conducted a combination regimen consisted of fludarabine, busulfan, and melphalan (Flu/Mel/Bu) for reduced intensity stem cell transplantation (RIST) conditioning. Patients and methods Among a total of 50 patients who underwent Flu/Mel/Bu conditioning between 2004 and 2012 in our institute, 32 patients with myeloid malignancies were retrospectively reviewed. Disease status was defined by WHO classification 2008. Therapy consisted of fludarabine 25 mg/m2 for five days (125mg/m2) and melphalan 50mg/m2 for two days (100mg/m2), both by intravenous infusion. Busulfan 2 mg/kg was administered orally for two days (4mg/kg) between 2004 and 2006, and intravenously at 1.6 mg/kg for two days (3.2mg/kg) between 2007 and 2012. Results Among the 32 eligible patients, 18 were female and 14 male. Seventeen patients were diagnosed with AML, 14 with MDS and one with chronic myelomonocytic leukemia (CMML). Median age was 59 years (32-66 years), and the median follow-up period was 1337 days (12-3043 days).Disease status of AML was complete remission (CR)1 (5), CR2 (10), CR3 (1) and CR4 (1), respectively, and all CR1 patients had poor risk factors. Disease status of MDS was RA (4), RARS (2), RCMD (1), RAEB-1 (5) and RAEB-2 (2), respectively. Three patients of RAEB conducted induction chemotherapy and two patients achieved CR. Donor sources consisted of 22 of unrelated bone marrow (URBM), 5 of related bone marrow (RBM), 3 related peripheral blood (RPB), and 2 of unrelated cord blood (URCB), respectively. Graft-versus-host disease (GVHD) prophylaxis consisted of tacrolimus plus short term methotrexate (TAC+sMTX) (25) and cyclosporin plus methotrexate (CsA+sMTX) (7). The cumulative incidence of grade II-IV acute GVHD was 31.7% and chronic GVHD was 85.9%. Five-year non-relapse mortality (NRM) was 15.9% and the 5-year relapse rate was 18.8%. One-year overall survival (OS) was 81.2% (95%CI 62.9-91.1), 5-year OS was 74.6% (95%CI 55.5-86.4), one-year progression free survival (PFS) was 78.1% (95%CI 59.5-88.9), 5-year PFS was 65.4% (95%CI 44.2-80.2). Fourteen patients were older than 60 years, and both 5-year OS and PFS of this group were 85.7% (95%CI 53.9-96.2). For AML, one-year OS was 82.4% (95%CI 54.7-93.9), 5-year OS 70.1% (95%CI 42.3-86.3), one-year PFS was 76.5% (95%CI 48.8-90.4) and 5-year PFS was 61.8% (95%CI 32.9-81.2). Five-year NRM was 11.8% and the 5-year relapse rate was 26.5%. For MDS, both one-year OS and 5-year OS were 78.6% (95%CI 47.2-92.5), and both one-year PFS and 5-year PFS were 69.8% (95%CI 37.8-87.6). Five-year NRM was 21.4% and the 5-year relapse rate was 8.7%.On the other hand, we conducted 32 allo-SCT for myeloid malignancies (22 of AML in CR and 10 of MDS) with conventional conditioning regimens of cyclophosphamide (Cy) and TBI or Bu and Cy between 2004 and 2012. Median age was 36.5 years (20-54 years), and the median follow-up period was 1191 days (38-3366 days).Disease status of AML was CR1 (14), and CR2 (8), respectively. Disease status of MDS was RA (3), RARS (2), RCMD (1), RAEB-1 (4), RAEB-2 (1), and MDS-U (1), respectively. Four patients of RAEB conducted induction chemotherapy and achieved CR. Donor sources were URBM (15), RBM (11), CB (5) and RPB (1), respectively. GVHD prophylaxis consisted of TAC+sMTX (17) and CsA+sMTX (15).The outcomes of Flu/Mel/Bu were not statistically inferior to those of conventional regimens with one-year OS of (81.2% vs. 87.1%, p=0.564) and 5-year OS of (74.6% vs. 78.0%, p=0.564), and one-year PFS of (78.1% vs. 83.9%, p=0.183) and 5-year PFS of (65.4% vs. 80.4%, p=0.183). Conclusions Flu/Mel/Bu was tolerable, and produced good outcomes and may be a candidate for curative conditioning regimen of RIST, especially for patients with myeloid malignancies in controlled status. Disclosures: No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Publisher: American Society of Hematology
