bluuebunny/crossref_metadata_2025 huggingface dataset:
Downregulation of THAP11 by Bcr-Abl Promotes c-Myc-Mediated CML Cell Proliferation.
Authors: Yokota Daisuke , Nakamura Satoki , Takemura Tomonari , Hirano Isao , Fujisawa Shinya , Takeshita Kaori , Ono Takaaki , Shigeno Kazuyuki , Ohnishi Kazunori
Published: October 2019

Abstract 3255

Poster Board III-1

[Background and Purpose] THAP11 (thanatos-associated protein 11) is one of the zinc-dependent, sequence-specific DNA-binding factors, which regulate cell proliferation, apoptosis and cell cycle. It has been reported that THAP11 is ubiquitously expressed in normal tissues and frequently downregulated in several tumors. In CML cells, we found that THAP11 expression was inhibited. We investigated the mechanisms of the suppression of THAP11 and its function in CML cell proliferation. [Methods] The cells used in this study were human CML cell lines, K562 and Meg01 cells. Primary CML cells were obtained from the bone marrow of CML (CP) patients. Human normal mononuclear cells (MNCs) were isolated from bone marrow of healthy volunteers after obtaining informed consents. For analysis of THAP11 mRNA expression, quantitative RT-PCR was performed in all cell lines treated with Abl kinase inhibitors (STI571, AMN107, and BMS354825). For proliferation analysis and the expression of c-Myc in CML cells, MTT assays, western blot and cell cycle analysis were performed in all cell lines transfected with Bcr-Abl siRNA, THAP11 siRNA, or THAP11 cDNA respectively. Moreover, THAP11 expression, c-Myc expression, and the colony counts of CFU-GEMM, CFU-GM, and BFU-E were analyzed in CML stem/progenitor cells transfected with Bcr-Abl siRNA or treated with Abl kinase inhibitors. [Results] In CML cell lines treated with Abl kinase inhibitors or transfected with Bcr-Abl siRNA, the expressions of THAP11 and c-Myc mRNA and protein were significantly increased compared to untreated cells. On the other hand, in CML cells transfected with the THAP11 cDNA, the c-Myc expression was suppressed. The overexpression of THAP11 induced G1 cell cycle arrest through p27 and p21 accumulation, and inhibited the CML cell proliferation. Moreover, in CML stem/progenitor cells obtained from patients with CML, the transfection with Bcr-Abl siRNA or treatment with Abl kinase inhibitors increased the expression of THAP11 mRNA and protein, and decreased the c-Myc expression and the counts of CFU-GEMM, CFU-GM and BFU-E. [Conclusion] Our results demonstrated that the Bcr-Abl suppressed the expression of THAP11, the depletion of THAP11 induced c-Myc expression, and induced the proliferation of CML cells through cell cycle progression. Moreover, induction of THAP11 expression inhibited the proliferation of CML stem/progenitor cells.

Disclosures:

No relevant conflicts of interest to declare.
FluBup-ATG-TBI for High-Risk or Advanced Adult ALL in Remission: A Retrospective Review of a Mature Cohort.
Authors: Daly Andrew , Stewart Douglas A. , Chaudhry Ahsan , Bahlis Nizar J , Brown Christopher , Duggan Peter , Geddes Michelle , Savoie Mary Lynn , Storek Jan , Yang Maggie , Russell James A.
Published: October 2019

Abstract 3384

Poster Board III-272

Purpose:

Hematopoietic stem cell transplantation (HSCT) is routinely offered to suitable candidates with high-risk or advanced acute lymphoblastic leukemia. In this report we describe our experience with a novel conditioning regimen, previously reported to confer low TRM and high OS in AML, in this population.

Patients and Methods:

Between 05/2000 and 06/2008 44 patients with high-risk (either adverse cytogenetics, age > 35 or high WBC at diagnosis) or advanced (>CR1) ALL received HSCT after myeloablative conditioning using Fludarabine 50 mg/m2 days -6 to -2, Busulfan 3.2 mg/kg days -5 to -2, and TBI 2 Gy x 2 doses. GVHD prophylaxis was with rabbit ATG 0.5 mg/kg day -2, then 2 mg/kg days -1 and 0, CyA (tapered on day +56) and short-course methotrexate. All patients were in remission at the time of transplant. Imatinib mesylate (Gleevec) was not used routinely before or after transplant for patients with BCR-Abl+ ALL. Median (range) follow-up of surviving patients is 4.3 (1.0 – 9.0) years. All patients were followed for at least 1 year after BCT.

Results:

The cohort consists of 32 patients with high-risk (median age 40 (19-64) years) and 12 patients with advanced (median age 25 (19-65) years) disease who received bone marrow (n=5), G-CSF mobilized blood stem cells (n=38) or umbilical cord blood stem cells (n=1) from 25 related (21 fully-matched) or 19 unrelated (16 fully-matched) donors. Median times to neutrophil and platelet engraftment were 14 (11-28) days and 18 (9-105) days, respectively. Five patients did not require platelet transfusion. Cumulative incidences of grade II-IV and grade III-IV acute GVHD were 53.2% (95% CI 36.3%-67.5%) and 20.6% (95% CI 3.5%-47.6%), respectively. Chronic GVHD complicated 55% (95% CI 38.4%-68.8%) of transplants. Six patients (13.6%) died in remission before day +100. Event-free and overall survivals at 5 years were 56.7% (95% CI 39.1%-71.0%) and 66.0% (95% CI 48.8%-78.6%), respectively. Nine patients (20%) died in remission, 6 (14%) died after relapse and two patients remain alive following second transplants for relapsed disease. Five of 11 patients age > 50, 8/12 patients with advanced disease and 13/23 patients with adverse-risk cytogenetics remain alive.

Conclusion:

We found encouraging results with FluBup-ATG-TBI in a cohort of patients with advanced or high-risk ALL. These results warrant comparison with other conditioning regimens in a randomized, multi-center study.

