bluuebunny/crossref_metadata_2025_split huggingface dataset:
Management of Perioperative Anticoagulation in Patients with VWD
Authors: Von Depka Mario , Detering Carsten , Döpke Stefanie , Ekhlasi-Hundrieser Mahnaz 
Published: October 2019

Abstract

Objectives: von Willebrand disease (VWD) is the most common hereditary bleeding disorder. This study reviews the management of perioperative anticoagulation in patients with VWD undergoing surgical procedures. Risk factors for VTE with von Willebrand factor (VWF) concentrate use are older age, previous VTE, obesity, surgery, hormone replacement therapy use, antifibrinolytic therapy use and high post infusion FVIII levels. Currently, there are few data from randomized clinical trials assessing efficacy and possible complications of perioperative VTE prophylaxis in VWD patients.
Methods: A total number of 116 surgeries were performed (minor: n=64 and major: n=52) in this retrospective, single-centre study. They were divided into groups of perioperative non-anticoagulation (n=54) and perioperative anticoagulation (n=62), who all received coagulation factor concentrate (CFC). Sub-analyses were done according to the type of concentrate used. Anticoagulation was performed using different low molecular weight heparins (LMWH) according to standard protocols or body-weight adapted doses in patients with either elevated body-mass-index or additional thrombosis risk factors. Blood samples had been collected pre- and post-surgery (up to 21 days) to analyse PT, aPTT, PFA, and trough levels of FVIII coagulant activity (FVIII:C), VWF activity (VWF:GPIbM) and antigen (VWF:Ag), respectively. Furthermore, the median doses of CFC/kg and the median total number of infusions were calculated. The rates of clinically overt thrombosis as well as bleeding were assessed during the post-operative phase.
Results: The majority of patients suffered from VWD type 1 (104), 9 patients with type 2A, 2 with type 2M and 1 with type 3 VWD. Humate-P (H) was used in 55 patients and 61 patients received Wilate (W). Using W, we found parallel curves for FVIII:C, VWF-antigen and VWF:GPIbM, respectively. Using H, less concordance between VWF:Ag and VWF:GPIbM was visible and FVIII:C tends to increase between D3 to D10 in spite of decreasing VWF:Ag and VWF:GPIbM. This observation was visible in minor as well as major surgical procedures. LMWH (Enoxaparin, Nadroparin and Certaparin) were used in doses between 30 and 100 mg/injection (mean 46.0 ± 18.5 mg/injection) and a mean of 32.2 ± 24.6 injections in total (range: 8-112). In one patient a significant haematoma occurred (1/116; 0.9%), also one thrombotic event was documented in a different patient (1/116; 0.9%).
Conclusion: Using standard dose LMWH in patients with no overt increased thrombosis risk as well as body-weight-adapted LMWH in high risk patients seem to be safe and effective in VWD patients receiving coagulation factor concentrate perioperatively. However, prospective randomized comparative studies are required to determine the optimal indication as well as type of anticoagulation according to the CFC treatment regimen in this setting.

Disclosures
No relevant conflicts of interest to declare.
Efficacy and Safety of Weight-Adjusted Extended Duration Tinzaparin for Prevention of Post-Operative Venous Thromboembolism after Bariatric Surgery
Authors: Siegal Deborah M. , Handa Pankaj , Kolesar Emma , Anvari Mehran , Tiboni Maria , Douketis James D. , Crowther Mark A. 
Published: October 2019

Abstract
Background: Patients having bariatric surgery are at moderate-to-high risk for post-operative venous thromboembolism (VTE), with pulmonary embolism representing the most common cause of post-operative death. The optimal dosing and duration of anticoagulant prophylaxis is uncertain.
Aim: To evaluate the efficacy and safety of extended-duration, weight-adjusted tinzaparin for post-operative thromboprophylaxis after bariatric surgery.
Methods: Retrospective cohort of 793 bariatric surgery patients who received routine post-operative weight-adjusted tinzaparin, 4,500-14,000 IU daily (75 IU/kg, rounded to the nearest pre-filled syringe) for 7 days after hospital discharge. The primary efficacy and safety outcomes were the frequency of VTE and major bleeding, respectively, within 30 days of surgery in patients receiving at least 1 dose of tinzaparin.
Results: 793 patients who underwent bariatric surgery from 2009 to 2012 were reviewed. There were 44 (5.5%) patients excluded from analyses: need for therapeutic-dose anticoagulation (n=16); no post-operative tinzaparin (n=27); surgery aborted (n=1). There were 646 and 645 patients included in the 30-day efficacy and safety analyses, respectively (mean age, 44.6 years [SD 9.7], median body mass index, 47.1 kg/m2 [range: 19.4-81.0]). An additional 106 patients had outcome data only during hospital admission. VTE occurred in 1/645 patients (0.3%; pulmonary embolism) after hospital discharge and in 1/752 patients (0.1%; superior mesenteric vein thrombosis) in hospital. Major bleeding occurred in 12/645 patients (1.9%). Trough anti-Xa levels measured 7-10 days post-operatively were undetectable in 143/190 (75.3%) patients. In the other 47 patients the median trough anti-Xa level was 0.12 IU (range: 0.10-0.41).
Conclusions: Weight-adjusted, extended-duration tinzaparin appears effective and safe for post-operative thromboprophylaxis after bariatric surgery. There was no drug accumulation with large doses of tinzaparin.