    Publication Date: 2013
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  • 3
    In: Blood, American Society of Hematology, Vol. 124, No. 21 ( 2014-12-06), p. 1654-1654
    Abstract: Background: A consensus is yet to be reached on therole of interim fluorine-18 fluorodeoxyglucose (18F-FDG) position emission tomography-computed tomography (PET-CT) for prognosis in diffuse large B-cell lymphoma (DLBCL), but it has been recently reported that maximum standardized uptake value (SUV max) reduction between baseline and midtherapy may be a better predictor of overall survival (OS) and event free survival (EFS). Patients and Method: 456 patients were newly diagnosed with DLBCL in our institute between February 2007 and January 2013. Primary central nervous system lymphoma or primary effusion lymphoma patients were excluded from this study. All patients who were not perfomed PET were also excluded. In order to determine the predictive value of interim PET-CT and SUV max reduction criteria on OS and EFS for patients with DLBCL, we retrospectively analyzed the PET-CT and SUV max data of 104 first-line DLBCL patients treated with 6 to 8 courses of R-CHOP or R-THPCOP therapy. PET-CT was performed at diagnosis (baseline), and after 2 to 4 courses (interim PET). Interim PET was scheduled two weeks after the second to fourth cycle of immunochemotherapy. A visual analysis of interim PET was done with the use of the Deauville criteria, and an analysis of the SUV max reduction criteria was calculated by [(SUV max reduction from baseline to interim PET)/SUV max at baseline]. For each PET image, the tumor with the most intense 18F-FDG uptake was identified among all foci with the use of a graded color scale. The hottest volumetric region was determined, and the SUV max was calculated. To assess the SUV max, the hottest tumor in any region or organ on interim PET was used for comparison, even if its location differed from the initial hottest tumor in baseline PET. Survival curves were estimated with Kaplan–Meier analysis and compared using the log-rank test. Results: One hundred and four patients were enrolled in this study, and their characteristics are detailed in Table 1. With a median follow-up of 38.5 months, the two-year overall survival and EFS were 83.3% and 73.6%. The results forinterim PET were 66.3% (69/104) negative and 33.7% (35/104) positive, respectively. Interim PET negative versus positive two-year OS was 85.0% (95%CI 0.739-0.917) versus 79.9% (95%CI 0.624-0.899) (P value 0.58), and the two-year EFS was 77.7% (95%CI 0.658-0.860) versus 65.7% (0.476-0.789) (P value 0.24), respectively (Figure 1). The SUV max cutoff values 66% estimated by receiver-operating-characteristic (ROC) analysis to distinguish treatment response were similar to other independent cohort studies at 2 to 4 cycles of induction treatment, and this threshold was chosen to analyze our series. For SUV max data, the two-year OS was 84.4% (95%CI 0.750-0.904) for a SUV max reduction ≥66.0% compared with 75.0% (95%CI 0.408-0.912) for a reduction of 〈 66.0% (P value 0.02). The two-year EFS for the former was 75.7% (95%CI 0.654-0.833) compared with 58.3% (95%CI 0.270-0.801) for the latter (P value 0.03) (Figure 2). Baseline characteristics according to SUV max reduction criteria [SUV max≥66%(n=92),and SUV max 〈 66%(n=12)] are summarized in Table 1. Conclusions: Analysis of the data in our study was unable to demonstrate the predictive value of interim PET for OS and EFS, but SUV max reduction criteria may improve the prognostic value of interim PET. However, the 66% cutoff value must be validated in a larger-scale prospective trial, and further investigation and the standardization of SUV max reduction criteria are needed. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Publisher: American Society of Hematology
    Publication Date: 2014
