bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Risk Of Recurrence In Women With Venous Thromboembolism While Using Estrogens: A Prospective Observational Cohort Study
Authors: Kyrle Paul A , Eischer Lisbeth , Eichinger Sabine 
Published: October 2019

Abstract

Objective
Because of the high recurrence risk, guidelines recommend indefinite anticoagulation in women with a first unprovoked proximal deep-vein thrombosis (DVT) and/or pulmonary embolism (PE). The optimal duration of anticoagulation in women who had venous thromboembolism (VTE) while using estrogens is unknown. We therefore compared the risk of recurrent VTE between women who used estrogens at the time of first VTE and women who did not.

Methods
This analysis was performed within the frame of the Austrian Study on Recurrent Venous Thromboembolism (AUREC), an on-going prospective observational study. Patients with a first objectively confirmed DVT of the leg and/or PE who had received anticoagulants for 3 to 18 months were included. Exclusion criteria were: age < 18 years; VTE associated with surgery, trauma, cancer or pregnancy; long-term anticoagulation; natural inhibitor deficiency; lupus anticoagulant; homozygosity or double heterozygosity for factor V Leiden and/or the prothrombin mutation. Women were advised to refrain from further estrogen use and were excluded in case of non-adherence. The study end point was recurrent symptomatic DVT and/or PE verified by imaging. The local ethics committee approved the study and all patients gave written informed consent.

Results
We followed 630 women (mean age 46 +/- 17 years) who had been treated with oral anticoagulants for 7 (+/- 3) months for an average of 69 (+/- 52) months. Recurrent VTE was recorded in 71 patients (11%). Recurrent VTE occurred in 22 (7%) of 333 estrogen users and in 49 (17%) of 297 non-users. After 1, 2 and 5 years, the cumulative probability of recurrence was 1% (95% CI 0-2), 1% (95% CI 0-2) and 6% (95% CI 3-9) among estrogen users and 5% (95% CI 2-7), 9% (95% CI 6-13) and 17% (95% CI 12-22) among non-users, respectively (p < 0.001, Figure). Compared to non-users, estrogen users had a relative risk (RR) of recurrent VTE of 0.4 (95% CI 0.2-0.8) after adjustment for age, site of first VTE and factor V Leiden carrier status. Compared to non-users in the respective age groups, the RR of recurrent VTE was 0.4 (95% CI 0.2-0.8) among estrogen containing contraceptive users and was 0.7 (95% CI 0.3-1.5) among women using estrogen containing hormone replacement therapy.

Conclusion
Women who had their first VTE while using estrogens have a low risk of recurrent VTE. The recurrence risk is particularly low in estrogen containing contraceptive users. We therefore propose that these women should receive anticoagulant therapy for no longer than 3 months.

Disclosures:
No relevant conflicts of interest to declare.
Packed Red Blood Cells Modulate Expression Of Dendritic Cell Signal Transduction and Antigen Presentation Pathways
Authors: Dean Melinda M , Ki Katrina , Tran Thu V , Baidya Shoma , Christensen Anne-Marie , Flower Robert L 
Published: October 2019

Abstract

Introduction
Clinical studies suggest packed red blood cell (PRBC) transfusion modulates recipients' immune responses, resulting in increased rates of infection, morbidity and mortality. In addition, there is evidence that increased length of storage of PRBC before transfusion is associated with worse outcomes. The mechanisms driving immunosuppression and transfusion-related poor patient outcomes remain largely undefined. As dendritic cells (DC) play a critical role in initiation and regulation of the immune response, this study investigated transfusion-mediated modulation of expression of DC genes involved in inflammation, antigen presentation and signal transduction.

Methods
Monocyte-derived DC (MoDC) were differentiated following exposure to PRBC (at either D2, D14, D28 or D42 of routine storage) in an in vitro transfusion model. Gene expression profiling of MoDC generated “post-transfusion” was assessed at each time point using quantitative real time PCR using ΔΔCT method (relative to GAPDH and matched “no transfusion control”) and data analysed using ANOVA (95% CI). Expression array (Applied Biosystems) consisted of 28 genes broadly categorised into 1) chemokines, cytokines and receptors, 2) antigen presentation, 3) signal transduction.

Results
PRBC transfusion significantly modulated expression of genes associated with signal transduction (CD86, CD44, TAP2, all p<0.05) and antigen presentation (NFκB2, RELA, RELB, all p<0.05). Modulation of gene expression was associated with the storage age of the PRBC to which the “recipient” was exposed. Networks of gene interactions were generated (Ingenuity Systems) to illustrate potential mechanisms of immune deregulation that are associated with transfusion-related DC modulation.

