bluuebunny/crossref_metadata_2025 huggingface dataset:
Acquired Von Willebrand Syndrome In Mitral Valve Leak
Authors: Pérez-Rodríguez Almudena , de Faria Maria Joana Costa Pinto Prego , Fraga Esther Lourés , Rodríguez-Trillo Angela , Cuenca José Joaquín , Batlle Javier , Fernanda López-Fernández Maria
Published: October 2019

Abstract 4657

Background

Several studies have shown that between 15% to 25% of patients with severe aortic stenosis present bleeding episodes that may be attributed to an acquired von Willebrand syndrome (AVWS). Until now, to our knowledge, no association of AVWS with mitral valve disfunction has been reported.

Design and Methods

Four patients with mitral valve leak presented acquired abnormalities of von Willebrand factor (VWF) and a bleeding history. Two of them presented severe bleedings requiring blood transfusions. All of them were within an adequate range of oral anticoagulation.

Results

Prior to surgery, these patients presented an APTT prolonged and in two of them the closure time determined by the platelet function analyzer (PFA-100®) (with COL/ADP and COL/Epi) was prolonged also. Factor VIII procoagulant activity (FVIII:C), VWF antigen (VWF:Ag), VWF ristocetin cofactor activity (VWF:RCo) and VWF collagen binding (VWF:CB) were considerably elevated and the VWF multimers in plasma, showed a lower relative proportion of the high molecular weight VWF multimers (HMWM), to some extent similar to type 2A congenital von Willebrand disease (VWD). In two of them, the VWF:RCo/VWF:Ag or VWF: CB/VWF:Ag ratios were less than normal range (>0.7) while in the other two were normal. After surgery, FVIII:C, and VWF properties were extremely increased and the ratios VWF:RCo/VWF:Ag and VWF: CB/VWF:Ag > 0.7. After surgery, FVIII:C, VWF:Ag, VWF:RCo and VWF:CB increased considerably. The ratios were > 0.7. The PFA-100® (COL/ADP and COL/Epi) was corrected in the two patients who had it prolonged. The multimeric VWF profile were also corrected in all of them.

Conclusions

The present study describes acquired VWF qualitative alterations for the first time in mitral valve leak. When such alterations are important they may be associated or to contribute to a bleeding diathesis. This problem was reported previously in aortic valve stenosis in relationship with a suspected very high shear stress. This situation may be not usually present in mitral valve stenosis, but it seems that it must occur in the presence of mitral valve leak. Consequently, the AVWS should be taken into account in patients with mitral valve leak that present a bleeding diathesis, not explained by an excess of oral anticoagulation.

ACKNOWLEDGEMENTS

This work was supported by the Fondo de Investigación Sanitaria, F.I.S. Carlos III, Ministerio de Sanidad, Spain (FIS PI# 07/0229), and Consellería de Innovación e Industria, Xunta de Galicia (INCITE08ENA916109ES).

Disclosures:

No relevant conflicts of interest to declare.
Biochemical Characterization of a Recombinant FIX Drug Candidate for Treatment of Hemophilia B
Authors: Turecek Peter , Vejda Susanne , Varadi Katalin , Rottensteiner Hanspeter , Boehm Ernst , Reiter Manfred , Kaliwoda Martin , Mundt Wolfgang , Ehrlich Hartmut J. , Scheiflinger Friedrich
Published: October 2019

Abstract 4658

Human coagulation factor IX (FIX) is a vitamin-K-dependent coagulation factor whose absence or dysfunction causes hemophilia B. Treatment of hemophilia B is based on replacement therapy using highly purified FIX concentrates.

Baxter has developed a recombinant factor IX for treating hemophilia B patients that is produced in a CHO cell-line using a serum and protein-free fermentation technology. The purification process avoids the use of immune-affinity chromatography and includes two viral reduction steps. The final drug product is formulated in the absence of proteins of animal or human origin. Baxter's recombinant FIX resembles commercially available rFIX in most characteristics with the exception of a significantly lower FIXa content, which might improve standardization compared to commercial rFIX products.

Preclinical and clinical lots of rFIX were characterized with respect to their hemostatic potency, efficiency of activation by FXIa and FVIIa in the presence of tissue factor, and capacity to bind to phospholipid vesicles. Three lots of commercial rFIX with different potencies and one lot of a plasma-derived FIX product were included in the study. Similarity could be shown between the preclinical and clinical lots of rFIX in all these assays. Furthermore, the functional and biochemical characterization of Baxter's recombinant FIX showed that it resembles the recombinant comparator product. The phase I clinical trial which has been initiated will now have to show whether Baxter's rFIX can become an alternative drug candidate product for treating patients suffering from hemophilia B.

Disclosures:

Turecek: Baxter Innovations GmbH: Employment. Vejda:Baxter Innovations GmbH: Employment. Varadi:Baxter Innovations GmbH: Employment. Rottensteiner:Baxter Innovations GmbH: Employment. Boehm:Baxter Innovations GmbH: Employment. Reiter:Baxter Innovations GmbH: Employment. Kaliwoda:Baxter Innovations GmbH: Employment. Mundt:Baxter Innovations GmbH: Employment. Ehrlich:Baxter Innovations GmbH: Employment. Scheiflinger:Baxter Innovations GmbH: Employment.
The Frequency and Clinical Significance of Thrombocytopenia Complicating Critical Illness: A Systematic Review
Authors: Hui Philip LY , Cook Deborah J , Lim Wendy , Fraser Graeme , Arnold Donald M.
Published: October 2019

Abstract 4664

Background:

The epidemiology of thrombocytopenia in critically ill patients has not been well characterized. The objective of this study was to systematically review the prevalence, incidence, risk factors for, and consequences of thrombocytopenia among critically ill patients.

