bluuebunny/crossref_metadata_2025_split huggingface dataset:
Molecular, Epigenetic, and Phenotypic Repolarization of T Lymphocytes From Chronic Lymphocytic Leukemia Patients Using 5-Aza-2′-Deoxycytidine
Authors: Dubovsky Jason A , Powers John , Gao Yang , Mariusso Luis F , Sotomayor Eduardo M. , Pinilla-Ibarz Javier 
Published: October 2019

Abstract
Abstract 4651
The improper polarization of helper and cytotoxic T lymphocytes is a trademark immunosuppressive strategy employed by chronic lymphocytic leukemia (CLL). Mounting data implicates chromatin regulation, namely promoter methylation, in the plasticity of naïve human T cells. Very recent in-vitro evidence indicates that this plasticity may be phenotypically replicated by using methylation inhibitors which are approved for clinical use in certain types of cancer, however the longevity and therapeutic potential of repolarized cells remains to be elucidated. These results beg the question: can the ineffective polarization of T lymphocytes in the context of CLL be effectively modulated using methylation inhibitors in a sustainable therapeutic fashion? To answer this question our laboratory has focused on repolarizing helper and cytotoxic T lymphocytes using 5-aza-2′-deoxycytidine (5A2) tracking changes in well characterized molecular and epigenetic signaling pathways involved in effective polarization. Moreover, we sought to investigate the consequences of methylation inhibitor treatment on lymphocyte survival, activation intensity, and naïve cell polarization. Our studies relied upon intracellular staining, methylation specific PCR, western blot, and phosflow anlaysis. The data indicates that 5A2 treatment can repolarize Th2 cells to effectively secrete interferon gamma, signal via T-bet, and achieve demethylation of critical Th1 specific promoters. We also show that this repolarization can be maintained. Moreover, we demonstrate that 5A2 can force Th1 polarization of naïve T cells despite a strong IL-4 stimuli and a lack of IL-12. Finally, we have identified a cytotoxic threshold for 5A2 treatment which is above the concentration necessary to achieve effective and sustained molecular and epigenetic repolarization. All in all our data seeks to define a modality in which improper or ineffective T cell polarization can be altered ex post facto and continued immunotherapeutic intervention may benefit from the naïve polarization of anti-cancer responses.

Disclosures:
No relevant conflicts of interest to declare.
Functional Characterization of a Recombinant FVIIa Drug Candidate
Authors: Turecek Peter L , Varadi Katalin , Vejda Susanne , Boehm Ernst , Rottensteiner Hanspeter , Schwarz Hans Peter , Mitterer Artur , Mundt Wolfgang , Ehrlich Hartmut J. , Scheiflinger Friedrich 
Published: October 2019

Abstract
Abstract 4653
Human coagulation factor VII (FVII) is a vitamin-K-dependent protein with a molecular weight of 50 kDa. Activation of FVII occurs by cleavage of the arginine 152 – isoleucine 153 peptide bond resulting in two disulfide-linked peptide chains. The therapeutic utility of rFVIIa is based on its capacity to trigger hemostasis independently from factor VIII and factor IX and thus even in the presence of factor VIII or factor IX inhibitors.
Baxter has developed a recombinant FVIIa (rFVIIa) that is synthesized by a genetically engineered Chinese hamster ovary (CHO) cell line. No materials of human or animal origin are employed in the manufacture, purification, or formulation of the final product, thus reducing the risk of transmission of adventitious agents. The growth medium is a chemically defined medium, and the downstream process does not use monoclonal antibodies for the purification of rFVIIa.
The rFVIIa drug candidate was functionally characterized in vitro and its features were compared with those of a commercially available rFVIIa. The overall hemostatic potency of rFVIIa was assessed by its FVIII bypassing activity in a human FVIII-deficient plasma with high-titer inhibitor by measuring the kinetics of thrombin generation and fibrin formation. The thrombin-generating capacity of the rFVIIa drug candidate was similar to that of commercial rFVIIa. The time and rate of fibrin formation was measured by thromboelastography, where a dose-dependent normalization of the impaired clotting times and fibrin formation was observed. A similar FXa-generating potency was found for rFVIIa and commercial rFVIIa when measured on the surface of TF-expressing fibroblasts, suggesting a full capability to bind to TF-bearing cells and trigger hemostasis on their surfaces. rFVIIa could be inactivated by anti-thrombin III-heparin in solution with relipidated TF with no relevant difference to the comparator product. TF pathway inhibitor effectively inhibited FXa generation in a cell-based activity assay, with a similar IC50 for rFVIIa and the comparator.
In summary, the functional characteristics of the rFVIIa drug candidate were very similar to those of commercially available rFVIIa.