Disclosures:

Daly: Hoffmann-Laroche: Advisory Board, Honoraria. Stewart:Hoffmann La Roche: Advisory Board, Honoraria, Research Funding.
Induction Chemotherapy Followed Immediately by Busulfan-Based Reduced Conditioning and Allografting in Elderly Patients with Advanced MDS or sAML.
Authors: Kröger Nicolaus , Zabelina Tatjana , Wolschke Christine , Lellek Heinrich , Stübig Thomas , Lestin Maite , Alchalby Haefaa , Ayuk Francis , Bacher Ulrike , Fehse Boris , Zander Axel R.
Published: October 2019

Abstract 3387

Poster Board III-275

<>Introduction:

Allogeneic stem cell transplantation after a dose-reduced conditioning has become a reasonable treatment option for elderly patients with MDS/sAML. For patients with high number of blasts prior transplantation, the risk of relapse is considerably. To reduce the risk of relapse after dose-reduced allograft we performed a study using an anthracycline based induction chemotherapy (amsacrine, cytosine-arabinoside, fludarabine) followed immediately by a reduced intensity conditioning therapy consisting of busulfan (8mg/kg).

Patients and Methods:

Between November 2005 and November 2008, 49 patients with MDS (n = 24), CMML (n = 8) and sAML (n = 17) and a median age of 61 years (r: 26 – 73) and a median number of 13% blasts were included. Stem cell source were unrelated (n = 43) or related donor (n = 6).

Results:

No graft failure was observed and the median time to leukocyte engraftment (> 1.0 × 109 /l) was only 10 days (r: 7 – 32). The incidence of acute graft-versus-host disease grade II to IV was 39 % and of grade III / IV was 14 %. Chronic GvHD was noted in 57 % of the patients, which was limited in 35 % and extensive in 32 % of the patients. After a median follow-up of 15 months (r: 3 – 35) the two-years estimated disease-free and overall survival was 49 % (95 % CI 33 – 65 %) and 54 % (95 % CI 39 – 69 %), respectively. The 1 year cumulative incidence of treatment-related mortality was 29 % (95 % CI 15 – 43 %). The 2 year cumulative incidence of relapse was 18 % (95 % CI 6 – 30 %). Patients with fully matched related or unrelated donor had a better survival than patients transplanted from mismatched donor (69 % vs. 37 %; p=0.06).

Conclusions:

A sequential approach using anthracycline based induction chemotherapy followed immediately by a busulfan based reduced conditioning regimen and allogeneic stem cell transplantation from related and unrelated donors resulted in a fast engraftment and a relative low risk of relapse in elderly patients with advanced MDS or sAML. To lower the therapy related mortality a careful donor selection is mandatory.

Disclosures:

No relevant conflicts of interest to declare.
Interim Restaging FDG-PET/CT to Guide Use of High Dose Sequential Induction Therapy with Rituximab-Dose-Intensive Cyclophosphamide, Etoposide, Cisplatin (RDICEP) and Rituximab-Carmustine, Etoposide, Cytarabine, Melphalan (RBEAM) and Autologous Stem Cell Transplantation (ASCT) for Poor Prognosis Diffuse Large B-Cell Lymphoma (DLBCL). ClinicalTrials. Gov Identifier: NCT00530179.
Authors: Stewart Douglas A. , Duggan Peter , Bahlis Nizar J , Daly Andrew , Geddes Michelle , Kloiber Reinhard , Fibich Christian , Macdonald David Blair , Bence-Bruckler Isabelle
Published: October 2019

Abstract 3414

Poster Board III-302

Purpose

Approximately 50-55% of patients (pts) with stage 3-4 DLBCL and elevated serum LDH achieve long-term event-free survival (EFS) following R-CHOP chemotherapy. A prospective, multicentre, phase II clinical trial was designed to evaluate the use of high dose sequential therapy with RDICEP then RBEAM/ASCT for such pts who have positive interim restaging FDG-PET/CT scans after 2 cycles of RCHOP.

Patients and Methods

Pts were eligible if they were HIV-, 18-65 years of age, and had stage 3-4 DLBCL with elevated serum LDH. An FDG-PET/CT scan was performed 10-15 days following the second cycle of R-CHOP. Pts with negative interim restaging PET scans completed 4 further cycles of RCHOP, whereas pts with positive PET scans (more than 1 site with uptake more intense than background liver) received one cycle of RDICEP (Rituximab 375mg/m2 day 1 and 8, C 1.75g/m2 d2-4, E 350mg/m2 d2-4 P 35mg/m2 d2-4, G-CSF d15-21 and autologous blood stem cell collection by apheresis d21or 22) followed by one cycle of RBEAM/ASCT (Rituximab 375mg/m2 day -6 and +14, BCNU 300mg/m2 d-6, E 200mg/m2 d-5to-2, A 400mg/m2 d-5to-2, M 140mg/m2 d-1).

Results

Of the 36 pts who have been accrued from 5/2007-7/2009, 33 pts have undergone interim PET/CT restaging following 2 cycles RCHOP. All 33 pts had IPI scores 3-5, the median age was 55 years (range19-65) with 7 pts (21%) older than 60 years, 20 (61%) had ECOG status 2-4, 28 (85%) had stage 4, 22 (67%) >1 extranodal site. At a median follow-up of 12mo, the 1 year OS and EFS rates for all 33 pts are 82% (95%CI = 66-98%) and 76% (95%CI = 58-83%), respectively. Interim restaging PET/CT was positive for 16 pts and negative for 17 pts. The EFS rates for PET+ and PET- pts are 78% (95%CI = 57-100%) and 66% (95%CI = 36-99%), respectively (logrank p=0.97). To date, 4 pts have died, 3 from lymphoma at 6, 10, 11 months post-RCHOP1, and 1 from sepsis and necrotic bowel on day 5 post-ASCT.