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  • 4
    In: Blood, American Society of Hematology, Vol. 124, No. 21 ( 2014-12-06), p. 2539-2539
    Abstract: Introduction: Allogeneic hematopoietic cell transplantation (HCT) has curative potential for a variety of hematologic malignancies. However, the success of HCT is heavily dependent on the disease and remission status. Armand et al. recently proposed a disease risk index (DRI) to assess the risk for allogeneic HCT based on the disease and remission status, and could be used to risk stratification on survival (Armand et al Blood 2012), but evaluation of its clinical significance is limited. HCT-comorbidity index (HCT-CI) was developed as a measure of pretransplant organ dysfunctions (Sorror et al Blood 2005), which has been shown to be associated with nonrelapse mortality (NRM). In order to clarify any association between pretransplant factors including DRI and HCT-CI, and respective overall survival (OS), NRM and relapse rate, we retrospectively reviewed the data of patients who underwent allogeneic HCT at our department. Patients and Methods: A total of 305 patients with hematologic malignancies who underwent initial allogeneic HCT at our department between January 2000 and April 2013 were included. After excluding patients with insufficient HCT data, we included a total of 244 patients (138 male, 106 female) with a median age of 49.5 (range 15-69) years. A total of 133 patients received myeloablative conditioning and 111 received reduced-intensity regimens. Stem cell sources were bone marrow (n=177), peripheral blood (n=32), combined peripheral blood and bone marrow (n=1), and cord blood (n=34). A total of 146 patients received tacrolimus-based regimens and 98 patients received cyclosporine-based regimens for GVHD prophylaxis. The DRI has four risk-based categories (low, intermediate, high, and very high) and the HCT-CI has three categories in order of ascending risk (0, 1-2 and ≥3). OS was calculated with the Kaplan-Meier method, compared among groups with the log-rank test, and multivariable Cox regression analyses were used to evaluate factors associated with OS. The cumulative incidence of NRM and relapse were calculated while treating relapse and death without relapse, respectively, as competing events, and competing risk regression analyses were used to evaluate risk factors associated with NRM and relapse. Results: The median follow-up for survivors was 4.7 years (range 0.1-14.2 years). Pretransplant disease risks in the DRI low, intermediate, high, and very high risk groups were 8%, 60%, 25%, and 7%, and 4-year OS in the same groups were 74%, 64%, 35%, and 12%, respectively (Figure 1, p 〈 0.001). Four-year OS among patients with a HCT-CI of 0, 1-2, and ≥3 were 63%, 52%, and 41%, respectively. Multivariable analysis showed a significant association with OS for DRI (high risk hazard ratio [HR] 2.62, p 〈 0.001; very high risk HR 5.26, p 〈 0.001 versus intermediate risk), HCT-CI (HCT-CI ≥3 HR 1.64, p=0.022 versus HCT-CI 0-2), 2-4 performance status (HR 3.10, p 〈 0.001), and donor-recipient ABO minor-mismatch (HR 2.00, p=0.005 versus ABO match). The cumulative incidence of 4-year NRM was 25%, and NRM was significantly associated with HCT-CI (HCT-CI ≥3 HR 2.27, p=0.003 versus HCT-CI 0-2) and 2-4 performance status (HR 3.89, p 〈 0.001). The cumulative incidence of 4-year relapse was 25%, and relapse was significantly associated with DRI (high risk HR 2.26, p=0.012; very high risk HR 8.11, p 〈 0.001 versus intermediate risk). Finally, we reclassified all patients into four risk groups incorporating DRI and HCT-CI: DRI low-intermediate plus HCT-CI 0-2 (group I), DRI low-intermediate plus HCT-CI ≥3, or DRI high plus HCT-CI 0-2 (group II), DRI high plus HCT-CI ≥3 (group III), and DRI very high (group IV). Four-year OS among patients with group I, II, III, and IV were 68%, 47%, 25%, and 12%, respectively (Figure 2, p 〈 0.001). Conclusions: Our results suggest that risk stratification with DRI and HCT-CI for the prognosis of relapse and NRM may be useful for patients undergoing allogeneic HCT. Larger and prospective studies are warranted to more precisely validate these findings. Figure 1 Figure 1. Figure 2 Figure 2. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