Disclosures
Siegal: Interactive Forums Inc.: Other: created educational slides; Daiichi Sankyo: Other: participated in an advisory board; Boerhinger Ingelheim: Other: participated in an advisory board; Portola Pharmaceuticals: Other: participated in an advisory board. Douketis:Bayer: Consultancy; Actelion: Consultancy; Biotie: Other: Advisory board; Sanofi-Aventis: Honoraria; The Medicines Company: Other: Advisory board; Janssen: Consultancy; Daiichi-Sankyo: Consultancy; Bristol-Myers Squibb: Consultancy, Honoraria; Pfizer: Honoraria; Boehringer Ingelheim: Consultancy, Honoraria.
Plasma Levels of Human Neutrophil Peptides and Complement Activation Markers in Patients with Acquired Autoimmune Thrombotic Thrombocytopenic Purpura
Authors: Cao Wenjing , Pham Huy Phu , Williams Lawrence A. , Ping Zheng , Lorenz Robin G. , Marques Marisa B. , Zheng X. Long 
Published: October 2019

Abstract
Acquired thrombotic thrombocytopenic purpura (TTP) is a potentially fatal syndrome, resulting from autoantibodies against the metalloprotease ADAMTS13. Autoantibodies bind and inhibit plasma ADAMTS13 activity, leading to exaggerated platelet agglutination and thrombus formation in small arteries and capillaries. However, plasma ADAMTS13 deficiency or the presence of anti-ADAMTS13 autoantibodies is not sufficient to cause the disease. Infection or inflammation often precedes the acute onset of TTP. We hypothesize that neutrophil activation or complement activation vial alternative pathway may be the trigger for TTP. In this study, plasma samples were collected prior to plasma exchange from 58 adult patients with acute TTP between 2006 and 2015 at the University of Alabama at Birmingham Medical Center. All patients were diagnosed having TTP on the basis of severe thrombocytopenia, microangiopathic hemolytic anemia, and severe deficiency of plasma ADAMTS13 activity (less than 10%) and the presence of ADAMTS13 inhibitors. All patients were treated by plasma exchange. Thirty plasma samples from healthy individuals were collected for controls. Plasma levels of human neutrophil peptides (HNP-1, -2, and -3) released from activated neutrophils and alternative pathway complement activation markers (iC3b, C4d, C5b-9, and Bb) were determined by enzyme-linked immunosorbent assays (ELISAs). We showed that plasma levels of HNP-1, -2, and -3 in TTP patients were elevated on average by 10-fold when compared with those in the healthy controls (44.12 ± 39.53 ng/ml vs. 3.54 ± 2.97 ng/ml, means ± SD) (p <0.0001). Whereas plasma levels of Bb and iC3b were only modestly increased in TTP patients (2.94 ± 1.82 µg/ml vs. 1.39 ± 0.34 µg/ml for Bb, p <0.0001, and 16.21 ± 9.71 µg/ml vs. 12.13 ± 3.07 µg/ml for iC3b, p =0.028). No significant difference was detected in the plasma levels of C4d and C5b-9 between TTP patients and healthy controls (p >0.05). To differentiate HNP-1 from HNP-2 or HNP-3, we developed a LC/MS mass spectrometric assay and showed that all subtypes of HNPs in TTP patients were significantly elevated when compared with those in the healthy controls (p<0.001). In general, there was good concordance between ELISA and LC/MS MS in the measurement of all three HNPs. These results demonstrate that innate immunity (i.e. activation of neutrophils) and to a lesser degree the activation of complements via alternative pathway may play a role in pathogenesis of TTP in light of severe ADAMTS13 deficiency. Our findings provide rationales for developing novel targeted therapies for acquired TTP.