Conclusions

Preliminary results of this phase II study are encouraging, and suggest that high dose induction therapy with RDICEP and RBEAM/ASCT might improve EFS for poor prognosis DLBCL with positive interim restaging PET/CT. Accrual will continue to target ≥ 35 PET+ and ≥35 PET- pts.

Disclosures

Stewart: Hoffmann La Roche: Advisory Board, Honoraria, Research Funding. Off Label Use: Rituximab with High Dose Chemotherapy and Autologous Stem Cell Transplantation for DLBCL. Bence-Bruckler:Hoffmann La Roche: Honoraria, Research Funding.
SUPER- BEAM (SB): Incorporation of Bortezomib, Thalidomide, Dexamethasone, Cisplatin and Rapamycin Into the BEAM Regimen for Multiple Myeloma (MM).
Authors: Waheed Sarah , Alsayed Yazan , Nair Bijay , Szymonifka Jackie , Shaughnessy John D , van Rhee Frits , Anaissie Elias , Barlogie Bart
Published: October 2019

Abstract 3421

Poster Board III-309

Introduction

While the prognosis of patients with MM has been drastically improved with the introduction of novel agents into the treatment armamentarium, relapse management remains a difficult task, especially in the post-transplant and high-risk MM setting, as defined by the presence of cytogenetic abnormalities (CA) and gene expression profiling (GEP).

Patients and Methods

We have explored the efficacy and safety of SB in 95 patients with MM. The regimen comprised BCNU at 300mg/m2 on day 1, etoposide 200mg/m2 on days 1-4; cytarabine 400mg/m2 on days 1-5; melphalan 140mg/m2 on day 5 plus: bortezomib 1.0-1.3mg/m2 on days 1+4, thalidomide 100-200mg on days 1-5, dexamethasone 20-40mg days 1-5, cisplatin 10-12.5mg/m2/d by continuous infusion on days 1-5, rapamycin 3mg on day 1 and 1mg on days 2-5; followed by autotransplant (AT) on day 6.

Results

Patient characteristics included age >=65, 19%; LDH>ULN, 43%; CA, 67%; GEP high-risk, 44%; GEP MF, 15%; GEP delTP53, 22%; prior AT, 75% including 46% with 2AT and 12% with 3 AT. CR and near-CR status was documented in 50%; TRM occurred within 100 days in 6%. Three-year estimates of overall survival (OS) and event-free survival (EFS) were 34% and 18%; OS/EFS were superior: (a) without CA – 60%/50% versus 15%/5% with CA (p=0.003/0.0005); (b) with GEP low-risk – 55%/20% versus 10%/15% with high-risk MM (p=0.009/0.01); (c) with GEP Hyperdiploidy and Low Bone disease molecular subgroups – 70%/35% versus 20%/10% in the remainder (p=0,001/0.002); and (d) absence of progression prior to SB – 52%/35% versus 16%/5% for the remainder (p=0.002/0.0006). Univariately significant adverse variables for OS and EFS included low albumin, B2M>=3.5mg/L, CA, GEP high-risk and relapse just prior to SB, of which B2M, CA and pre-SB survived on multivariate analysis for the 78 patients with all variables; among the 57 with GEP data, OS was inferior with GEP high-risk MM (HR=3.55, p=0.007) whereas EFS was inferior with high LDH (HR=3.44, p=0.004), CA (HR=2.79, p=0.02) and pre-SB relapse (HR=2.94, p=0.035).

Conclusions

We conclude that SB is a safe salvage regimen that was well-tolerated for the majority in the outpatient setting. Although beneficial to patients lacking CA and GEP high-risk status, the poor outcome also with SB in the presence of these adverse features emphasize the urgent need to discover agents/strategies effective in this setting.

Disclosures

van Rhee: Genzyme Corporation: Honoraria, Membership on an entity's Board of Directors or advisory committees, Research Funding.
Long Term Results of BL22 (CAT-3888) in Multiply Relapsed Hairy Cell Leukemia.
Authors: Kreitman Robert J , Wilson Wyndham H , Stetler-Stevenson Maryalice , Noel Pierre , FitzGerald David J. , Pastan Ira
Published: October 2019

Abstract 3442

Poster Board III-330

BL22 is a 63 kDa anti-CD22 recombinant immunotoxin containing truncated Pseudomonas exotoxin and variable domains from an anti-CD22 antibody. Patients with multiply relapsed/refractory hairy cell leukemia received BL22 and achieved 19 (61%) complete remissions (CRs) and 6 (19%) partial responses (PRs) in phase I testing, and 17 (47%) CRs plus 9 (25%) PRs in phase II testing (n=36), for overall response rates (ORR) of 72-81%. The average dose/cycle was the same for phase I and II (29 vs 33 ug/Kg x3). The dose for phase II was 40 ug/Kg x3 initially and 30 ug/Kg x3 for retreatment, but retreatment was held if patients had hematologic remission (HR, neutrophils ≥ 1500/mm3, Hgb ≥ 11 g/dL, and platelets ≥ 100,000/mm3) after cycle 1. Disease-free survival (DFS, CR duration) for phase II has not yet been reached at a median of 32 (range 4-62) months, with 12 (71%) of 17 CRs still ongoing. Considering all 36 CRs from phase I and II testing, median DFS was 33 (3-112) months with 15 (42%) of 36 CRs ongoing. Patients in CR usually underwent bone marrow biopsy every 6 months for 2 years and yearly thereafter, and after relapsing usually remained in HR. In fact, the median HR duration of these patients has not yet been reached at 42 (range 4-112) months, with 24 (67%) of the 36 patients remaining in HR or CR. Outcomes were better for those with pre-BL22 spleens measuring ≤ 200 mm in height than those with either prior splenectomy or spleens > 200 mm, in terms of CR (68% vs 34%, p=0.007), ORR 95% vs 48%, p=0.000003), and DFS (median 69+ vs 27 mo, p=0.002). In contrast, CR rates or DFS was not related to whether patients had <1 (n=28) or >1 (n=8) years of response to their last course of purine analog (p=0.5-0.75). Of 69 patients who received BL22, 8 (12%) had a completely reversible hemolytic uremic syndrome (HUS) and all maintained normal renal function after a median 80 (9-112) months of follow-up. We conclude that BL22 is highly active producing durable remissions in chemoresistant HCL, particularly in patients with limited disease burden. Testing is underway with a high-affinity version of BL22, called HA22 (CAT-8015).