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    Publisher: American Society of Hematology
    Publication Date: 2014
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  • 5
    In: Blood, American Society of Hematology, Vol. 124, No. 21 ( 2014-12-06), p. 1636-1636
    Abstract: Background: Previous studies have shown that the absolute lymphocyte count (ALC) in peripheral blood at diagnosis may be an independent prognostic factor of IPI for patients with diffuse large B-cell lymphoma (DLBCL). In the rituximab era, the U.S. National Comprehensive Cancer Network International Prognostic Index (NCCN-IPI) was developed to improve the risk stratification of DLBCL in comparison to the existing IPI. Therefore, the aim of this study was to clarify the impact of ALC at diagnosis on event free survival (EFS) and overall survival (OS) on analysis performed with factors included in NCCN-IPI. Patients and methods: We retrospectively reviewed the ALC of 413 patients with newly diagnosed DLBCL treated with R-CHOP at our hospital between January 2005 and March 2013. Primary central nervous system lymphoma patients were excluded from this study. ALC was determined in all patients from complete blood count with differential white blood count at the time of diagnosis, and prior to therapy administration. EFS and OS were estimated according to the Kaplan-Meier method. Multivariate analysis was performed with the proportional hazard Cox model. Results: The median ALC was 1.2x10E9/L (range, 0.06-9.0). We set an ALC cut-point at 1.0x10E9/L based on previous studies. The median follow-up duration was 40 months. Baseline characteristics according to ALC ( 〈 1.0x10E9/L[n=145] and 〉 1.0x10E9/L[n=268]) are summarized in Table1. Patients with ALC 〈 1.0x10E9/L had a significantly poorer EFS and OS than patients with ALC 〉 1.0x10E9/L (5-year EFS, 37.0% versus 68.9%, p 〈 0.001; 5-year OS, 46.3% versus 80.0%, p 〈 0.001). On multivariate analysis performed with factors included in IPI and NCCN-IPI, ALC remained an independent predictor of EFS (IPI: hazard ratio [HR] 1.95; 95% confidence interval [CI] 1.43-2.68; p 〈 0.001, NCCN-IPI: HR 1.94; 95%CI 1.42-2.65; p 〈 0.001) and OS (IPI: HR 2.35; 95%CI 1.61-3.42; p 〈 0.001, NCCN-IPI: HR 2.29; 95%CI 1.57-3.33; p 〈 0.001) (Table2). Importantly, within the poor R-IPI group, ALC distinguished patients with different 5-year EFS (24.4% versus 50.4%, p 〈 0.001) and OS (35.7% versus 65.7%, p 〈 0.001). For the high NCCN-IPI group also, ALC distinguished patients with different 5-year EFS (14.8% versus 39.8%, p 〈 0.01) and OS (17.5% versus 54.5%, p 〈 0.001) (Figure1). Conclusions: According to our results, ALC 〈 1.0x10E9/L is an adverse prognostic factor and independent of IPI and NCCN-IPI. ALC might be more successful in identifying high-risk patients in which IPI and NCCN-IPI analysis was unrevealing. Our results suggest that other therapeutic strategies may be more effective in high-risk patients with DLBCL. Disclosures No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Publisher: American Society of Hematology
    Publication Date: 2014
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  • 6
    In: Blood, American Society of Hematology, Vol. 116, No. 21 ( 2010-11-19), p. 2304-2304