Methods:

We searched MEDLINE, EMBASE, the Cochrane Registry for controlled trials (until May 2010), and the Online Computer Library as well as bibliographies of relevant studies to identify investigations designed to examine the frequency, risk factors and/or outcomes associated with thrombocytopenia among patients admitted to the intensive care unit (ICU). We selected studies, abstracted data and assessed methodological quality in duplicate, independently. Heterogeneity of design and analysis precluded statistical pooling of results.

Results:

We identified 23 studies (12 prospective) enrolling 6,568 patients from medical, surgical, mixed, cardiac or trauma ICUs. Prevalent thrombocytopenia (on ICU admission) occurred in 8.3 – 67.6% of patients; incident thrombocytopenia (developing during the course of the ICU stay) occurred in 13.0 – 44.1% patients. High illness severity, organ dysfunction, sepsis and renal failure were common risk factors. Only 1 study using multivariate analysis examined whether thrombocytopenia was associated with major bleeding but found no association. Six out of 8 studies using multivariate analysis found that thrombocytopenia increased the risk of death.

Conclusion:

The frequency of thrombocytopenia during critical illness varies widely based on case mix and definition. Thrombocytopenia appears to increase the risk of death after adjustment for confounding factors. The association between thrombocytopenia and bleeding in the ICU has not been adequately examined. Although thrombocytopenia was associated with poor outcomes in most studies, randomized trials of platelet transfusions or other interventions aimed at increasing the platelet count are needed to determine whether improvement of thrombocytopenia can modify these risks.

Disclosures:

No relevant conflicts of interest to declare.
Reticulated Platelets Quantification In An Automatic CBC Counter and Its Role In Thrombocytopenia Etiology Assessment
Authors: Simoes Ana Teresa , Barreira Rogério , Oliveira Ana Cristina , Pinheiro Ana Cristina , Ferreira Patrícia , Duarte Marta , Salvado Ramón , Ribeiro Maria Leticia , Santos Susana Almeida
Published: October 2019

Abstract 4672

INTRODUCTION:

Reticulated platelets (rP) are young platelets, recently released from the bone marrow, and contain more cytoplasmic RNA elements than mature platelets. Considerable amount of evidence suggests that rP analysis may be useful in the investigation of platelet disorders. Automatic quantification methods have been developed; however, the methodology has not yet been standardized and validated in clinical practice.

STUDY PURPOSE

To analyze the rP percentage (rP%) in 168 samples, using software version v3 in Cell Dyn® Sapphire, in correlation with clinical data.

MATERIAL AND METHODS:

168 samples analyzed in Cell Dyn® Sapphire, between March and August 2010, using CBC+RET methodology in the determination of: rP%, reticulocytes % (Ret%), platelet optical counts (PLTo) and mean platelet volume (VPM). Normal control samples (C) n=73; Patients: 20 with auto-immune haemolytic anemia (IHA); 8 with thrombocytopenic thrombotic purpura (TTP); 45 with immune thrombocytopenia (IT), 4 with bone marrow aplasia (BMA) and 18 thrombocythopenias of unknown cause (TUC). Mean difference between rP% in controls vs TUC samples calculated by t-student. Statistical analysis by StatView 5.0.

RESULTS:

C samples: median rP % =1.6, range=0.2-4.1, mean PLTo= 263×109/L, median MPV=8.5fL. IHA: median rP%= 1.9, range =0.6 – 12.5, mean PLTo=284×109/L, median MPV= 8.6, mean Ret%=12. TTP: median rP%=5.1, range=0.3-16.5, mean PLTo=185×109/L, median MPV=8.5. IT: median rP%=3.8, range=0.4-27.8, mean PLTo=56.8×109/L, median MPV=11.4. BMA: median rP%=4.1, range=0.3-6, mean PLTo=66.2×109/L, median MPV=11. TUC: median rP%=2.7, range=0.2-13.6, mean PLTo= 80.5×109/L, median MPV=11.1 Mean difference between rP% in C vs TUC samples: t=2.5, p=0.02.

CONCLUSIONS:

Median rP% values obtained in control group are in agreement with published data, despite the variation in the “normal” values between the authors. MPV and rP% in destructive thrombocytopenias (IT and TTP) are higher than in control group, in accord to immature platelets release by the bone marrow. IHA patients, despite the high Ret%, have a median rP% similar to the control group, demonstrating that rP counts are not over-estimated by the high number of reticulocytes. In BMA rP% is higher than expected, however the number of samples analyzed is very low. The statistically significant mean difference between rP% in Control vs TUC samples makes the immune cause for the thrombocytopenia very likely. Due to its simplicity, ready availability and information provided by this methodology, it has a good potential to be a useful tool in the investigation of thrombocytopenias of unknown cause.

Disclosures:

No relevant conflicts of interest to declare.
Rituximab Is An Effective Therapy for Relapsed Thrombotic Thrombocytopenia Purpura
Authors: Jajeh Ahmad
Published: October 2019

Abstract 4679

Rituximab, a monoclonal antibodies designed against a CD20 glycoprotein expressing lymphocytes. This drug has been implemented in the treatment of B Cell lymphoproliferative disorders as well as other non-neoplastic diseases such as autoimmune disorders. Rituximab has produced response in auto-immune thrombocytopnea, auto-immune hemolytic anemia and refractory inhibitors to coagulation factors. This abstract present our experience with rituximab in the treatment of relapsed Thrombotic Thrombocytopenia Purpura and Hemolytic Uremic Syndrome TTP/HUS. A total of five patients diagnosed with relapsed TTP/HUS with mean age of fifty years, all are black females. Median time to relapse is 5 months, range from 3 weeks to two years. All patients were treated initialy with plasmaphoresis and steroids, average plasmaphoresis treatment before relapse is seventeen sessions, range twelve to twenty five. All patients were retreated with plasmaphoresis and steroids at relapse and all received Rituximab weekly therapy, range two doses to six. All patient achieved good response and durable remission with out relapse. Dose used is 375 mg/M2. Peripheral smears were monitored on daily bases, same as CBC, LDH, BUN and creatinin. Most interestingly that red cell fragmentation were significantly reduced after one week of starting rituximab. Fragmented red cell/schiztocytes were reduced to 3 schiztocytes per 5000 red cell on high power field light microscope. Most patient received first dose of rituxan after at least seven to nine sessions of plasmaphoresis given.