Disclosures:
Turecek: Baxter Innovations GmbH: Employment. Varadi:Baxter Innovations GmbH: Employment. Vejda:Baxter Innovations GmbH: Employment. Boehm:Baxter Innovations GmbH: Employment. Rottensteiner:Baxter Innovations GmbH: Employment. Schwarz:Baxter Innovations GmbH: Employment. Reiter:Baxter Innovations GmbH: Employment. Mitterer:Baxter Innovations GmbH: Employment. Mundt:Baxter Innovations GmbH: Employment. Ehrlich:Baxter Innovations GmbH: Employment. Scheiflinger:Baxter Innovations GmbH: Employment.
A Case of Clopidogrel Induced TTP Recurring After Four Years
Authors: Methuku Nanda K. , Chandra Abhinav B. , Belur Anuradha , Dabrowski Lech 
Published: October 2019

Abstract
Abstract 4678

Case description -
A 61 year old woman was started on clopidogrel after having PTCA with stent placement in February 2006. Four weeks after starting clopidogrel she developed thrombocytopenia with platelet nadir of 17,000. Her LDH was 700 IU/L and she was anemic with hemoglobin of 7.4 gm/dl with elevated reticulocyte count. Peripheral blood smear showed schistocytes and diagnosis of TTP secondary to clopidogrel was made. She did not have renal insufficiency. Clopidogrel was discontinued and patient was started on plasmapheresis with recovery of platelet counts. Early attempts in weaning plasmapheresis resulted in drop in platelet count and Rituximab was given to the patient weekly for four weeks. Subsequently, patient was weaned off plasmapheresis. For four years patient was followed periodically with CBC showing platelet counts greater than 250,000. In May 2010, four years after initial event patient was admitted to hospital for abdominal pain and found to have splenic infarcts. Subsequently, she also developed bilateral cerebral infarcts. Platelet count had decreased to less than 100,000. Her LDH was elevated at 419 IU/L. Reticulocyte count was 2.3%. Peripheral blood smear revealed significant number of schistocytes. There was no renal insufficiency or fever. Trans-esophageal echocardiogram (TEE) was done that did not reveal any vegetations. Patient was diagnosed as having recurrence of TTP and started on plasmapheresis with recovery in platelet counts. Pt was also treated with Rituximab.

Discussion-
We describe a case of TTP initially occurring within weeks of starting clopidogrel. Patient was treated with plasmapheresis and Rituximab and clopidogrel was discontinued. Patient had recurrence after four years as manifested by infarcts in multiple organs, with mild thrombocytopenia, elevated LDH and significant number of schistocytes on peripheral blood smear. It is very uncommon for clopidogrel associated TTP to recur after such a prolonged period of 4 years. Most cases of clopidogrel associated TTP have mild thrombocytopenia. This patient had severe thrombocytopenia on first presentation of TTP but had mild thrombocytopenia on recurrence. This case illustrates the importance of extended follow up and high index of suspicion for TTP as delays in initiation of plasmapheresis has a poor clinical outcome.

Disclosures:
No relevant conflicts of interest to declare.
Anagrelid In Essential Thrombocythaemia Treatment of a Patient Under 12 Months
Authors: Laguna Pawel , Romiszewski Michal , Klukowska Anna , Pawelec Katarzyna , Matysiak Michal 
Published: October 2019

Abstract
Abstract 4680
Essential thrombocythaemia (ET) is a chronic myeloproliferative disorder. It occurs mostly in adults and rarely in young children. We present the case of ET in an 8 –month- old girl, who was treated with Anagrelid for 13 months.
The girl was admitted to the hospital because of elevated platelet count. She had stomach pains periodically. On admission the child's condition was good but on palpation her stomach was sore and her spleen was enlarged. Laboratory investigations showed an elevated platelet count (1.5m) and hypercalaemia (6.3mEq/l). Thrombopoetin was normal.
Acesan was added to the treatment. A morphology performed after 1 month revealed further elevation of platelets (up to 2.65m). On the basis of the laboratory investigations, bone-marrow biopsy and trepanobiopsy, we eliminated an oncological disease and infectious diseases of connective tissues.
On bone-marrow investigation, acquired mutation of JAK2 (V617F) tyrosine kinase was diagnosed, which confirmed ET.
On account of the growing number of platelets, which was life - threatening, we decided to administer Anagrelid, starting with a dose of 0.25 mg per day. The number of platelets decreased to 1.4m, so the dose was increased to 0,25 mg twice daily after 3 weeks.
At present, after 13 months of Anagrelid treatment, the number of platelets in the child ranges between 400 and 650.
The patient comes to our, department for monthly check-up of morphology, biochemical tests, ECG and echocardiography.
In international literature there is no information on the use of Anagrelid in ET treatment of children under 12 months old. However, on the basis of our observations and initial results of treatment, it seems that the above protocol for Anagrelid administration is safe for infants of this age.