    Abstract: Abstract 2304 Background: Hemorrhagic cystitis (HC) is a major complication after allogeneic hematopoietic stem cell transplantation (SCT), and is life-threatening in severe cases. Early onset HC is usually due to toxic effects of conditioning regimen, while late onset HC is caused by viral or bacterial infection, and GVHD. In order to find out risk factors and to analyze prognosis of late onset HC, we retrospectively reviewed data in our institution from 1996 through 2009. Patients and Methods: We analyzed 232 cases having underwent allogeneic SCT (AML; 65, MDS; 42, ALL; 44, ML; 44, others; 37), excluding 22 patients who received second allogeneic transplantation. The median age at SCT was 47 years old (range, 13–72), and median follow-up of survivors was 1529 days (range, 99–4822) after SCT. One-hundred and eleven patients received from related donor, and 121 from unrelated. Conditioning was myeloablative in 139 cases, and reduced intensity (RIC) in 93. Myeloablative regimen was cyclophosphamide (CY) plus total body irradiation (TBI) in 79 cases, and busulfan (BU) plus CY in 54, while RIC regimen was fludarabine (Flu) plus melphalan (Mel) plus BU in 52, and Flu plus Mel plus TBI in 12. Disease status at SCT was controlled (partial or complete remission) in 164 cases, and progressive in 68. Prophylaxis for acute GVHD consisted of calcineurin inhibitor (cyclosporine; 106 cases, tacrolimus; 126 cases), short term methotrexate (158 cases), and corticosteroid (73 cases). Acute GVHD occurred in 121 cases, 92 of them were grade II-IV, and 74 were treated with corticosteroid. Cytomegalovirus (CMV) antigenemia was positive in 82 cases after engraftment. Definition of late onset HC was microscopic or macroscopic hematuria with urinary symptoms occurring after 7 days from SCT. Urine culture and virological analysis (polymerase chain reaction of CMV, adenovirus [ADV], and BK virus DNA) were carried out in all cases with HC. Treatment was mainly hydration and intravenous hemostat, including intravenous immunoglobulin (IVIg) and antiviral agents. Results: HC was reported in 43 cases on a median of 36 days (range, 7–469) after SCT, and 31 of them (72.1%) recovered after a median of 21 days (range, 2–252). Cumulative incidence of HC after 1 year from SCT was 21.6% (95% confidence interval [CI] 18.3–24.7%). In univariate analysis, risk factors identified included age over 45 years old (p=0.04), progressive disease status at SCT (p=0.04), myeloablative conditioning (p=0.04), acute GVHD grade II-IV treated with corticosteroid (p=0.01), and positivity of CMV antigenemia after SCT (p 〈 0.01). On multivariate analysis, older age (p 〈 0.01, hazard ratio [HR] 2.63), myeloablative conditioning (p 〈 0.01, HR 4.32), and positivity of CMV antigenemia (p 〈 0.01, HR 3.61) remained independent predictors. Each HC case was graded according to National Cancer Institute Common Terminology Criteria for Adverse Events (grade 1; 12, grade 2; 19, grade 3; 12, grade 4 to 5; none), and we defined moderate as grade 1 or 2, and severe as grade 3 or more. In subgroup analysis of patients with HC, myeloablative conditioning (p=0.11, Odd's ratio [OR] 5.24) and positivity of ADV in urine (p=0.10, OR 3.17) tended to be associated with severe HC. Recovery rate from HC was inferior and median duration from onset to recovery was longer in severe HC (25.0%, 121 days) with significance (p 〈 0.01) compared with moderate HC (90.3%, 17.7 days). Using of IVIg or antiviral agent (vidarabine, gancyclovir, or foscarnet) did not improve recovery rate in both groups. Overall survival (OS) at 1 year after day 35 landmark had no significance between cases with and without HC (59.7%, 56.7%, p=0.70), significantly poor in severe cases (16.7%, p 〈 0.01) compared to cases with moderate HC and cases without HC. Treatment-related mortality (TRM) at 1 year after day 35 landmark was 25.0% with HC and 29.0% without HC (p=0.35), while 59.1% in patients with severe HC (p=0.01). Conclusion: Five significant risk factors of HC such as older age, progressive disease status, myeloablative conditioning, acute GVHD, and CMV antigenemia were found out in this study. OS and TRM of patients with all the grade of HC were equivalent to those without HC, while severe HC significantly elevated TRM and shortened OS. Severity of HC was related to myeloablative regimen and ADV in urine. Any successful attempts to prevent severe HC have not been established yet, and novel prophylactic and pre-emptive strategies are warranted. Disclosures: No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