Disclosures

Kreitman: NIH: Patents & Royalties. Off Label Use: BL22 is a recombinant immunotoxin which targets CD22+ cells. FitzGerald:NIH: Patents & Royalties. Pastan:NIH: Patents & Royalties.
A Study of Aberrant DNA Methylation in Refractory and Relapsed Childhood Acute Lymphoblastic Leukaemia.
Authors: Kapatai Georgia , Nyangoma Stephen O , Kearns Pamela
Published: October 2019

Abstract 3456

Poster Board III-344

Refractory and relapsed Acute Lymphoblastic Leukaemia (ALL) raise difficult clinical challenges and represent a significant cause of death in childhood and adolescence. This study examines epigenetic changes in relapsed paediatric ALL to increase our understanding of the mechanisms of treatment failure in these patients. Aberrant DNA methylation, a frequent phenomenon in paediatric ALL, has been identified as an independent factor of poor prognosis. In this study, using a CGI microarray consisting of 12K clones representing 5411 unique chromosomal sequences spread throughout the genome, we analysed a cohort of 22 childhood B-precursor ALL patient samples, 11 diagnosis samples and 11 relapse samples (unmatched) from patients who experienced a hematologic relapse, and developed genome-wide methylation profiles of leukaemic cells, at diagnosis and relapse. Comparison of these profiles identified 227 genes with statistically significant methylation changes – increase and decrease of methylation levels - between the two patient groups. The methylation status and the methylation change of 5 genes (ECRG4, CDH5, IRX3, FOXD3 and ERBB4) were independently verified, using pyrosequencing, in 6 ALL patients, 3 diagnosis and 3 relapse samples. The relationship between methylation and expression of one of these genes (ECRG4) was examined in ALL cell lines (CEMC1 and Jurkat) before and after 5-Aza-2'-deoxycytidine treatment. More genes are currently under investigation. Methylation studies on the repetitive element LINE-1 showed a significant increase in methylation of repetitive sequences in relapse samples (mean 79% vs. 89%, respectively; p=0.003, Mann Whitney test). Chromosomal mapping of the 227 genes revealed significant clustering of differentially methylated targets in chromosomes 11, 13 and 15, chromosomal locations that were previously found to be associated with ALL prognosis. Functional gene relation analysis revealed a connection between 11 genes (FOXD3, HPGD, NR4A2, CAST, TBRG4, XRCC6, NAB2, SEZL6, CDH13, NR2F1, SP1) that show increased methylation in relapse ALL and NR3C1, the gene coding for glucocorticoid receptor. Currently we are investigating the role of methylation in glucocorticoid resistance in ALL cell lines. Our data indicate an underlying epigenetic mechanism responsible for the chemoresistance observed in relapsed paediatric ALL patients.

Disclosures

No relevant conflicts of interest to declare.
Prospective Evaluation of An Agvhd-Specific Proteomic Pattern in More Than 340 Patients.
Authors: Mischak-Weissinger Eva M. , Holler Ernst , Schleuning Michael , Beutel Gernot , Greinix Hildegard , Dickinson Anne M , Ferrara James LM , Buchholz Stefanie , Krons Annika , Kontsendorn Julia , Hahn Norbert , Zander Axel R. , Kolb Hans-Jochem , Metzger Jochen , Ganser Arnold
Published: October 2019

Abstract 347

Allogeneic hematopoietic stem cell transplantation (allo-HSCT) is a curative treatment for many hematologic malignancies or hematopoietic dysfunction syndromes, but the application is still limited due to major complications, such as severe graft versus host disease (GvHD) and infectious complications. Diagnosis GvHD is based on clinical features and biopsies, a non invasive, unbiased laboratory test does not exist.

To date the proteomic pattern specific for aGvHD was evaluated blindly on 961 samples collected from 345 patients undergoing allo-HSCT at MHH and 7 additional clinics, including the University of Michigan. The majority of the patients included were transplanted for hematological malignancies (n=329), 16 for hematopoietic failure syndromes, mainly severe aplastic anemia. Conditioning regimens included dose reduced conditioning regimens (FLAMSA and ClaraC for the majority of the patients of MHH), as well as standard conditioning regimens (TBI+Cy or Busulfan+Cy) for about 35% of the patients. GvHD-prophylaxis was cyclosporine A (CSA) and mycophenolate (MMF) or CSA metothrexate (MTX) as appropriate. In addition, about 80% percent of the patients received ATG (antithymocyte globulin) prior to HSCT.

A peptide pattern of 31 peptides - either absent/decreased (15) or present /increased (16) - was previously published (Weissinger et al. 2007). Prospective and blinded evaluation of the patients included in this diagnostic analysis for early recognition of patients at risk for aGvHD development revealed the correct classification of patients developing aGvHD about 7 days prior to the development of clinical symptoms for aGVHD with a sensitivity 76% and specificity of about 85% (fig.1). Additional data obtained from patients transplanted until September 2009 will be reported. Based on these data a pre-emptive therapy multicenter trial, administering steroids upon positivity of the proteomic pattern has been initiated now in 10 German centers, testing the efficacy of the pre-emptive therapy on incidence and severity of aGvHD and a possible benefit on overall survival of the patients.