Disclosures:
No relevant conflicts of interest to declare.
Efficacy of Lower Dose Rituximab by Adding Fresh Frozen Plasma for the Treatment of Idiopathic Thrombocytopenic Purpura
Authors: Sun Mei , Gu JIan , He Bin , Xie Xiao-Yan , Chen Bao-An , Pei Xiao-Ping 
Published: October 2019

Abstract
Abstract 4682

OBJECTIVE:
To evaluate the effectiveness and safety of lower dose rituximab by adding fresh frozen plasma in the treatment of patients with idiopathic thrombocytopenic purpura (ITP).

METHODS:
A prospective study was performed at Subei People,Hospital Affiliated to Yang Zhou University involving the use of lower dose rituximab combining fresh frozen plasma in 15 patients who had previously been treated with steroids, intravenous immunoglobulin (IVIG) or splenectomy. Fifteen patients with refractory ITP which were unresponsive to or relapse after the above said treatment were treated with two hundreds millilitre of fresh frozen plasma (FFP) followed with rituximab (100 mg/m2 per week for four weeks) and oral Medrol (24mg per day). Whole blood cell count in all cases and B cells of CD19 (+)/CD20 (+) were detected in eight cases before and after rituximab therapy.

RESULTS:
Complete response (CR) was achieved in 7 patients (46.67%), response (R) in 13(86.67%), and non-response (NR) in 2 (13.33%). The median follow-up time was 4 weeks. The median response and CR time were 7 and 14 days, respectively. CD19 (+) CD20 (+) B cells significantly decreased (P < 0.01). There were no severe adverse effects during rituximab therapy.

CONCLUSION:
Lower dose rituximab by adding fresh frozen plasma is effective and safe for ITP. Our therapeutic regimen is superior to other relative studies; however, a randomized control trial is needed to confirm the results of our study.

Disclosures:
No relevant conflicts of interest to declare.
Thrombotic Thrombocytopenic Purpura Associated with Systemic Lupus Erythematosus and Antiphospholipid Syndrome: A Case Report
Authors: He Yang , Yang Weiwen , Ruan Changgeng 
Published: October 2019

Abstract
Abstract 4685

Background:
The occurrence of thrombotic thrombocytopenic purpura (TTP) in patients with systemic lupus erythematosus (SLE) is rare TTP frequence. In pregnancy women is increased. TTP with simultaneous SLE, and antiphospholipid syndrome (APS) occurring in the same patient is extremely rare.

Case report:
We described a 41-year-old Chinese female with TTP in her third trimester (33+4 gestational weeks) of pregnancy who suffered from SLE. and APS at the same time. The clinical features and positive laboratory abnormalities included fever, repeated abortion in first trimester, palpitation, dyspnea, orthopnea, hypertension (180/113mmHg), proteinuria, renal dysfunction, thrombocytopenia (platelets count 12×109/L), hemolytic anemia (Hb 67g/L), positive Coombs test, positive lupus test, positive antinuclear antibody, anti-Sm antibody, anti-insulin antibody, anti-ENA antibody and anti-RNP antibody. The level of plasma ADAMTS13 activity was below 5%, and ADAMTS13 inhibitor was present in the patient's plasma. This patient was treated with a combination of corticosteroids and plasma infusion and plasmapheresis. At 33+5 weeks of gestation she delivered a healthy infant via cesarean section. The patient improved quickly with these therapies.

Conclusion:
This is a rare TTP case with simultaneous SLE, APS and preeclampsia. Her overlaping clinical manifestations and laboratory abnormalities made the diagnosis very difficult. These data indicate that plasma therapy and corticosteroids can improve the prognosis of TTP associated with SLE and APS. In addition, the termination of pregnancy may be also helpful for controlling the development of the disease.