Conclusions
This study reports, for the first time, transfusion-driven modulation of DC gene expression that was associated with the storage age of PRBC transfused. These results provide a basis for the early assessment of immune function that we plan to translate into clinical studies to evaluate the efficacy of diagnostic tools to predict poor patient outcomes.

Disclosures:
No relevant conflicts of interest to declare.
Pleiotrophin Improves Survival Following Radiation-Induced Myelosuppression and Mediates HSC Expansion Via Induction Of Ras Signaling
Authors: Himburg Heather A , Doan Phuong L. , Quarmyne Mamle , Nakamura Mai , Chao Nelson J. , Chute John P. 
Published: October 2019

Abstract
Discovery of the mechanisms through which the bone marrow microenvironment stimulates hematopoietic regeneration following myelosuppression could lead to therapies to accelerate hematopoietic reconstitution in patients receiving chemotherapy, total body irradiation and stem cell transplantation. We have previously shown that treatment with pleiotrophin (PTN), a heparin-binding growth factor which is secreted by BM endothelial cells (ECs), causes a 10-fold expansion of murine long term-HSCs in culture (Himburg et al. Nat Med 2010). More recently, we demonstrated that PTN-deficient mice have a >10-fold deficit in LT-HSCs and hematopoietic regenerative capacity compared to PTN+/+ mice, suggesting an important role for PTN in maintaining the HSC pool in vivo (Himburg et al. Cell Reports 2012). In keeping with this, 100% of PTN-deficient mice died prior to day +30 following 700 cGy total body irradiation (TBI) compared to 30% mortality in irradiated, PTN+/+ mice (P<0.0001). In order to determine the therapeutic potential of PTN as a systemically deliverable agent to accelerate hematopoietic recovery following myelosuppression, we irradiated C57Bl6 mice with 700 cGy TBI followed by every other day dosing with PTN or saline through day +14. At day +30, 80% of the PTN-treated mice were alive compared to 30% survival in the irradiated, saline-treated group (P=0.03). Interestingly, systemic administration of PTN was equally effective at promoting the survival of irradiated mice when administered as late as 96 hrs post-TBI, suggesting that PTN promotes HSC regeneration after injury. Mechanistically, PTN treatment significantly decreased HSC apoptosis following ionizing radiation exposure (P=0.007), which may explain, at least in part, the mitigative effects of PTN treatment on survival from radiation injury. Furthermore, we have discovered that PTN treatment strongly induces elements of the Ras/MEK/ERK signaling cascade in HSCs, including phosphorylation of Grb2 and ERK1/2, while also increasing levels of the ERK1/2 target genes, phospho-Erf1 and Fra-1. Consistent with this finding, PTN-deficient mice have significantly decreased levels of phospho-ERK1/2 in BM HSCs, suggesting that deficient Ras/MEK/ERK signaling may explain, at least in part, the HSC deficit observed in PTN-deficient mice. Importantly, pharmacologic inhibition of Ras or MEK1/2 proteins or genetic inhibition of KRas in BM KSL cells significantly abrogated PTN-mediated expansion of BM KSL cells and CFU-GEMMs in culture (P<0.01 for total KSL cells and CFU-GEMMs). Taken together, these results suggest that PTN-mediated expansion of HSCs may be dependent upon activation of the Ras/MEK/ERK pathway and provide the basis for further studies to delineate the role of this pathway in mediating PTN effects on the HSC pool in vitro and in vivo.

Disclosures:
No relevant conflicts of interest to declare.
Myelodysplastic Syndrome-Derived Stromal Cells Support Leukemia-Initiating Cells and Contradirectly Eliminate Normal CD34+ Clonogenic Cells
Authors: Horiguchi Hiroto , Kobune Masayoshi , Kikuchi Shohei , Iyama Satoshi , Takada Kohichi , Murase Kazuyuki , Ono Kaoru , Kamihara Yusuke , Hashimoto Akari , Tatekoshi Ayumi , Hayashi Tsuyoshi , Sato Tsutomu , Miyanishi Koji , Sato Yasushi , Takimoto Rishu , Kato Junji 
Published: October 2019

Abstract

Introduction
The failure of normal hematopoiesis in myeloid neoplasm could be induced by a variety of mechanism. Regarding myelodysplastic syndrome (MDS)/acute leukemia (AML), aberrant hematopoietic stem/progenitor cells with exhibiting ineffective hematopoiesis and impaired differentiation ability gradually substitute it for normal hematopoietic stem/progenitor cells during a long term as a consequent of replacement of stem cell niche. However, it has not yet been clarified precise mechanism how MDS stem/progenitor cells could replace normal hematopoietic stem/progenitor cells.