Disclosures
No relevant conflicts of interest to declare.
NIPA As a Novel Regulator of Aging and Stress Response of the Primitive HSC Pool
Authors: Kreutmair Stefanie , Istvanffy Rouzanna , Klingeberg Cathrin , Dierks Christine , Peschel Christian , Oostendorp Robert A.J. , Duyster Justus , Illert Anna Lena 
Published: October 2019

Abstract
Accumulation of DNA damage in hematopoietic stem cells (HSCs) is associated with aging, bone marrow failure and development of hematological malignancies. Although HSCs numerically expand with age, their functional activity declines over time and the protection mechanism from DNA damage accumulation remains to be elucidated.
NIPA (Nuclear Interaction Partner of ALK) is highly expressed in hematopoietic stem and progenitor cells, especially in the most primitive long-term repopulating HSCs (CD34-Flt3-Lin-Sca1+cKit+). Loss of NIPA leads to a significant exhaustion of primitive hematopoietic cells, where Lin-Sca1+cKit+ (LSK) cells were reduced to 40% of wildtype (wt) littermates (p<0.001). All LSK-subgroups, LT-HSCs (p<0.001), ST-HSCs (CD34+Flt3-LSK; p<0.01) and MPPs (CD34+Flt3+LSK; p<0.05) of NIPA deficient animals are affected and failed to age-related increase, whereas the lineage differentiation of Nipako/ko progenitor cells showed no gross differences. Myeloid depression by 5-FU treatment led to severely reduced HSC self renewal in Nipako/ko mice independent of age (p<0.001). Moreover, weekly 5-FU activation showed reduced survival of Nipako/ko vs. wt animals (11 vs. 14.5 days).
To further examine the role of NIPA in HSC maintenance and exhaustion, we performed in vivo repopulationexperiments, where Nipa deletion causes bone marrow failure in case of competition, as Nipako/ko cells contributed to less than 10% of transplanted BM cells 6 month after transplantation (TX). According to this, colony formation assays and limiting dilution transplantation showed a dramatic reduction of competitive repopulation units (p<0.0001) in Nipako/ko animals. Serial LSK transplantation assays revealed loss of Nipa-deficient LSKs shortly after TX, whereas long-term repopulation capacity seemed to be maintained, suggesting a role of NIPA in critical stress response.
To further investigate the stress response in Nipa-deficient HSCs, we irradiated LSKs with 3 Gy and stained for DNA-Damage foci by pH2ax. Remarkably, loss of NIPA led to significant higher numbers of pH2ax foci in irradiated HSCs (46% > 6 foci vs. 17% > 6 foci in wt cells) and highly increased the rates of apoptotic cells especially in the primitive CD34-LSK population.
Taken together our results highlight the importance of the DNA damage response at HSC level for lifelong hematopoiesis and establish NIPA as a novel regulator of aging and stress response of the primitive HSC pool.

Disclosures
No relevant conflicts of interest to declare.
A Human Bone Marrow-Derived Stromal Cell Population with Hemogenic Potential
Authors: Mokhtari Saloomeh , Colletti Evan Joseph , Sanada Chad , Lamar Zanetta S. , Simmons Paul J , Atala Anthony , Krause Diane S , Zanjani Esmail D. , Porada Christopher D , Almeida-Porada Graca 
Published: October 2019

Abstract
During ontogeny, definitive hematopoietic stem/progenitor cells (HSC) are thought to arise from vascular endothelial cells, through an endothelial-to-hematopoietic transition, a natural process that occurs in unique, specialized embryonic hemogenic endothelial cells. Developmental studies, and experiments using pluripotent stem cells in an effort to recapitulate this process and thereby gain a better understanding of the emergence of definitive hematopoiesis, have collectively led to the prevailing view that the hemogenic endothelium constitutes a transient population of cells within the embryo that rapidly disappears during development and is absent in the adult. Herein, we provide the first evidence that at early time points of gestation, prior to the establishment of hematopoiesis, a unique subpopulation of Stro-1+ cells present within the inner part of the developing human bone marrow co-expresses APLNR, a marker of angiogenic mesoderm. Moreover, these Stro-1+APLNR+ cells express multiple other markers described for hemogenic endothelium, and subsequently contribute to the vasculature, cartilage, and bone. Importantly, we also show that cells expressing these same markers of primitive mesoderm/hemogenic endothelium persist at low frequency within the adult marrow. These adult-derived cells can be extensively expanded in vitro without loss of potential, but lack hematopoietic colony-forming potential in vitro. However, upon transplantation into a fetal microenvironment, clonally-derived populations of these adult Stro1+ isolated stromal progenitors (SIPs) not only contribute to the vasculature and nascent BM niches, but also efficiently generate, at a clonal level, hematopoietic stem cells (HSC) that are capable of robust, multilineage hematopoietic reconstitution, with generation of both myeloid and lymphoid cells upon serial transplantation. In conclusion, our studies have thus uncovered the latent potential of a highly expandable population of seemingly vestigial adult human somatic cells, whose ontogenic history includes a phenotype identical to that described for hemogenic endothelium. We have also shown that, if provided with the appropriate/necessary inductive factors, these unique adult cells are capable of giving rise to hematopoietic cells that fulfill the gold standard criteria for bona fide HSC. Therefore, these cells could potentially be more amenable to reprogramming technologies, to produce HSC that could be used to treat/cure a broad variety of blood diseases.