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    Publisher: American Society of Hematology
    Publication Date: 2010
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  • 7
    In: Blood, American Society of Hematology, Vol. 124, No. 21 ( 2014-12-06), p. 29-29
    Abstract: PU.1 is an Ets family transcription factor, which is essential for differentiation of both myeloid and lymphoid lineages. It was previously reported that conditional knockout of the upstream enhancer region (URE) located in 14 kb 5’ of the murine PU.1 gene resulted in down-regulation of PU.1 expression in granulocytes and B lymphocytes by 80% compared to that of wild type and induced acute myeloid leukemia and CLL-like diseases in mice. Therefore, down-regulation of PU.1 in myeloid and B cell lineages results in hematological malignancies. We previously reported that PU.1 is down-regulated in 5 out of 7 myeloma cell lines as well as primary myeloma cells from a subset of myeloma patients; that the promoter and the URE located in 17 kb 5’ of the human PU.1 gene that is homologous to that in 14 kb 5’ of murine PU.1 gene are highly methylated in these cell lines; and that conditionally expressed PU.1 with tet-off system induces cell growth arrest and apoptosis in myeloma cell lines, U266 and KMS12PE, suggesting that the down-regulation of PU.1 is necessary for myeloma cell growth. Here, to evaluate tumor suppressor activity of PU.1 in mature B and plasma cells in vivo, we generated Cγ1-Cre PU.1 knockout mice by crossing Cγ1-Cre and PU.1-loxP mice. We confirmed that PU.1 alleles were both conditionally deleted in the maturation stages of B cells from post germinal center B to plasma cells. By 18-24 months of age, about 77.7% (10 of 13) of the knockout mice had developed serum M proteins. To induce B cell differentiation to plasma cells, those mice were immunized with NP-CGG and 76.9% (20 of 26) of the mice developed serum M protein. ELISA of sera from those mice revealed that IgG was not elevated compared to those from the PU.1-loxP mice, which was thought because Cγ1-Cre locus fails to produce IgG1. Instead, a small number (5 of 20) of the mice showed relatively large amounts of IgM and/or IgA. When 11 such mice were sacrificed, 7 had developed splenomegaly and/or intestinal B cell lymphoma. Immunostaining revealed that B220+ cells had infiltrated into the tumors and various organs including the spleen, liver, and bone marrow. Those cells were monoclonal for κ chain and partly CD138 positive. When we transplanted those tumor cells into Rag2-/- Jak3-/- immunedeficient mice, all the mice died within 3 weeks. Thus, PU.1 apparently functions as a tumor suppressor in mature B cells and its deletion in late B cell maturation stages produces B cell lymphoma with M proteinemia. The remaining 4 mice developed high titer IgM and/or IgA levels and flow cytometry of bone marrow cells and splenocytes revealed that those cells were monoclonal for κ chain and positive for B220 and IgM and/or IgA, suggesting that those mice suffered from multiple myeloma or monoclonal gammopathy with undetermined sighnificance (MGUS). These data strongly suggest that conditional knockout of PU.1 in post germinal center B and plasma cells results in B cell lymphoma and plasma cell neoplasms related to multiple myeloma. Disclosures No relevant conflicts of interest to declare.