Methods
In an attempt to analyze the supporting activity of bone marrow (BM) stromal cells, we first established the MDS/AML-derived stromal cells and healthy volunteer (HV)-derived-stromal cells. Next, MDS/AML-derived CD34+ cells or normal CD34+ cells were cocultured with established stromal cells using cytokines including stem cell factor, thrombopoietin, flt3-ligand in the presence of notch ligand (for normal CD34+ cells) or IL-3 (for AML/MDS derived cells). Subsequently, we analyzed clonogenic cells after 2 weeks coculture, 5 week cobblestone area-forming cells (CAFC) and repopulating cells in immunedeficient mice (NSG mice).

Results
The support of clonogenic cells after 2 weeks coculture and 5 weeks CAFCs was observed after coculture with normal CD34+ cells and HV-derived stromal cells. Furthermore, these cocultured cells engrafted into immunedeficient mice. Interestingly, the number of colony-forming units (CFU) mixed cells (MIXs) and CAFC derived from CD34+ cells was drastically reduced after coculture with MDS/AML-derived stromal cells. Nevertheless, MDS/AML-derived stromal cells support the proliferation of leukemia-initiating cells (L-ICs) and L-ICs were detected after third replating. These results indicate that MDS/AML-derived stromal cells preferentially support leukemia stem/progenitor cells, but not normal CD34+ cells. We compared the mRNA expression between (HV)-derived-stromal cells, MDS/AML-derived stromal cells and 5-aza-dC-treated stromal cells. The expression of several factors including hedgehog-interacting protein (HHIP) was reduced in MDS/AML-derived stromal cells. 5-aza-dC treatment restored the expression in some of genes and the stromal supporting activity for normal CD34+ cells partially recovered.

Conclusion
These results suggest that reduction of several gene expressions was detected in MDS/AML stromal cells by changes of methylation status. The epigenetic alteration of stromal genome may be involved in the progression of myeloid disorders.

Disclosures:
No relevant conflicts of interest to declare.
Evidence That GPI-AP-Specific CTLs Are Not Involved In The “escape” Of Piga mutant Hematopoietic Stem Cells In Aplastic Anemia
Authors: Katagiri Takamasa , Maruyama Hiroyuki , Ohtake Shigeki , Saito Chizuru , Hosokawa Kohei , Ogawa Seishi , Nakao Shinji 
Published: October 2019

Abstract

Background
Hematopoietic stem cells (HSCs) harboring PIGA mutations acquire a survival advantage under immune pressure compared to normal HSCs in patients with acquired aplastic anemia (AA). Cytotoxic T cells (CTLs) specific to glycosylphosphatidylinositol-anchored proteins (GPI-APs) are reportedly involved in this survival advantage, because PIGA mutant HSCs cannot present GPI-AP-derived peptides via class I HLAs. However, there is no convincing evidence that CTLs specific to GPI-AP-derived peptides are involved in the “escape” hematopoiesis by PIGA mutant HSCs. We recently demonstrated that 31.4-99.4% HLA-A allele-lacking leukocytes (HLA-LLs) were detectable in approximately 13% of AA patients as a result of escape hematopoiesis by HSCs with uniparental disomy in the short arm of chromosome 6, and that some patients possessed both GPI-AP-deficient (GPI-AP-) leukocytes and HLA-LLs (Katagiri, et al. Blood 2011). We hypothesized that if GPI-AP-derived peptides serve as a target for CTLs that elicit the development of AA, HLA-LLs may always be detectable only in the GPI-AP+ leukocyte population, because PIGA mutant HSCs do not require the lack of HLA class Is for the escape from the attack by GPI-AP peptide-specific CTLs.

Objectives and Methods
To examine this hypothesis, the GPI-AP expression was analyzed in various leukocyte lineages in 32 (nine at diagnosis and 23 previously treated) AA patients possessing HLA-LLs by a flow cytometry (FCM) analysis with liquid fluorescent aerolysin.