Disclosures:

Krons: mosaiques-diagnostics GmbH: Employment. Metzger:mosaiques-diagnostics GmbH: Employment.
Changes in Platelet Activation Associated with Hypertensive Disorders of Pregnancy.
Authors: Bouchard Beth A. , Schonberg Adrienne , Badger Gary J. , Bernstein Ira M.
Published: October 2019

Abstract 3504

Poster Board III-441

Pre-eclampsia is associated with increased rates of platelet clearance, changes in platelet function, and platelet activation. The goal of the current study was to assess changes in basal levels of platelet activation in individuals with pregnancies with hypertensive complications, and to compare those levels to individuals with uncomplicated pregnancies. In this study, three indices of platelet activation, platelet CD63 expression (% CD63), monocyte/platelet aggregate (% MP) formation, and neutrophil/platelet aggregate formation (% GP) were measured in whole blood by flow cytometry using specific, fluorescently-labeled monoclonal antibodies in 20 nulliparous women of reproductive age who subsequently conceived singleton pregnancies. Subjects were re-examined in early (11 - 16 wks) and late pregnancy (31 - 34 wks), as well as ∼15 months postpartum. Three of these women were diagnosed with hypertensive complications (1 gestational hypertension with intrauterine growth restriction, 2 pre-eclampsia), and delivered at term. However, all subjects were normotensive at all study points. The data are expressed as the mean ± SEM. P < 0.05 is considered significant. Subjects with uncomplicated pregnancies were 29.4 ± 0.77 years old with a BMI of 23.6 ± 0.83 kg/m2 at the time of the pre-pregnancy studies. Subjects with pregnancies with hypertensive complications were 32.0 ± 1.5 years old with a BMI of 21.3 ± 0.88 kg/m2 at the time of the pre-pregnancy studies. No differences in the platelet activation markers, % CD63 and % GP, were observed in this study. In contrast, a strong trend for increased % MP formation in pre-pregnancy measurements in those women destined to develop hypertensive complications of pregnancy was observed (p=0.062): Women with pregnancies with hypertensive complications had substantially more % MP formation (20.9 ± 10.3%) than women with uncomplicated pregnancies (7.1 ± 4.2%). In individuals with uncomplicated pregnancies, an increase in the % MP formation over pregnancy was observed over time, and this platelet activation marker remains elevated (20.2 ± 4.4%, p=0.015) in postpartum measurements when compared to pre-pregnancy values. A markedly different pattern was observed in women with pregnancies with hypertensive complications with postpartum values below the pre-pregnancy values (5.3 ± 12.2% vs. 20.9 ± 10.3%). These combined observations suggest that hypertensive complications of pregnancy are associated with alterations in the pattern of platelet activation in pregnancy and postpartum.

Disclosures:

No relevant conflicts of interest to declare.
Decitabine Increase Platelet Count by Two Mechanisms.
Authors: Wang Jianhui , Li Zongdong
Published: October 2019

Abstract 3507

Poster Board III-444

Thrombocytopenia is frequently associated with the myelodysplastic syndromes (MDS). 5-aza-2'-deoxycytidine (Decitabine) has been used to treat MDS with an approximately 20% response rate in thrombocytopenia. In this study, we have investigated the effect of Decitabine on platelet count in mouse. We report here that enhanced platelet release and maturation of megakaryocyte are two mechanisms involved in Decitabine induced elevation of platelet count. We first noted that a 30% of platelet count increase was found in the Balb/c mice 12 hours after the injection of Dectiabine at a clinically relevant dose (15 mg/m2) suggesting an instant platelet release from spleen or from megkaryocyte of bone marrow. The effect of Decitabine on megakaryocyte maturation was studied in in vitro differentiation of mouse bone marrow cells and megakaryoblastic cell line L8057. Decitabine (2.5 mm) is able to induce L8057 cells to differentiate into a megakaryocyte like polyploidy cells with positive marker of acetylcholinesterase and αIIb integrin. High expression of αIIb integrin was also found in the primary bone marrow cells cultured with both thrombopoietin and Decitabine as compared to thrombopoietin alone. The demethylation-induced transcription of GP6 has been reported in thrombopoietin induced megakaryocyte differentiation. Since Decitabine is a DNA demethylation reagent, we have investigated the GP6 expression in Decitabine treated L8057 cells and have found upregualtion of GP6 expression. Although the role of DNA demethylation in megkaryoctye differentiation still needs to be verified, our current data support that Decitabine is able to drive magakaryocyte maturation.

Disclosures:

No relevant conflicts of interest to declare.
A New Immunodeficient Mouse Strain, NOD/SCID IL2Rγ−/− SGM3, Promotes Enhanced Human Hematopoietic Cell Xenografts with a Robust T Cell Component.
Authors: Wunderlich Mark , Chou Fu-Sheng , Mizukawa Benjamin , Link Kevin A , Mulloy James C.
Published: October 2019

Abstract 3524

Poster Board III-461

Several strains of immunodeficient mice have been employed in the effort to model both normal and aberrant growth and differentiation of human hematopoietic cells in vivo. Despite numerous successes in this field, room remains for improvement. For example, robust models of in vivo differentiation and expansion of human erythroid and T cells are lacking or challenging at best. With the goal to further improve in vivo human hematopoietic cell study, we have crossed the NOD/SCID IL2Rγ knockout mice (NSG) with the NOD/SCID-SGM3 mice (NS-SGM; transgenic expression of human cytokines SCF, GM-CSF, and IL-3) to create a new strain of immunodeficient mice (NSG-SGM3). Uncultured umbilical cord blood CD34+ cells (UCB) were used to initiate xenografts in these three strains as well as NOD/SCID (NS) mice for comparison. Although xenografts with high human composition can be set up in NS mice, the graft, which is B-lymphoid biased without any measurable T cell component, tends to fade over time. In comparison, NSG mice offer longer sustained engraftment with a predominantly B-lymphoid bias and an inconsistent minor T cell component that is absent in NS mice. These mice also display increased levels of human cells in the periphery, allowing for easier measurement of the graft. Similar to NS grafts, NS-SGM3 grafts are initiated at high levels of chimerism, decrease significantly over time, lack T cells, and have a relatively low peripheral involvement. However, the graft consists of a more balanced myeloid/lymphoid distribution. The NSG-SGM3 combines some of the useful properties of the parent strains. Grafts remain at high, sustained levels through 16 weeks while maintaining a fairly balanced myeloid/lymphoid contribution. Chimerism in the NSG-SGM3 mice is readily detectable in the periphery as well as the BM. Remarkably, as early as 8 weeks post-transplant, we observed consistently high human T cell engraftment and expansion far superior to what we observed in NSG mice in parallel experiments using the same UCB samples. Flow cytometric analysis revealed the presence of numerous T cell sub-populations, suggesting significant recapitulation of human T cell development and function in NSG-SGM3 mice. Although it remains unclear as to why the unique combination of SGM3 cytokine expression and IL2Rγ knockout promotes enhanced human T cell engraftment and expansion, this mouse presents an improved opportunity to examine human T cell development and function in a xenograft setting as well as modeling T cell associated diseases including HIV infection, GVHD, and T cell leukemia.