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    Publisher: American Society of Hematology
    Publication Date: 2014
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  • 8
    In: Blood, American Society of Hematology, Vol. 118, No. 21 ( 2011-11-18), p. 3608-3608
    Abstract: Abstract 3608 CD56 expression is reportedly associated with an adverse prognosis in patients with acute promyelocytic leukemia (APL) (Murray et al, 1999, Ferrara et al, 2000), but the prognostic significance of CD56 has not been elucidated in multivariate analysis. Recently, however, Spanish group demonstrated that CD56 expression was a significant adverse prognostic factor for relapse in APL treated with ATRA combined anthracycline-based regimens in multivariate analysis (Montesinos et al, 2011). We tried to evaluate the significance of CD56 in APL on more intensive regimen composed by ATRA, anthracycline and cytosine arabinocide (Ara-C). Newly diagnosed APL patients were registered to the JALSG APL97 from May 1997 to June 2002. The detail of the treatment protocol was previously reported (Asou et al, 2007). In brief, induction therapy was composed of ATRA and chemotherapy including idarubicin and Ara-C. The dose and duration of induction therapy were based on initial white blood cell (WBC) count. After 3 courses of consolidation therapies, patients were randomly allocated either to receive 6 courses of intensified maintenance chemotherapy or to observation. Leukemia blasts were stained by anti-CD45 monoclonal antibody (mAb), gated by CD45 expression and analyzed by flow cytometer. Cells were additionally stained by fluorescein conjugated mAb against each surface antigen including CD56. Surface markers were defined as positive if more than 20% of APL cells expressed a specific antigen. Clinical characteristics were compared by the chi-square test or the Fisher's exact test for categorical data and the Wilcoxon rank-sum test for continuous data. Overall survival (OS) and event-free survival (EFS) were estimated by the Kaplan-Meier method and the log-rank test. Cumulative incidence of relapse (CIR) was compared by the Gray test for comparisons. Multivariate analyses were also performed. Clinical outcomes were renewed on March 2010 and the median follow up period is 7.5 years. Among 302 patients enrolled, 239 patients were assessable. Twenty three patients (9.6%) were CD56-positive (CD56+) APL. CD56 expression was significantly associated with lower platelet count, serious DIC and expressions of CD2, 7, 34, HLA-DR. (P = 0.04, 0.04, 0.03, 0.04, 〈 0.001 and 〈 0.001, respectively) The CR rate and mortality during induction therapy were not significantly different compared with CD56− APL. CIR and EFS showed inferior trend in CD56+ APL (24.3% vs. 39.1%, P = 0.08 and 64.8% vs. 47.8%, P = 0.08, respectively), whereas cumulative incidence of extramedullary relapse and OS were not different between two groups (P = 0.27 and P = 0.52, respectively). In patients whose initial WBC count was more than 3.0 × 109/L, EFS and CIR for CD56+ group were significantly inferior (28.9% vs. 53.8%, P = 0.03 and 63.6% vs. 30.8%, P = 0.008, respectively), while in patients whose initial WBC count was under 3.0 × 109/L, EFS and CIR were not significantly different (P = 0.99 and P = 0.44, respectively). In multivariate analysis, CD56 expression was an independent prognostic factor for EFS in patients whose initial WBC count was more than 3.0 × 109/L (HR = 3.10; 95% CI, 1.4–7.1, P = 0.007). CD56+ APL was associated with lower platelet count, serious DIC and expressions of CD2, 7, 34 or HLA-DR. Thus, CD56 expression was an independent adverse prognostic factor for EFS in APL treated by ATRA combined with anthracycline and Ara-C regimen, especially in APL whose initial WBC count is more than 3.0 × 109/L. The prognostic significance of CD56 expression should be continuously monitored in future. Disclosures: Takeshita: Takeda: Research Funding; Novaltis: Research Funding. Naoe:Kyowa-Hakko Kirin.: Research Funding; Dainipponn-Sumitomo Pharma.: Research Funding; Chugai Pharma.: Research Funding; Novartis Pharma.: Honoraria, Speakers Bureau; Zenyaku-Kogyo: Research Funding; Otsuka Pharma.: Research Funding.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
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    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2011
    detail.hit.zdb_id: 1468538-3
    detail.hit.zdb_id: 80069-7
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  • 9