In conclusion: Rituximab is an effective therapy in relapsed TTP/HUS, larger study needed to confirm efficacy. Consideration to give Rituximab in first line therapy as well as maintenance therapy is a valid point.

Disclosures:

No relevant conflicts of interest to declare.
Patrolling Murine Monocytes Are Defined by Their Expression of the Orphan Nuclear Receptor, Nur77 (nr4a1)
Authors: Hubbeling Harper , Maltzman Jonathan , Moran Amy , Hogquist Kristin , Cunningham Nicole , Punt Jenni A.
Published: October 2019

Abstract 4723

‘Resident’ or ‘patrolling’ monocytes have recently been described as a unique subset of myeloid cells that differ in phenotype and function from classic inflammatory monocytes. Resident monocytes are distinguished by their ‘crawling’ migration on endothelial surfaces, their immunosuppressive activity, and their unique pattern of surface protein expression: Ly6Clo, CX3CR1hi, CCR2-, LFA-1+, and PDL-1+. Using a Nur77-eGFP reporter mouse, we show that all murine patrolling monocytes, but not conventional monocytes express Nur77+ (also known as NR4A1), a member of the Nur orphan nuclear receptor family that regulates cell fate decisions several cell types and inhibits leukemogenesis. Nur77 is an immediate early response gene, and we have recently determined that it is a sensitive indicator of antigen receptor signaling among T cells. We therefore propose that its expression among patrolling monocytes is an indication that these cells are actively receiving signals, perhaps via interactions with the endothelium. We are currently working to identify these signals and our preliminary data suggest that adhesion molecules play an influential role.

We thank the Arnold and Mabel Beckman Foundation (HH), and NSF (MCB-0744570 (JP and NC)) for their support for this work.

Disclosures:

No relevant conflicts of interest to declare.
Learn While You Wait: An Experience of a Novel, Interactive VHA e-Notebook Education Program for Multiple Myeloma
Authors: Yacoub Abdulraheem , Mehta Paulette
Published: October 2019

Abstract 4743

We evaluated a novel e-notebook -based education program for patients with multiple myeloma. The program was developed by the Myeloma Initiative in the Veteran's Administration (MIVA) consortium to educate patients while they wait for clinic appointments or procedures. The goals of this study were to assess feasibility of the program in a busy clinic setting, patients’ gain in knowledge from the program, patients’ acceptance of the e-notebook format, and potential use of program in other populations and in different formats. Study subjects were patients with multiple myeloma being treated at the Central Arkansas Veterans Healthcare System McClellan Hospital. Eligibility criteria were diagnosis of multiple myeloma, ability to follow instructions in the program and complete a paper questionnaire. Study subjects underwent the program while awaiting their oncology visit and then completed a survey consisting of ten multiple-choice questions designed to measure the goals of the study. The scope of the questions were: patients’ perceived benefit of the education program in improving their understanding of their disease (three questions), patients’ primary source of medical education, patients’ preferences in regard to educational formats (three questions), and multiple myeloma knowledge (three questions). Program use did not adversely affect patient flow in our busy oncology clinic, and was well accepted by the patients. Eight patients were enrolled in the study. All patients considered their oncologist to be their primary source of information about their disease. All patients stated that the education program helped them better understand the information provided by their oncologists and better understand multiple myeloma treatment options. Five patients (63%) answered that the program motivated them to learn more about myeloma. Seven patients (88%) recommended that this program be expanded to spouses or family members, and seven patients (88%) advocated creating a video format of the program for repeat viewing. Five patients (63%) stated they plan to view the content again at a later time. Of all multiple myeloma knowledge questions, 71% were answered correctly and 50% of the patients attributed their correct answers to information they learned in the education program. We conclude that this e-notebook -based multiple myeloma education program is well accepted, can be adopted in a busy clinic and has the potential for further applications. It is an effective and motivating tool for patient education in adjunct with physician efforts. It can be a productive utilization of waiting room time.

Disclosures:

No relevant conflicts of interest to declare.
IDENTIFICATION of Infrared SPECTRAL Signature of INDUCED Pluripotent STEM CELLS (iPSC)
Authors: Sandt Christophe , Feraud Olivier , Turhan Ali G. , Dumas Paul , Bennaceur-Griscelli Annelise
Published: October 2019

Abstract 4792

Recent technological advances in cell reprogramming by generation of induced pluripotent stem cells (iPSC) offer major perspectives in disease modelling and future hopes for providing novel stem cells sources in regenerative medicine. However, research on iPSC still requires refining the criteria of the pluripotency stage of these cells and exploration of their equivalent functionality to human embryonic stem cells (hESC). In this work, we report that the use of the Synchrotron-based FTIR microspectroscopy allows following the infrared spectral modification of the differentiated cells during the reprogramming process as well as the comparison between iPSC with hESC. The model that we studied consisted on the use of human ES cell line H9 grown on murine embryonic fibroblasts in the presence of bFGF. We have generated mesenchymal stem cells from the H9 cell line (H9-MSC) and we used them to generate iPSC (iPSC-H9) by the enforced expression of pluripotency genes Oct4, Sox2, Lin28 and Nanog. We have also followed the same approach on murine cells by generating murine iPSC from murine ES cells by retrovirus mediated gene transfer of Oct4, Sox2, c-Myc and Klf4. iPSC were characterized by expression of pluripotency markers, and teratoma assays. Infrared Spectral fingerprints of the original H9, MSC-H9 and iPSC-H9 as well as differentiated murine fibroblasts and murine iPSC were acquired at sub-cellular resolution using a synchrotron-powered infrared microscope. In murine system, the spectral signature of iPSC has been compared to that of D3, a well-characterized murine ES cell line. The spectral signature of iPSC and ESC displays a marked difference with those of the differentiated cells used before reprogrammation regardless the origin of the target cell (mesenchymal stem cells or murine fibroblasts). We unambiguously demonstrate for the first time to our knowledge, that the human and murine iPSC retrieve the same chemical composition with an indistinguishable spectral signature from their embryonic stem cells counterparts. Importantly, the spectral signatures were found to be specific to each of the cell line, as evidenced using pattern recognition methods and illustrated the genetic biodiversity of each iPSC and ESC. Thus, in addition to the classical pluripotency markers, FTIR microspectroscopy signature could be a rapid methodology to evaluate the pluripotency after somatic cell reprogramming.