Disclosures:

No relevant conflicts of interest to declare.
Survival Advantage with Hypomethylating Agents In Patients with Higher Risk Myelodysplastic Syndrome
Authors: Park Young-Hoon , Lee Je-Hwan , Lee Kyoo Hyung , Lee Jung-Hee , Kim Dae-Young , Kim Sung-Doo , Lim Sung-Nam , Choi Yunsuk , Seol Miee , Lee Young-Shin , Kang Young-A , Jeon Mijin
Published: October 2019

Abstract 1859

Background:

Two hypomethylating agents, azacitidine and decitabine, are active in myelodysplastic syndrome (MDS) and both drugs have been approved for the treatment of MDS in Korea. In this retrospective analysis, we tried to analyze the effects of hypomethylating agents on the clinical outcomes of the patients with MDS.

Methods:

A total of 385 patients, who were diagnosed as MDS at the Asan Medical Center between July 1992 and March 2010, were included in this study. We divided the patients into three treatment groups such as hypomethylating therapy (HMT, n=92), intensive chemotherapy (IC, n=15), and supportive care (including low-dose cytarabine and immunosuppressive treatment, SC, n=278). Primary end points of this study were overall survival (OS) and progression-free survival (PFS). PFS was defined as time from diagnosis to AML progression or death. All survival data were censored at the time of hematopoietic cell transplantation (HCT) to eliminate the influence of HCT on survivals. The difference of survivals was compared using time updated Cox model because times from diagnosis of MDS to treatment (HMT or IC) were various.

Results:

Baseline clinico-laboratory features were not significantly different between 3 treatment groups (HMT vs. IC vs. SC) in regard to sex, age, WHO subtype, and IPSS risk category. Among 83 patients in the HMT group, 39 (47.0%) attained any response to HMT including complete response, partial response, marrow complete response, and hematologic improvement. Survival data were the followings: OS, HMT vs. IC vs. SC, median 62.5 vs. 16.7 vs. 21.0 months; PFS, median 37.7 vs. 13.8 vs. 17.8 months. Multivariate analyses by stratified time updated Cox model showed that hazard ratio (HR) of HMT compared to SC was 1.029 (95% CI, 0.670–1.582, P=0.895) for OS and 0.539 (95% CI, 0.362–0.803, P=0.002) for PFS. When we performed subgroup analyses in lower risk disease (IPSS Low/Intermediate-1) and higher risk disease (IPSS Intermediate-2/High), the effects of HMT on survivals were different by the risk stratification of MDS. In lower risk MDS, there were no survival benefits of HMT compared to SC (OS, median 53.9 vs. 39.8 months, HR 1.570, P=0.108; PFS, median 37.7 vs. 52.5 months, HR 0.880, P=0.633), whereas both OS and PFS were significantly longer in HMT compared to SC (OS, median 137.9 vs. 5.9 months, HR 0.289, P=0.043; PFS, median 80.2 vs. 0.9 months, HR 0.154, P<0.001).

Conclusion:

Our data suggested that hypomethylating therapy could improve the survivals (OS and PFS) of the patients with higher risk MDS.

Disclosures:

No relevant conflicts of interest to declare.
PI3K/AKT Pathway as a Potential Therapeutic Target In Myelodysplastic Syndrome
Authors: Rezende Denise C , Pacheco Lorena Zaida , Pelloso Luis Arthur F. , Chauffaille Maria L. , Silva Marçal C.A , Kimura Elisa , Casaes-Rodrigues Rafael L , Segreto Helena , Yamamoto Mihoko , Kerbauy Daniella Marcia Maranhao Bahia
Published: October 2019

Abstract 1871

Introduction:

PI3K/AKT pathway is involved in cell growth, proliferation and apoptosis. A key downstream effector is the phosphorylated serine-threonine Akt (p-AKT). Constitutive activation of PI3K/AKT has been observed in solid tumours and leukemic cells. Inhibition of PI3K/AKT activity, results in apoptosis in cell lines (CL) after treatment with different compounds, e.g. deguelin, a natural product from the leguminous Mundulea sericea, with antitumour effects. Aims: To evaluate PI3K/AKT activation in MDS patients and its therapeutic potential in MDS.

Methods:

PI3K/AKT activation was evaluated by flow cytometry (FC) using an alexa-fluor 488-antibody Ser 473 p-AKT (Cell Signalling Technology). A triple immunostaining procedure using CD45-PerCP and CD34-PE was used for p-AKT expression in CD34+ primary samples. The p-AKT activity was determined using Kolmogorov-Smirnov test (D). CD34+ cells from healthy donors and Jurkat cells were used as negative and positive controls respectively. Apoptosis (determined by Annexin V and PI/7AAD) and cell cycle arrest (using RNAse and PI) were determined following treatments with LY294002 (50uM), and deguelin (100-500nM) in P-39 myeloid leukemia cell line, with constitutive PI3K/AKT activation. Apoptosis was determined in bone marrow mononuclear cells and CD34+ cells from MDS patients with the same treatments. To evaluate in vivo activity of deguelin, we used a xenotransplant model. Briefly, NODSCID mice were injected intrafemurally with P-39 CL and 12 days post transplant a three week-course of treatment, every other day, was started (deguelin 4mg/Kg, n=3 vs vehicle, n=3).