    In: Blood, American Society of Hematology, Vol. 92, No. 4 ( 1998-08-15), p. 1374-1383
    Abstract: A novel cell line was established from a patient with a leukemic-state nasal angiocentric natural killer (NK) cell lymphoma with systemic skin infiltration. The morphology of the leukemic cells was large-granular-lymphocyte (LGL), and their immunophenotype was CD2+, CD3−, CD5+, CD7+, CD16−, CD56+, and CD57−. The presence of Epstein-Barr viral (EBV) genome was shown in specimens from the patient’s nose, skin, and peripheral blood by in situ hybridization using an EBV-encoded small RNA-1 probe or by Southern blotting using a terminal-repeat probe of the EBV genome. Leukemic cells were cocultured with a mouse stromal cell line (SPY3-2) in the presence of 100 U/mL recombinant human interleukin-2 and a novel stromal cell-independent cell line, NK-YS, was established. The NK-YS cells showed LGL morphology and expressed surface CD2, CD5, CD7, CD25, CD56, and CD95. The NK-YS cells retained cytotoxicity against K562 and Jurkat cells. A Southern blotting using a terminal-repeat probe of EBV showed that NK-YS and fresh leukemic cells had a clonal EBV genome, whereas the T-cell receptor β and γ chain genes of NK-YS were not rearranged. In an immunocytochemical analysis, the NK-YS cells showed a type-II latent infection of EBV. The NK-YS cells preserved the original characteristics of NK cell lymphoma/leukemia and will be a useful tool for the study of biological characteristics of EBV-associated nasal angiocentric NK cell lymphoma/leukemia. © 1998 by The American Society of Hematology.
    Type of Medium: Online Resource
    ISSN: 1528-0020 , 0006-4971
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    RVK:
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 1998
    detail.hit.zdb_id: 1468538-3
    detail.hit.zdb_id: 80069-7
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  • 10
    In: Blood, American Society of Hematology, Vol. 114, No. 22 ( 2009-11-20), p. 1477-1477
    Abstract: Abstract 1477 Poster Board I-500 Human embryonic stem (ES) cells differentiate into three lineages in vitro and in vivo as mouse ES cells. They are therefore highly promising source of various cells/tissues in the regenerative medicine. The current protocols, however, remain to be optimized for the induction of the cells/tissues required. We have recently reported that the lentiviral transduction of TAL1/SCL gene to ES cells derived from the common marmoset, a small nonhuman primate, enables efficient differentiation into hematopoietic progenitor cells even in the absence of stromal cells (Kurita et al. Stem Cells.24:2014-22, 2006). Such culture condition without any stromal cells is considered to facilitate clinical application of ES cell-derived cell/tissues therapy in the regenerative medicine. The present study addressed whether the strategy is also effective in human ES cells. First, we determined optimal culture conditions to induce multilineage hematopoietic differentiation in a human ES cell line, khES-1, kindly provided by Dr. Nakatsuji, Kyoto University, Japan, as assessed by the expression of Brachyury, Flk1 and CD34. We found that the addition of BMP4 and VEGF augmented hematopoietic differentiation of embryoid bodies, and determined optimal concentrations of the cytokines. We established four human ES cell lines stably expressing TAL1/SCL gene by lentiviral transduction. The TAL1/SCL transduction further increased the hematopoietic differentiation under the optimal culture condition as assessed by the expression of CD34, CD235a and CD133. We also observed increased number of hematopoietic progenitor cells derived from two of the TAL1/SCL expressing human ES cell lines by colony-forming assays. Hematopoietic differentiation of the TAL1/SCL expressing ES cells in vivo is also being investigated by transplantation into irradiated immune deficient mice. These results suggest that the combination of optimal culture conditions and lentiviral TAL1/SCL gene transduction is a highly effective strategy to obtain hematopoietic stem cells from human ES cells in the absence of any stromal cells. Disclosures: No relevant conflicts of interest to declare.
    Type of Medium: Online Resource
    ISSN: 0006-4971 , 1528-0020
    RVK:
    RVK:
    Language: English
    Publisher: American Society of Hematology
    Publication Date: 2009
    detail.hit.zdb_id: 1468538-3
    detail.hit.zdb_id: 80069-7
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