Disclosures
No relevant conflicts of interest to declare.
The Role of Tetraspanin CD37 in B-Cell Malignancy
Authors: Beckwith Kyle A. , Williams Erich I. , Wright Mark D. , van Spriel Annemiek B. , Byrd John C. , Muthusamy Natarajan 
Published: October 2019

Abstract
Members of the tetraspanin superfamily of transmembrane proteins have roles in a variety of processes including cellular adhesion, migration, activation, proliferation, survival, and solid tumor metastasis. Although numerous tetraspanin proteins are expressed by immune cells, only a few have expression that is restricted to these cells. CD37 is not found in non-hematologic tissues and is highly expressed by mature B-cells, with greater than 15 times higher expression compared to other immune cells (Deckert et al., 2013). Recently there has been significant interest in CD37 as a therapeutic target in non-Hodgkin lymphoma and Chronic Lymphocytic Leukemia (CLL), as evidenced by the ongoing clinical development of four different anti-CD37 therapies. However, very little is known about the biological function of this protein. Ligation by the CD37-targeting peptide otlertuzumab leads to induction of both pro-survival Akt activation and pro-apoptotic SHP1 signaling, although cellular death is heavily favored (Lapalombella et al., 2012). Furthermore, CD37 is critical to normal plasma cell development, clustering VLA-4 (α4β1 integrin) on the surface of germinal center B-cells to allow integrin-mediated activation of Akt (van Spriel et al., 2012). In CLL, expression of the VLA-4 subunit CD49d is associated with more aggressive disease and promotes homing and adhesion to supportive cells within lymphoid tissues. This prompted us to explore the role of CD37 expression in B-cell malignancy using a CD37-deficient Eµ-TCL1 mouse model of CLL. For these studies, we monitored mice that were CD37 wild type, heterozygous knockout, or homozygous knockout (all hemizygous for the TCL1 transgene). While no significant difference was observed in the time until appearance of leukemia in peripheral blood, loss of CD37 led to decreased survival of TCL1 mice (p=0.032). In the same timeframe, non-TCL1 mice with CD37 deficiency did not exhibit impaired survival. While CD37 is predominantly seen in B-cells, the protein serves a role in other immune cells despite substantially lower expression. Our results are consistent with the recent reports on impaired dendritic cell migration in CD37 deficient mice, which is associated with decreased anti-tumor immunity in the context of a subcutaneously injected mouse cell line expressing a foreign human antigen (Gartlan et al., 2013). Our data support a potential role for CD37 in anti-tumor immune response against the syngeneic leukemia that arises spontaneously in the TCL1 model of CLL. Given the complex nature of this global knockout model, other functions of CD37 may be obscured. Ongoing studies will examine engraftment of CD37-deficient or wildtype TCL1 leukemia into healthy mice to elucidate whether the B-cell intrinsic role of CD37 is supportive of the leukemia, independent of an extrinsic role in anti-tumor immunity.
Figure 1. Figure 1.

Disclosures
Byrd: Acerta Pharma BV: Research Funding.
Tyrosin Kinase Inhibition Restores the Membrane Localization of FLT3-ITD
Authors: Reiter Katrin S. , Polzer Harald , Krupka Christina , Vick Binje , Maiser Andreas , Subklewe Marion S. , Jeremias Irmela , Leonhardt Heinrich , Spiekermann Karsten , Greif Philipp A. 
Published: October 2019