Disclosures:

No relevant conflicts of interest to declare.
hOCT1 Gene Expression Might Predict Molecular Response In Patients with Chronic Myeloid Leukemia
Authors: Zamora Lurdes , Cabezon Marta , Boqué Concha , Marce Silvia , Ribera Jordi , Dominguez Diana , Granada Isabel , Grau Javier , Xicoy Blanca , Fernandez Cristalina , de Sevilla Alberto Fernandez , Gallardo David , Ribera Josep-Maria , Millá Fuensanta , Feliu Evarist
Published: October 2019

Abstract 4838

Introduction:

Chronic myeloid leukemia (CML) is a clonal hematopoietic malignancy characterized by the presence of BCR/ABL fusion gene. The resulting protein has a high tyrosine kinase (TK) activity. The first-line treatment for CML is Imatinib, which allow the achievement of cytogenetic and molecular response in most of patients with CML in chronic phase. However, some patients do not respond to this treatment or lose their initial response. Imatinib has been reported to be incorporated into the cell through hOCT1 transporter (human organic cation transporter).

The aim of this study was to determine whether the expression of hOCT1 at diagnosis of CML influenced the achievement of molecular response.

Patients and Methods:

We analyzed hOCT1 gene expression by quantitative PCR in 42 patients at diagnosis and 18 months after treatment with Imatinib. We compared the expression with the presence of compleat molecular response (CMR) at 18 months. We consider CMR when the Ratio (BCR-ABL/ABL)×100 was <0.1% (after International Scale correction). For statistical analysis methods we have used Kolmogorov-Smirnov and Mann-Whitney nonparametric methods.

Results:

Of the 42 patients, 2 were in hematological response, 22 were in cytogenetic response and 18 in CMR at 18 months. We found a higher hOCT1 gene expression at 18 month than at diagnosis (53.3 versus 29.6, p<0.001) in all patients (Figure 1). We have found some tendency of higher hOCT1 expression at diagnosis in patients with CMR at 18 months than in those who did not had (25.5 versus 18.8, p = 0.07) (Figure 2).

Conclusions:

Partially funded by FICJ-P-EF-09, RD06/0020/1056 de RTICC and Novartis.

We want to thank Dr. David Marin for providing us plasmid for quantitative analysis.

Disclosures:

No relevant conflicts of interest to declare.
Fas and Fas Ligand Expression In Children with Acute Lymphoblastic Leukemia, Benign Hematological Diseases and Solid Tumors
Authors: Stiakaki Eftichia , Martimianaki Georgia , Kaparou Maria , Perdikogianni Chryssoula , Kalmanti Maria
Published: October 2019

Abstract 4853

Fas/FasL is a key pathway of cellular apoptosis. Fas receptor is expressed on membranes of both normal and neoplastic cells while, Fas ligand (FasL) mainly on activated T-lymphocytes. Fas-FasL abnormalities have been detected in malignancies and autoimmune diseases and implicated in resistance to treatment. The aim of the study was the determination of the levels of Fas, FasL and their coexpression in bone marrow cells of children with acute lymphoblastic leukaemia (ALL) at diagnosis, during the course of treatment and the comparison with relevant levels in other hematological and neoplastic diseases. Expression levels of Fas and FasL were determined with flow cytometry in children with ALL at diagnosis (ALLd, n=13), on day 15 of treatment (d15, n=6), on day33 (ALLd33, n=6), during consolidation (ALLhr, n=12) and at the end of therapy (ALLet, n=7) as well as in children with Langerhans Histiocytosis (LCH, n=4), cytopenias (Cytp, n=8) and solid tumors without bone marrow infiltration at diagnosis (STd, n=5) and on therapy (STther, n=6).

Results:

The lowest levels of Fas expression were detected at diagnosis of ALL and were gradually statistical significantly increased until remission of day 33 (ALLd vs ALLd33: 8.02±1.94 vs 24.04±6.11, p=0.035). At consolidation Fas levels were found to be decreased compared to day33 (16.1±4.18) and were again increased at the end of therapy (ALLd vs ALLet: 8.02±1.94 vs 24.96±7.95, p=0.024). On the contrary, FasL levels were gradually decreased and finally increased to similar to diagnosis levels at the end of treatment (ALLd: 4.59±1.41, ALLet: 5.89±1.99). In solid tumors at diagnosis Fas levels were similar to the ones while on chemotherapy (STd vs STth:16.04±2.2 vs 15.19±6.4). The highest FasL levels were detected in the group of STd with the relevant levels on treatment being lower in comparison (STd vs STther: 10.91±3.32 vs 2.92±0.79, p=0.052). In LCH both Fas and FasL levels were found to be as low as at ALL diagnosis. In cytopenias no significant difference was observed between groups for either Fas (11.05±4.49) or FasL (3.01±0.62). As for Fas+FasL+ coexpression no difference was evident between ALLd, ALLet and STd or STther [ALLd (0.73±0.38), ALLet (0.64±0.17), STd (0.68±0.095), STther (0.66±0.38)]. The lowest coexpreesion levels were observed in the group of cytopenias with statistical significant difference compared to STd (0.68±0.095 vs 0.26±0.08, p=0.031). In conclusion, at diagnosis of ALL Fas levels were expressed in lowest levels that were found to be gradually increased at remission and at the end of treatment. This finding probably correlates with the apoptotic process of the leukemia clone and possibly with response to treatment.