Results:

P-39 CL showed constitutive PI3K/AKT activation with levels significantly higher than in CD34+cells from controls (median±SD= 0.73.

Disclosures:

No relevant conflicts of interest to declare.
The Optimal Duration of Anticoagulant Therapy In Patients with Cancer-Related Deep Vein Thrombosis: The Advantage of Using Residual Vein Thrombosis (the Cancer-DACUS Study)
Authors: Siragusa Sergio , Malato Alessandra , Mascheroni Doris , Ageno Walter , Bucherini Eugenio , Spadaro Pietro , Imberti Davide , Saccullo Giorgia , Coco Lucio Lo , Rotondo Stefano , Santoro Rita , Micco Pierpaolo Di , Oriana Vincenzo , Urbano Oreste , Prantera Tullio , Recchia Francesco , D'Alessio Andrea , Gallucci Pina , Ghirarduzzi Angelo , Cormaci Oreste , Caracciolo Clementina , Abbadessa Vincenzo
Published: October 2019

Abstract 190

Type and duration of anticoagulation is still matter of debate in cancer patients with acute Deep Vein Thrombosis (DVT) of the lower limbs. Residual Vein Thrombosis (RVT) has been proven to be effective for assessing the optimal duration of oral anticoagulants in non cancer patients (Siragusa S et al Blood 2008:112:511-5). In the present study we evaluate the role of a RVT-based management of anticoagulation with Low-Molecular Weight Heparin in cancer patients with acute DVT.

Materials and Methods.

Patients with active cancer and a first episode of DVT were treated with LMWH for 6 months (the first month at full dosage followed by dose reduction of 25% in the next 5 months). At the end of treatment, they were managed according to RVT findings: those with RVT were randomized to continue anticoagulants for 6 additional months (Group A1) or to stop it (Group A2), while patients without RVT stopped LMWH (Group B). Outcomes were recurrent venous thromboembolism and/or major bleeding; patients were followed up for one year after LMWH discontinuation.

Results.

Over a period of 36 months, 409 patients were evaluated; 62 were excluded (refusal, need for continuing anticoagulation, etc). In total, 347 were included in the study (Table 1). RVT was detected in 242 (69.7%) patients; recurrent events occurred in 21.9% of those randomized to discontinue and 14.2% of those who continued LMWH. In patients without RVT (105, 30.3%), recurrent events occurred in 3 cases (2.8%) (Table 2 and Figure 1). The adjusted Hazard Ratio (HR) for age and sex between RVT groups (Group A2 vs A1) was 1.58 (95% confidence interval [CI], 0.85–2.93; P=.145). The adjusted HR between group A1 versus RVT-negative group (B) was 4.54 (CI 2.3–6.66; P =.028). Five major bleeding events occurred in Group A1 and two events both in Group A2 and B (Table 2). Overall, 89 (25.6%) patients died due to cancer progression after a median follow-up of 10.2 months after heparin withdrawn.

Conclusions.

The Cancer DACUS is the first ever study evaluating an individual marker for assessing duration of anticoagulation in active cancer population. Final results of the study show that absence of RVT identifies a group of patients at low risk for recurrent thrombosis who can safely stop LMWH after 6 months.

Disclosures:

No relevant conflicts of interest to declare.
Hydroxyurea Treatment In 1075 Patients with Essential Thrombocythemia and Occurrence of Extra-Hematological Adverse Events: A Preliminary Report of the Registro Italiano Trombocitemia (RIT)
Authors: Gugliotta Luigi , Papayannidis Cristina , Piccin Andrea , Tieghi Alessia , Vianelli Nicola , Scalzulli Potito Rosario , Candoni Anna , Dragani Alfredo , Patriarca Andrea , Lunghi Monia , Ciancia Rosanna , Martinelli Vincenzo , Liberati Anna Marina , Specchia Giorgina , Imovilli Annalisa , Vannucchi Alessandro M. , Di Gaetano Rosa , Randi Maria Luigia , Passamonti Francesco , Palmieri Fausto , Mazzucconi Maria Gabriella , Mastrullo Lucia , Cimino Giuseppe , Pierri Ivana , Cacciola Emma , Lucchesi Alessandro , Luraschi Annalisa , Martinelli Giovanni , Baccarani Michele
Published: October 2019

Abstract 1973

Background.

Hematological and extra-hematological adverse events associated to Hydroxyurea (HU) treatment in patients with chronic myeloproliferative neoplasms (MPN) are object of particular interest in the Ph-negative MPN since they can significantly limit the HU use.

Objective.

To evaluate the extra-hematological adverse events associated to HU treayment in a large cohort of Essential Thrombocythemia (ET) patients.

Material and Methods.

One thousand and seventy-five out of the 2005 ET patients registered in the RIT were treated with HU and are the object of this report. The patients, 641 females (59.6%) and 434 males (40.4%), diagnosed according to the PVSG or WHO criteria in 54 hematological centers of the RIT, started treatment with HU as first (92%) or second (8%) line, at median age of 65 years.The mean duration of HU treatment was 3.3 years. The HU treatment was withdrawn in 221 (20.5%) patients after a mean of 3.0 years.The administered dose of HU was 0.25–3.0 g/day (median 1), and a mean cumulative dose was 1113 g. The extra-hematological adverse events (EHAEs) observed during the HU treatment were distinguished in HU related AEs (HU-EHAEs), ET related AEs (ET-EHAEs) and HU or ET unrelated AEs (U-EHAEs).Results. During the HU treatment (3587 pt-y) 378 EHAEs were reported in 207 (19.3%) patients, being the HU-EHAEs 244 (6.8/100 pt-y) in 170 (15.8%) patients. In detail, the HU-EHAEs were: dermatological in 108 (48.3%) cases (38 hyperpigmentation, 26 leg ulcers, 22 maculo-papular rash, 10 lichenoid eruptions, 5 skin cancer, 4 alopecia); gastro-intestinal in 80 (32.8%) cases (37 nausea/vomiting, 30 diarrhea, 13 gastro-intestinal intolerance); systemic in 35 (14.3%) cases (28 fever, 7 fatigue); neurological in 19 (7.8%) cases (headache); miscellanea in 2 (0.8%) cases.