Disclosures:
No relevant conflicts of interest to declare.
Hematopoietic Reconstitution of Mice Cord Blood Transplantation Combined with Bone Marrow Transient Hematopoietic Progenitor Cell
Authors: Shi Jun , Ikeda Kazuma , Yosinobu Maeda , Yuan Yinghua , Yu Yehua , Tanimoto Mitsune 
Published: October 2019

Abstract
Abstract 4688
Umbilical cord blood has been increasingly used as a source of hematopoietic stem cells. However, fetal blood recipients had slower hematopoietic engraftment and impaired immune reconstitution. To accelerate myeloid and lymphoid recovery, we used an animal model of newborn blood modeled for cord blood (CB), along with transiently reconstituting progenitor cells from congenic bone marrow with recipients, as sources of stem cells. According to the previous reports that murine transiently reconstituting progenitor cells express the c-kit molecule, but not Sca-1 and lymphohematopoietic lineage markers, Lin-sca-1-c-kit+(c-kit+) were isolated by MACS method. c-kit+ cells population consisted of exclusively of medium- or large-sized blast-like cells, which displayed relatively low proliferative potential in vitro than Lin-sca-1+ (sca-1+) population. After transplantation of CB from DBA/2 mice (H-2d/d, CD45.2), with or without graded numbers of either c-kit+ or sca-1+ cells isolated from BDF1 mice (H-2d/b, CD45.1) bone marrow into lethally irradiated CD45.2 congenic BDF1 mice, hematopoietic engraftment were dynamic investigated. The intermingled transplantation of CB and c-kit+ cells or sca-1+ cells at the dosages of 1×104 or 2.5×104 or 5×104 to recipient mice leads to the quantity of white blood cells and platelets increased to 1×109/L and 1×1012/L at day12, whereas the injection of CB alone resulted in day17. By 2 weeks post-transplantation, congenic BM-derived cells were dominantly found in granulocytes and B lymphocytes, while host cells were dominantly found in T lymphocytes in CB transplantation combined either with c-kit+cells or sca-1+ cells. In cotransplantation with CB and c-kit+cells – engrafted surviving mice, the degree of donor CB cells in the peripheral blood increased progressively over time, while congenic donor BM-derived cells decreased gradually. After 60 weeks cotransplantation with CB and c-kit+ cells, a complete chimerism frequency of CB–derived cells continued to maintain in granulocytes and B lymphocytes, while T lymphocytes were dominantly derived from CB. On the other hand, congenic bone marrow or host-derived cells were the dominant population and CB-derived cells in the peripheral blood were less than 10% after 60 weeks cotransplantation with CB and sca-1+cells. In conclusion, the cotransplantation of CB and congenic c-kit+ cells was able to accelerate early hematopoietic recovery due to congenic marrow cells. But complete or main chimerism of cord blood was formed without or with fewer residual cells of host origin and congenic BM origin in long-term multilineage reconstitution. Thus, this cotransplant model in vivo may be to bring useful information for improving hematopoietic and immune reconstitution in fetal blood recipients.

Disclosures:
No relevant conflicts of interest to declare.
Cytokine Expressions In Peripheral Blood Mononuclear Cells During Graft-Versus-Host Disease After Allogeneic Hematopoietic Stem Cell Transplantation
Authors: Ye Fang , Qiao Zhenhua , Yang Tao , Zhang Li , Jia Ning 
Published: October 2019

Abstract
Abstract 4708

Objective
To investigate the relationship between cytokines and human graft-versus-host disease (GVHD) after allogeneic hematopoietic stem cell transplantation (allo-HSCT).

Methods
In 21 patients undergoing allo-HSCT, the plasma concentrations of cytokines (interleukin 2(IL-2), soluble interleukin 2 recepter (sIL-2R), interferon-gama (IFN-γ), interleukin 4(IL-4), transforming growth fator-beta1(TGF-β1)) were measured by using sandwich enzyme-linked immunological assay (ELISA) and the gene messanger RNA expressions of these five cytokines were analysed by using semi-quantitate reverse transcriptase-polymerase chain reaction (RT-PCR).