Disclosures:

No relevant conflicts of interest to declare.
Biphasic Roles for the Soluble Guanylyl Cyclase (sGC) In Platelet Activation In Mice
Authors: Zhang Guoying , Xiang Binggang , Skoda Radek C. , Smyth Susan S. , Du Xiaoping , Li Zhenyu
Published: October 2019

Abstract 486

The role of intracellular secondary messenger cGMP in platelet activation has been controversial, with both stimulatory and inhibitory roles reported. The platelet cGMP is believed to be predominantly synthesized by soluble guanylyl cyclase (sGC), which is activated by nitric oxide (NO). To specifically determine the role of sGC-dependent cGMP synthesis in platelet function and in vivo thrombosis and hemostasis, we produced mice harboring a “floxed” sGC beta1 allele. In the “floxed” sGC beta1 mice (sGC beta1fl/fl), the exons 7 and 8 of sGC beta1 gene and an inserted Neo cassette were flanked with three LoxP sites. Platelet-specific deletion of sGC beta1fl/fl allele was accomplished through breeding of the sGC beta1fl/fl mice with pf4-Cre recombinase transgenic mice. Immunoblotting showed the complete absence of this protein in sGC beta1fl/fl/Cre platelets. Mice lacking sGC beta1 in platelets appeared to develop normally and had normal blood counts, including platelets. Blood pressure of platelet-specific sGC deficient mice was comparable to that of wild-type littermates. Inactivating the sGC beta1 gene in platelets abolished cGMP production induced by either NO donors or platelet agonists that are known to activate endogenous NO synthesis, confirming that both the platelet agonist-induced and NO donor-induced platelet cGMP production are predominantly mediated by sGC. Platelets lacking sGC exhibit a marked defect in aggregation and secretion in response to low doses of platelet agonists, collagen and thrombin. Importantly, tail-bleeding times were significantly prolonged in the platelet-specific sGC deficient mice compared with the wild-type littermates. In a FeCl3-induced carotid artery thrombosis model, time to occlusive thrombosis was prolonged in the platelet-specific sGC deficient mice, compared to wild type littermates. Thus, the agonist-stimulated sGC activation is important in promoting platelet granule secretion and aggregation. On the other hand, NO donor SNP-induced inhibition of platelet activation was abolished in sGC-deficient platelets. However, at high concentrations (>100μM), SNP inhibited platelet activation in both wild type and sGC deficient mice, indicating that both cGMP-dependent and -independent mechanisms are involved in NO donor-induced inhibition of platelet activation. Together, our data demonstrate that sGC contributes to both agonist-induced platelet activation and NO donor-induced platelet inhibition.

Disclosures:

No relevant conflicts of interest to declare.
Immunodulatory Capacity of Mesenquimal STROMAL CELLS IS CONTROLLED by NON-Canonical NF-Kb PATHWAY
Authors: Haddad Rodrigo , Saldanha-araujo Felipe , Araujo Amélia , Covas Dimas T , Zago Marco A. , Panepucci Rodrigo Alexandre
Published: October 2019

Abstract 4865

Introduction:

Mesenquimal Stromal Cells (MSCs) possess immunosuppressive properties, becoming these cells a promising subject of study for future approaches in cell-based therapy. During co-culture with activated lymphocytes and probably through T-cell derived cytokines, MSCs are activated in order to become suppressive. Once MSCs are triggered, they acquire an inhibitory profile increasing the release of immunoregulatory factors, such as IDO, IFNγ, PGE2, NO and TGF-β, responsible for T-cell inhibition. In addition, expression of adhesion molecules, such as ICAM-1, and generation of regulatory T cells subsets, such as CD69+ T cells, are important in immunosuppressive property of human MSCs. However, the exact mechanisms underlying the immunomodulatory functions of MSCs remain largely unknown. NF-kB comprises a family of inducible transcription factors that serve as important regulators of the host immune and inflammatory responses. The NF-kB signals are activated via canonical (mediated mainly by RelA-p50) and/or non-canonical (mediated by RelB-p52) pathways in response to diverse stimuli. Given the immunomodulatory properties of MSCs and the possible involvement of NF-Bk pathway in this effect, here, it was explored the role of non-canonical NF-KB signaling in the immunomodulatory capacity of MSCs cocultured with activated T cells.

Methods:

siRNAs targeting RelB were transfected into MSC with lipofectamine. siRNAs C- were used as negative control and siRNA-FITC as transfection control. After 24 hours, immunomagnetically purified CD3+ T cells were stained with CFSE, activated by anti-CD2/CD3/CD28 beads and cultured in the presence of MSCs. After 3 days, flow cytometry was performed to observe the transfection efficiency, T cell proliferation and percentage of CD69+ T regulatory cells. Additionally, RNA from MSCs was extracted and RelB and ICAM-1 mRNA expression were quantified by Real-time PCR.

Results:

The transfection efficiency was around 75% and RelB mRNA level was reduced by 80% in MSCs transfected with siRNA RelB compared to siRNAs C- transfected cells. Compared to MSCs previously transfected with siRNA C- and co-cultured with activated T cells, MSCs transfected with siRNA RelB resulted increasing of 22% in T cell proliferation and decreasing of 9,2% and 30% in the CD69+ regulatory T cells generation and ICAM-1 expression respectively.

Conclusion:

Non-canonical NF-kB pathway, mediated by RelB, may be partially involved in acquisition of the inhibitory profile by decreasing T cells proliferation, and increasing the expression of ICAM-1 and the generation of CD69+ regulatory T cells.