Abstract
The fms-related tyrosine kinase 3 (FLT3) receptor has been extensively studied over the past two decades with regards to oncogenic alterations that do not only serve as prognostic markers but also as targets for therapy in acute myeloid leukemia (AML). Internal tandem duplications (ITDs) became of special interest in this setting as they are associated with unfavorable prognosis. Due to sequence-dependent protein conformation changes, FLT3-ITD tends to auto-phosphorylate and displays a constitutive intracellular localization. As tyrosine kinase inhibitors (TKIs) efficiently block FLT3 auto-phosphorylation, we set out to study the effect of TKI on the subcellular localization of the FLT3 receptor and its mutants.
We visualized FLT3 WT and its mutants (N676K, D835Y, W51 (ITD), NPOS (ITD), K644R, W51-K644R) in transiently transfected U2OS cells, using anti-FLT3 immunofluorescence in combination with confocal microscopy. Treatment with 50nM of the TKI AC220 for 6 hours restored the membrane localization of both ITD (Figure 1 A) and D835Y. The TKI-induced surface expression of FLT3-ITD was confirmed by flow cytometry (mean fluorescence intensity, MFI) of BaF3 cells stably expressing FLT3 WT or mutants and in the homozygous (ITD/ITD) AML cell line (MV4-11). Similar results were obtained in cells from an AML patient derived xenograft (pdx) with FLT3-ITD and loss of heterozygosity (LOH) (Figure 1 B). Western blot analysis showed increased glycosylation of FLT3-ITD after TKI-treatment (Figure 1 C) suggesting that auto-phosphorylation prevents physiological processing of the receptor, which is required for maturation and surface expression. To monitor the subcellular localization of FLT3 and its mutants in vivo we have generated a FLT3-GFP fusion. Proliferation assays in BaF3 cells confirmed that the C-terminal GFP-tag does not alter the function of FLT3. Upon FLT3-ligand (FL) stimulation of FLT3-GFP we observed FLT3-internalization by life cell imaging in U2OS cells. Conversely, TKI treatment increased cell surface expression of FLT3-GFP.
Our findings have translational implications when considering FLT3 as a target for immunotherapy in AML, since FLT3 surface expression might facilitate antigen recognition. Especially FLT3-ITD positive patients with LOH may benefit from a combined TKI-immunotherapy approach. Therefore, we currently study the effect of AC220 on FLT3 antibody-dependent cellular cytotoxicity (ADCC), mediated by natural killer (NK) cells. Taken together, we discovered that TKI alters the subcellular localization and protein processing of specific FLT3 mutants. However, the complex interplay between the individual posttranslational modifications and their role in FLT3 traffic remains elusive.

Disclosures
Krupka: AMGEN Research (Munich): Research Funding. Subklewe:AMGEN Research (Munich): Research Funding.
Activation of Simultaneous Apoptosis and Necroptosis to Eradicate Drug Resistant Leukemia
Authors: McComb Scott , Aguadé-Gorgorió Julia , Marovca Blerim , Harder Lena , Cario Gunnar , Eckert Cornelia , Schrappe Martin , Stanulla Martin , von Stackelberg Arend , Bourquin Jean-Pierre , Bornhauser Beat 
Published: October 2019

Abstract
Dysregulation of apoptotic pathways provides an indiscriminate mechanism for refractory acute lymphoblastic leukemia (ALL) to escape cell death induced by many chemotherapeutic compounds. Here we have assessed the potential of SMAC mimetic (SM) compounds to short circuit cell death resistance by blocking the pro-survival cellular inhibitor of apoptosis (cIAP) proteins. By screening a large set of patient-derived precursor B-cell ALL samples in an ex vivo model of the leukemia microenvironment we detect exquisite sensitivity to two different SM compounds, Birinapant and LCL161, in about one third of ALL samples. Strong ex vivo SM activity correlated with potent in vivo anti-leukemic efficacy against de novo refractory and relapsed ALL xenografts. Intriguingly, we find that although SM-sensitivity is independent of TNF and TNFR1 levels, expression of TNFR2 is highly predictive of response to SM in two separate cohorts of ALL samples, suggesting that TNFR2 expression may represent a promising biomarker for identifying SM-sensitive cells. Downstream, we employ a novel and powerful multi-colour Lenti-CRISPR approach to show that simultaneous disruption of both apoptotic and necroptotic genes is necessary to block SM-induced death. In contrast, disruption of RIP1 alone was adequate to block SM-induced apoptosis and necroptosis. Surprisingly, RIP1 loss had no significant impact on response to standard anti-leukemic therapies, supporting a view that the RIP1-dependent death pathway is not likely to be selected against in leukemia cells that have failed to respond to front line therapy. These results provide the first evidence that SM compounds can circumvent apoptotic escape in drug-refractory ALL through parallel activation of both RIP1-dependent apoptosis and necroptosis. Furthermore, our data strongly support further development of SM as anti-leukemic agents for treatment in resistant disease.

Disclosures
No relevant conflicts of interest to declare.