Results
1. The concentrations and gene messanger RNA expressions of IL-2, sIL-2R and IFN-γ in the patients with GVHD were significantly higher than those without GVHD (P<0.01); these three cytokines expression levels in the patients with aGVHD were higher than with cGVHD and without GVHD (P<0.05); The levels of IL-4 showed no difference between the patients with or without GVHD (P>0.05); the levels of TGF-β1 in the patients with GVHD were significantly declined (P<0.01) but in those without aGVHD were obviously increased (P<0.05). After effective treatment, unnormal IL-2, sIL-2R, IFN-γ and TGF-β1 expressions recovered to the levels before transplantation. 2. The results of multivariate logistic regression modles and Kaplan-Meier survival curves analyses showed: age, sex, infection, HLA-matched, blood type, conditioning regiment and the times of absolute neutrophil counts and platelets recovering to normal, had no association with the incidence of GVHD (P>0.05). A multivariate COX survival function modle analysis showed IL-2, sIL-2R and TGF-β1 are independent prognostic factors for GVHD (=11.149, P<0.0001). 3. The results of IL-2, sIL-2R, IFN-γ and TGF-β1 measured by RT-PCR and ELISA for detecting peripheral blood cytokines expression levels had no difference.

Conclusions
1. IL-2, sIL-2R, IFN-γ and TGF-β1 play important roles in the development of GVHD after allo-HSCT. Monitoring the changes of IL-2, sIL-2R, IFN-γ and TGF-β1 expression levels (especially IL-2, sIL-2R and TGF-β1) might provide predictive markers for GVHD, especially aGVHD. 2. Different conditioning regiments resultes in different IL-2, sIL-2R, IFN-γ and TGF-β1 expression levels. 3. IL-4 expression had no effect on the developing of GVHD after allo-HSCT. 4. The sensitivity between RT-PCR and ELISA for detecting peripheral blood cytokines expression levels had no difference.

Disclosures:
No relevant conflicts of interest to declare.
The Effect of Rhg-CSF on MDSC In Bone Marrow and Peripheral Blood Cells
Authors: Ling Yiwen , Liu Qifa , Fan Zhiping , Wu Xiuli , Liu Can , Li Xuan , Sun Jing 
Published: October 2019

Abstract
Abstract 4709

Objective:
As granulocyte colony-stimulating factor, recombinant human granulocyte colony stimulating factor (rhG-CSF) is widely used in neutropenic patients. In addition to stimulating the growth of granulocyte, rhG-CSF can promote hematopoietic stem cells from bone marrow (BM) to peripheral blood (PB) and has the effect of immune regulation. Myeloid-derived suppressor cells (MDSC) are a group of heterogeneous cells, derived from bone marrow progenitor cells and immature myeloid cells. Recently, MDSC is researched in the field of solid tumor, but not in the field of hematopoietic stem cell transplantation. Here, we investigate rhG-CSF's effect on MDSC in healthy donors’ BM, PB and the relationship between MDSC and graft-verse-host disease (GVHD).

Methods:
We obtained the BM and PB samples before mobilization and the BM APB and peripheral blood stem cell collection (PBSC) on the 5th day after the rhG-CSF mobilization from 12 healthy donors, respectively. Then we used the flow cytometry to check the absolute number of MDSC. Finally, we analyzed the relationship between the number of MDSC and the incidence of GVHD.

Results:
In normal physiological conditions, the MDSC could be detected in healthy donor's PB and BM. In PB, the proportion of MDSC in the mononuclear cells was 1.35 ± 0.35%. In BM, the proportion was 2.44 ± 1.11%. The proportion in BM is higher than that in PB, the difference was statistically significant (P=0.047). On the 5th day after the rhG-CSF mobilization, the MDSC ratio of mononuclear cell in PB were 4.01 ± 1.82%. In BM, the ratio was 4.38 ± 2.19%. The difference between the ratio of MDSC in BM and PB was no significant (P=0.076). The number of mobilized peripheral blood MDSC was significantly higher than that before mobilization (P=0.015), while the difference between the numbers of bone marrow MDSC cells before and after mobilization was not significant (P=0.083). The numbers of MDSC in collection and the incidence of GVHD had a significant negative correlation (P=0.048).

Conclusion:
MDSC could be detected in the healthy donors’ PB and BM, the numbers of MDSC in BM were higher than that in PB. The rhG-CSF could mobilize more MDSC from BM to the peripheral blood, and the increased s of MDSC in PB after rhG-CSF mobilization might be related to the low incidence of GVHD in hematopoietic stem cell transplantation.
Supported by National Natural Science Foundation of China (30971300), Science and Technology Planning Project of Guangdong Province of China (2009A030200007) and China Postdoctoral Science Foundation (200902332, 20080440776).

Disclosures:
No relevant conflicts of interest to declare.