Disclosures:

No relevant conflicts of interest to declare.
Comparison Between CD20-Negative and CD20-Positive Diffuse Large B Cell Lymphoma; Characteristics and Clinical Outcome
Authors: Jo Jungmin , Yoo Changhoon , Ahn Yongchel , Park Seong Joon , Kim Shin , Lee Dae Ho , Kim Sang We , Lee Jungsin , Suh Cheolwon
Published: October 2019

Abstract 4891

Background

CD20 is a non-glycosylated phosphoprotein expressed on the surface of all mature B-cells. CD20 is expressed on all stages of B cell development except the first and last. However, complete lack of CD20 expression occurred in a few cases without previous rituximab (R-) treatment. The immunohistochemostry (IHC) studies which we used were not perfect for confirmation of expression. However, we intended to investigate characteristics and clinical outcome of CD20-negative diffuse large B-cell lymphoma (DLBCL), not detected with usual method, and to compare with CD20-positive.

Methods

The records of Non-Hodgkin's Lymphoma patient registry were reviewed in Asan Medical Center. Between September 2003 and February 2009, a total of 407 patients were diagnosed DLBCL and 16 patients (3.9%) out of 407 confirmed CD20-negative DLBCL by IHC. The rest of patients (n=391) were CD20-positive and unconfirmed cases were excluded. Retrospective analysis of complete response (CR), disease-free survival (DFS), and overall survival (OS) was performed.

Results

The median age was 60.5 years old (range 31–81) in CD20-negative patients. Ten patients were males. The Ann Arbor stage was I in 3 patients, II in 3 patients, and III or IV in 10. Six patients were low risk group, 7 patients in intermediate, and 3 in high risk group according to international prognostic index (IPI). Most of patients (62.5%) received cyclophosphamide, doxorubicin, vincristin, and prednisone (CHOP) chemotherapy in CD20-negative and 295 patients (75.4%) with R-CHOP in CD20-positive DLBCL. The Baseline characteristics was not different in both groups except Hans classifier (p=0.02).

With a median follow-up time of 32.3 months (range 0.5–83.4), the CR rate was 73.5%, the 3-year OS was 69.5%, and 3-year DFS 74.2% in all patients. CD20-positive and CD20 negative groups had a CR rate of 73.7%, 68.8% (p=0.146), respectively, a 3-year OS of 70.7%, 40.0% (p=0.003), a 3-year DFS of 75.4%, 44.6% (p< 0.001), respectively. The 3-year OS and DFS also had significant difference with adjusted IPI (p<0.001, respectively).

Conclusions

CD20-negaive DLBLC was not infrequently with usual IHC method (around 4%). The survival outcome was poor compared with CD20-positive DLBLC because of high relapse rate. It was caused without rituximab treatment in CD20-negative. Development of novel target agents like rituximab should be explored to improve outcome and maintain the CR status of CD20-negative DLBCL.

Disclosures:

No relevant conflicts of interest to declare.
A Refractory Stage IVb Thymoma Responded Completely to a Combination of Dasatinib and Prednisone but Poorly to Either One Alone
Authors: Lian Eric C. Y. , Matsangou Maria , Liu Diana
Published: October 2019

Abstract 4925

Introduction:

Thymomas are rare tumors for which there is little randomized evidence to guide treatment and therefore, therapeutic options to cure advanced malignant thymomas are limited. We hereby report the first clinical outcome of combined dasatinib and prednisone therapy in a patient with heavily treated stage IVB malignant thymoma.

Case Presentation:

A 41 year old white male presented with fatigue, fever and granulocytopenia. He soon developed myasthenia gravis and aplastic anemia. Computer tomography showed a mediastinal mass, pleural thickening, splenomegaly and mesenteric nodes; biopsy of mediastinal mass revealed a malignant thymoma. Prednisone for 6 weeks along with antithymocyte globulin and cyclosporine were started for myasthenia gravis and aplastic anemia. Chemotherapy for unresectable thymoma was soon initiated with ADOC (Cisplatin, Doxorubicin, Vincristine, and Cyclophosphamide), but only small partial response was noted. After failing to show response to another three lines of therapy, we commenced therapy with dasatinib 75 mg twice daily. Thymoma showed partial response to dasatinib initially but soon became resistant and progressed. When prednisone 100 mg daily was added, a complete response was observed.

Conclusion:

A patient with stage IVB thymic carcinoma failed to respond well 3 lines of combination chemotherapy, and neither to prednisone or dasatinib alone. When dasatinib is combined with prednisone, it induced a complete response.

Disclosures:

Off Label Use: Dasatinib is an oral dual BCR/ABL and Src family tyrosine kinase inhibitor. It is a potent inhibitor of Imatinib resistant KIT activation loop mutants and induces apoptosis in mast cells and leukemic cells. At higher concentrations, Dasatinib also inhibits c-Kit, PDGFR and EphedrinA2. it has currently FDA approval for treatment of adults in all phases of chronic myeloid leukemias with resistance or intolerance to prior therapy, including Gleevec. The second indication for Dasatinib, is for the treatment of Ph+ALL. A number of Phase II trials have shown activity of this agent to various solid tumors including breast and lung cancer. Imatinib showed activity in thymomas, thus the rationale for using Dasatinib in the treatment of our patient.
Effects of 5-Bromotetrandrine and Daunorubicin on the Expression of Survivin In K562/AO2 Cells
Authors: Chen Bao-An , Cai Xiao-hui , Wang Jun , Gao Chong , Ding Jia-hua , Cheng Jian , Sun Yun-yu , Song Hui-hui , Bao Wen , Chen Ping-shen , Zhang Yu , Tang Meng
Published: October 2019

Abstract 4947

Objective:

The aim of this study was to investigate the expression of survivin and the apoptosis induced by DNR and BrTet in the leukemic cells K562/A02.