Conclusion.

This preliminary analysis in 1075 ET patients of the Registro Italiano Trombocitemia (RIT) treated with HU shows that the extra-hematological adverse events referred to the HU (HU-EHAEs) occurred with not negligible rate (6.8/100 pt-y) and need to be object of attention in the management of ET patients.

Disclosures:

No relevant conflicts of interest to declare.
A Germinal Center- Derived B Cell Lymphoma Model
Authors: Phan Ryan T , Nguyen Khang , Romero Sonia , Nicolson Alice , Nham Phillipp , Nguyen Dany V.
Published: October 2019

Abstract 2008

Most human B-cell lymphomas represent mature phenotypes of germinal center (GC) or post-GC origin and are frequently associated with chromosomal translocations, often involving the rearrangement of immunoglobulin (Ig) loci to various cellular oncogenes, leading to oncogenic activation. The mechanisms underlying these processes, however, are not well understood. Several studies suggest that these genetic lesions arise from errors of physiologic DNA rearrangements in GC B cells, namely class switch recombination (CSR) and somatic hypermutation (SHM). Here we report the generation of a mouse model in which DNA breaks are physiologically instituted in mature B cells, yet inefficiently repaired via specific deletion of DNA repair gene XRCC4 in GC B cells, thus effectively creating an in vivo environment for errors in DNA rearrangements. These activated B cells exhibit significant increased chromosomal IgH locus breaks and reduced CSR. In p53-deficient background, these mice develop B-cell lymphoma from 5.5 to 16 months. These clonally developed tumors characteristically harbor chromosomal translocations and phenotypically resemble mature phenotypes. Many of these tumors bear mutated V genes, suggesting that those cells have transited through GC. Thus, this mouse model mimics human B-cell lymphoma and might be useful for the development of therapeutic interventions in B-cell lymphoma.

Disclosures:

No relevant conflicts of interest to declare.
An Important Role for Rac1 and RhoA In Regulating Platelet Microparticle Formation and Phosphatidylserine Exposure
Authors: Delaney Michael Keegan , Liu Junling , Zheng Yi , Du Xiaoping
Published: October 2019

Abstract 2023

Platelets activated by physiological agonists such as thrombin and collagen shed procoagulant microparticles (MPs) and externalize the procoagulant phospholipid phosphatidylserine (PS), both of which are critical to hemostasis and play an important role in inflammation. To date, the signaling mechanisms that regulate agonist-induced MP formation and PS exposure in platelets remain unclear. In this study, we demonstrate that the small GTPases Rac1 and RhoA play important roles in regulating the procoagulant activity of platelets. Rac1 null (-/-) mouse platelets or human platelets treated with the Rac1 inhibitor NSC23766 (NSC) displayed a significant defect in MP formation and PS exposure induced by various agonists. Furthermore, Rac1-/- platelets and NSC-treated human platelets displayed a defect in procoagulant activity as demonstrated by a prolonged coagulation time following recalcification of citrated PRP. The stimulatory role of Rac1 in platelet MP formation and PS exposure is distinct from the known function of Rac1 in facilitating platelet granule secretion and secretion-dependent amplification of platelet aggregation, because supplementation of the granule content ADP rescued the defect in platelet aggregation caused by Rac1 inhibition, but failed to rescue the defect in MP formation caused by Rac1 inhibition. In contrast to Rac1, RhoA plays an inhibitory role in regulating platelet procoagulant activity, because treatment of platelets with the Rho inhibitor C3-toxin (C3) significantly enhanced agonist-induced MP formation, PS exposure, and procoagulant activity. The enhancing effect of C3 on platelet procoagulant activity is not caused by an overall enhancement of platelet activation because C3 significantly inhibited platelet secretion and aggregation. Thus, our data demonstrates that while Rac1 and RhoA both play important stimulatory roles in platelet granule secretion and aggregation, they play opposing roles in MP formation and PS exposure in platelets. Rac1 is important for stimulating platelet MP formation, PS exposure, and procoagulant activity, which is antagonized by RhoA.

Disclosures:

No relevant conflicts of interest to declare.
Hepcidin Independent Anemia of Inflammation
Authors: Rivera Seth , Vaulont Sophie , Shprung Dana , Limbrick Donald , Ganz Tomas
Published: October 2019

Abstract 2050

Anemia of inflammation (AI, anemia of chronic disease) is a common complication of a wide variety of inflammatory conditions including infections and connective tissue diseases. Because the iron regulatory protein hepcidin is increased during inflammation and overexpression of hepcidin recapitulates many of the features of AI, hepcidin is currently believed to be the primary mediator of AI. Furthermore, Sasu and colleagues showed that blocking hepcidin could improve AI (Blood, Vol. 115, No. 17, pp. 3616-24). To test whether hepcidin is indeed necessary for the development of AI, we used two models of AI in hepcidin knockout mice. The first model, using heat-killed Brucella abortus to model brucellosis, is the same as that used by Sasu and colleagues. The second model uses complete Freund's adjuvant to model Mycobacterial disease. We showed that absence of hepcidin did not protect against the development of anemia. Furthermore, we show that this was an iron restricted anemia arguing that iron sequestration can occur independent of hepcidin. This study indicates that hepcidin may not be the central mediator in all AI. Future research will be needed to determine why the anti-hepcidin treatment used by Sasu and colleagues was able to improve the anemia.

Disclosures:

Ganz: Intrinsic Life Sciences: Employment, Membership on an entity's Board of Directors or advisory committees.