Methods:

In a typical experiment, the K562/AO2 cells were treated with daunorubicin (DNR), 5-bromotetrandrine (BrTet), or DNR and BrTet for 48 hours, and the cells treated without any drugs were used as control group. Cell proliferation was analyzed by MTT assay. Cells apoptosis and the concentration of DNR within the cells were measured by Flow cytometry (FCM). The expressions of mRNA and protein of survivin were determined by semi-quantitative reverse transcription PCR (RT-PCR) and Western blot, respectively.

Results:

The results of MTT assay indicated that DNR and BrTet were both able to inhibit the proliferation of K562/AO2 cells in dose-dependent manner. The fresh evidence from flow cytometry showed that a higher apoptosis rate could be induced and a higer concentration of DNR could be detected in K562/AO2 cells by DNR and BrTet as compared with those by DNR or BrTet in the same concentrations(P<0.01). RT-PCR revealed that the expression of survivin mRNA, a higer expression in K562/AO2 cells with acquired resistance to adriamycin than that in parent K-562 cells, decreased in the DNR and BrTet group (P<0.05), but there was no obvious change in other groups(P>0.05). Western bolt demonstrated that the expression of survivin protein was much lower in the DNR and BrTet group(P<0.05).

Conclusion:

BrTet could increase the concentration of DNR and reverse the multidrug resistance(MDR) in the K562/AO2 cells. Survivin may play an important role in apoptosis induced by DNR. Survivin could be a target for the treatment of MDR in haematopoietic malignancies.

Disclosures:

No relevant conflicts of interest to declare.
Reduction of Imatinib Concentration After Gastric Bypass Surgery
Authors: Liu Hongtao , Artz Andrew S
Published: October 2019

Abstract 4948

Imatinib mesylate is an orally bioavailable inhibitor of several protein tyrosine kinases, including BCR-ABL, which was approved for BCR-ABL–positive CML. This oral agent has approximately 98% oral bioavailability independent of a meal, and it is eliminated by biliary excretion and hepatically metabolized. Better clinical outcomes have been associated with higher steady state imatinib trough levels for both CML and GIST. Because imatinib is absorbed, excreted, and possibly recycled in the gastrointestinal tract, gastrointestinal disorders impairing absorption may hinder achieving therapeutically effective imatinib levels. We present the case of a woman with CML who underwent biliopancreatic diversion with duodenal switch for weight loss leading to a marked reduction in imatinib concentration, and as a consequence, inadequate suppression of her CML. This is a 54 year old obese African American woman who achieved complete hematological remission (CHR), and complete cytogenetics remission (CCyR) and good molecule response with a BCR-ABL/c-ABL ratio of 0.015 (1.5%) with close to therapeutic imatinib trough level at 965 ng/mL prior to gastric bypass surgery on imatinib 400mg daily. The patient underwent laparoscopic biliopancreatic diversion with duodenal switch, concomitant cholecystectomy, and appendectomy. Her adjusted imatinib steady state trough level declined to 166ng/mL, 17% of the pre-surgery level and below the concentration required to suppress BCR-ABL activity leading to a 0.5 log increase in BCR-ABL/c-ABL ratio of 0.083 (8.3%). Escalation of imatinib to 400 mg BID led to imatinib trough level of 734 ng/mL and decrease of BCR-ABL/c-ABL ratio to 0.024 (2.4%), a value comparable to the ratio achieved while on imatinib 400mg daily prior to the gastric bypass surgery. Eventually, our patient lost almost 50kg (close to 45% of her original weight) within a year period. Interestingly, the imatinib level increased to 2124 ng/mL on imatinib 400mg twice a day after the substantial weight loss, supporting the correlation of imatinib level and actual body weight. Her PCR BCR-ABL/c-ABL from peripheral blood still revealed a ratio of 0.072 (7.2%) with high therapeutic imatinib level, and a secondary generation TKI, nilotinib, was initiated at 400mg twice a day. The detail of the clinical course was listed in table below.

Disclosures:

No relevant conflicts of interest to declare.
Biologic Features and Clinical Outcomes According to Age Groups In Myelodysplastic Syndrome
Authors: Lee Seungbum , Lee Je-Hwan , Lee Jung-Hee , Kim Dae-Young , Kim Sung-Doo , Lim Sung-Nam , Choi Yunsuk , Park Young-Hoon , Seol Miee , Lee Young-Shin , Kang Young-Ah , Jeon Mijin , Lee Kyoo-Hyung
Published: October 2019

Abstract 4963

Myelodysplastic syndrome (MDS) is a disease of the elderly, but can also affect younger people. Age is known to be an important prognostic factor in MDS but age variable is not included in most prognostic scoring systems because it is not thought as a disease-related variable. Many reports have showed that MDS is seen one to two decades earlier in Far Eastern countries than Western countries. We retrospectively investigated the differences in biologic features and clinical outcomes according to different age groups in Korean patients with MDS. Primary end points of our study were overall survival (OS) and progression-free survival (PFS). PFS was defined as time from diagnosis to AML progression or death. About one third of the patients received intensive treatment including chemotherapy, hypomethylating treatment or hematopoietic cell transplantation. Therefore, all survival data were censored at the start of intensive treatment to eliminate the influence of the treatments on clinical outcomes. A total of 403 patients, 248 males and 155 females, were included in this study. Median age was 54 years. We divided the patients into three age groups: ≤50 years (n=181), 51 to 60 (n=81), and over 60 (n=141). Baseline biologic features were significantly different according to three age groups: with increasing age, more male preponderance (P=0.009), more BM blast percentage (P<0.001), more advanced WHO subtype (P<0.001), higher proportion of high risk cytogenetic features (P=0.052; ≤60 vs. >60, P=0.011), poorer ECOG performance status (P=0.004), higher IPSS risk group (P=0.019). Five-year survival probabilities were significantly different according to age groups (≤ 50 vs. 51–60 vs. > 60; OS, 66.8% vs. 28.5% vs. 12.2%, P<0001; PFS, 58.5% vs. 37.9% vs. 12.3%, P<0.001). Survivals were also significantly different according to age groups in both IPSS Low/INT-1 (P<0.001 for OS, P=0.001 for PFS) and IPSS INT-2/High risk group (P=0.026 for OS, P=0.069 for PFS). Cox proportional hazards models also demonstrated that age group was an independent prognostic factor for survivals: ≤ 50 vs. 51–60 and > 60; OS, RR 2.3 (P=0.037) and RR 4.6 (P<0.001); PFS, RR 1.3 (P=0.449) and RR 2.0 (P=0.012).

Conclusion:

Biologic features and clinical outcomes were significantly different among age groups in MDS. Clinical outcomes were better in younger age group independently of biologic features. Survivals (OS & PFS) were better in younger age group and survival differences by age groups were observed in both lower and higher risk MDS, suggesting that age stratification should be considered in treatment decision and clinical trial design.

Disclosures:

No relevant conflicts of interest to declare.
Primary Immune Thrombocytopenia (ITP) Guidelines for Bone Marrow Aspirate Reduce the Diagnostic Rate of Myelodysplastic Syndromes
Authors: Vincelli Iolanda , Oliva Esther Natalie , Ronco Francesca , Cufari Patrizia , Oliva Bianca , Gatto Valentina , Laganà Carmelo , D'Errigo Maria Grazia , Nobile Francesco
Published: October 2019

Abstract 4974

Introduction:

Primary immune thrombocytopenia (ITP) is an acquired immune-mediated disorder characterized by isolated thrombocytopenia, defined as a peripheral blood PLT count <100 Gi/L, and the absence of any obvious initiating and/or underlying cause of the thrombocytopenia (Rodeghiero et al, Blood, 2009). Guidelines consider bone marrow (BM) aspirate informative in patients with thrombocytopenia > 60 years of age, those with systemic symptoms or abnormal signs or in some cases where splenectomy is considered (Provan et al, Blood, 2009).

Methods:

We performed a retrospective chart analysis between January 2008 and June 2010 of all patients referred to our clinic for isolated thrombocytopenia but with a PLT count 100– 149 Gi/L. According to recent guidelines, these patients are not to be considered thrombocytopenic and do not require further investigation. The aim of the study was to evaluate the validity of omitting BM analysis in these cases.

Results:

Twenty-three cases (13 males/10 females) of mean age 58 ± SD 19 years were evaluated at our clinic for a PLT count below normal lab range values. At the time of evaluation none had bleeding symptoms. PLT counts ranged from 101 to 149 Gi/L, mean 123 Gi/L. Four patients had an enlarged spleen. After initial screening, 2 patients had a complex autoimmune disorder and 1 case had HCV hepatitis. The remaining 20 patients had a bone marrow (BM) aspirate performed: a diagnosis of myelodysplastic syndrome (MDS, WHO classification refractory thrombocytopenia) was obtained in 13 cases (65%) and BM biopsy was performed in 12, completed by cytogenetics in 9 cases (7 normal, 1 del20q, 1 –Y). Patients diagnosed with MDS were significantly older (66 ± SD 13 vs 47 ± SD 21 years, p = 0.017), but 4 cases (31%) were < 60 years of age (44, 49, 51 and 55 years of age, respectively).

Conclusions:

The most recent guidelines, which lower the PLT threshold to 100 Gi/L from 150 Gi/L for the investigation of causes of thrombocytopenia, reduce the diagnostic rate of MDS. In our retrospective review, 65% of patients would not have had an early diagnosis of MDS. Furthermore, BM aspirate should be considered irrespective of age, since one third of the patients in our case review had MDS with PLT > 100 Gi/L as a single cytopenia and age under 60.

Disclosures:

No relevant conflicts of interest to declare.
Analysis of Related Factors of Malignant Clone Burden of Patients with Myelodysplastic Syndromes
Authors: Shao Zonghong , Yue Lanzhu , Fu Rong , Wang Huaquan , Li Lijuan , Ruan Erbao , Qu Wen , Liang Yong , Wang Guojin , Wang Xiaoming , Liu Hong , Song Jia , Wu Yuhong , Xing Limin , Wang Jun , Guan Jing
Published: October 2019

Abstract 4979

Objective

To investigate the related factors of the malignant clone burden of patients with Myelodysplastic syndromes (MDS), and explore the risk factors of cytogenetic evolution of MDS.

Methods

Seventy-three cases of MDS patients were enrolled in this study who received treatment in the hematological department of General Hospital of Tianjin Medical University from 2004 to 2009. Statistics methods such as t-test, Chi-square test and Logistic regression analysis were used to investigate the correlation between the malignant clone burden and its related factors such as bone marrow dysplasia, peripheral blood and iron metabolism indexes. Furthermore, the risk factors of cytogenetic evolution of MDS were also analyzed.

Results

Odd number-nucleus erythrocytes, double-nucleus granulocytes, hypolobated neutrophils were significantly correlated to high malignant clone burden (P<0.05). PAS positive patients and patients with dysplasia in three myeloid lineages, blasts in peripheral blood exhibited significantly higher malignant clone burden (P<0.05). Patients with abnormal karyotype presented significantly higher level of serum ferritin and lower level of unsaturated iron binding capacity (UIBC) than those with normal karyotype (P<0.05). Odd number-nucleus erythrocytes, megaloblastic granulocytes and high myeloid differentiation index (DI) are risk factors indicating cytogenetic evolution of MDS patients.

Conclusion

The indexes of high malignant clone burden of MDS include: odd number-nucleus erythrocytes, double-nucleus granulocytes, hypolobated neutrophils, dysplasia in three myeloid lineages, blasts in peripheral blood, positive PAS, high level of serum ferritin and low level of UIBC. The risk factors of cytogenetic evolution of MDS include: odd number-nucleus erythrocytes, megaloblastic granulocytes and high myeloid DI.

Disclosures:

No relevant conflicts of interest to declare.