Results
A total of 0.01%-50% GPI-AP- granulocytes (GPI-AP- Gs) were detected in 22 (69%) of the 32 HLA-LLs (+) patients. Of the 22 patients possessing both HLA-LLs and increased GPI-AP- Gs, HLA-LLs were detectable in GPI-AP+ cells alone in 19 patients (86%). However, in the remaining three patients, HLA-LLs were shown in both GPI-AP+ and GPI-AP- populations. To determine which mutation occurs first in HSCs with a PIGA mutation and 6pUPD, the lineage diversity of GPI-AP- HLA-LLs was determined in the three patients. In two of the three patients, the lineage diversity of GPI-AP- cells (G/monocytes (M)/T cells (T)/B cells (B)/NK cells (NK)) and G/M/T) was greater than that of the HLA-A-lacking cells (G/M/B/NK and G/M) suggesting that PIGA mutations occurred earlier in the maturation of HSCs than did 6pUPD. The lineage diversity was the same in the GPI-AP- cells and HLA-LLs in one patient

Conclusions
The presence of HLA-LLs in the GPI-AP- leukocyte population and lower lineage diversity in HLA-LLs than GPI-AP- leukocytes suggest that CTLs specific to GPI-APs are not involved in the escape of PIGA mutant HSCs in AA, and that other mechanisms, such as a lower sensitivity to myelosuppressive cytokines than wild-type HSCs, may contribute to the survival advantage of PIGA mutant HSCs.

Disclosures:
No relevant conflicts of interest to declare.
Ikaros Mutation Confers Integrin-Dependent Survival Of Pre-B Cells and Progression To Acute Lymphoblastic Leukemia
Authors: Jena Nilamani , Joshi Ila , Yoshida Toshimi , Qi Xiaoqing , Zhang Jiangwen , Georgopoulos Katia , Van Etten Richard A. 
Published: October 2019

Abstract
Deletion of the IKAROS DNA-binding domain generates dominant-negative isoforms that interfere with the transcriptional activity of the IKAROS family and correlate with poor prognosis in human precursor B cell acute lymphoblastic leukemias (B-ALL). In this study, we defined the role of the Ikaros family during pre-B cell differentiation, the stage from which human B-ALLs arise, by conditionally inactivating IKAROS DNA binding in the immediate precursors of pre-B cells in mice. We demonstrate a novel niche-dependent phase in early pre-B cell differentiation that supports self-renewal and proliferative expansion. Expression of dominant-negative IKAROS arrests cells in this state by augmenting integrin and MAPK signaling and attenuating pre-B cell receptor signaling and differentiation. Up-regulated genes in Ikaros mutant pre-B cells were highly enriched in pathways involved in focal adhesion and remodeling of the actin cytoskeleton. The mutant pre-B cells had increased β1 integrin-mediated adhesion and elevated levels of activated focal adhesion kinase (FAK), whereas treatment with a small molecule FAK inhibitor greatly reduced pre-B cell stromal adhesion and selectively induced apoptosis in Ikaros mutant but not WT pre-B cells. Transplantation of polyclonal Ikaros mutant pre-B cells into recipient mice resulted in long-latency oligoclonal pre-B-ALL, demonstrating that loss of IKAROS contributes to multistep B-leukemogenesis. The highly proliferative and aberrantly self-renewing phenotype of Ikaros-deficient pre-B cells illuminates mechanisms underlying human IKAROS mutant B-ALL and suggests new therapeutic strategies for treatment of this aggressive leukemia.

Disclosures:
Van Etten: Bristol Myers Squibb: Consultancy; Deciphera Pharmaceuticals: Consultancy; TEVA Pharmaceuticals: Consultancy, Research Funding.
The Acute Promyelocytic Leukemia-Oncoprotein PML-Raralpha Blocks Senescence and Disrupts The Atrx/Daxx Chromatin Remodeling Complex To Promote Leukemia
Authors: Korf Katharina , Wodrich Harald , Haschke Alexander , Evans Ron M. , Sternsdorf Thomas M. 
Published: October 2019

Abstract
Although Acute Promyelocytic Leukemia (APL) has become a curable disease due to in-depth understanding of the underlying molecular processes, its investigation has provided unique and valuable insights into the processes involved in leukemogenesis. Therefore we use it as a model disease. 99% of APL-patients express a PML-RAR fusion protein. While involvement of RAR has proven indispensable for oncogenicity, the role of the PML domain is far less clear. In our previous study (Sternsdorf et al., Cancer Cell, 2006) we found that substitution of PML with heterologous self-interaction domains suffices to induce leukemias, but drastically decreases oncogenic potency of the resulting fusion proteins. In this study, we have chosen the inverse strategy: we have modified the PML domain to create a more active artificial model oncoprotein by adapting PR to its biological environment: As the typical model organism for APL studies is the mouse, we have replaced the human PML domain with the murine PML domain. This oncoprotein (mPR) creates APL-type leukemias in mice with higher penetrance and shorter latency than its human counterpart, hPR. We have used this system to study immediate early effects of expression of the model oncoprotein.
While proliferating murine bone marrow cells go into senescence ex vivo, expression of mPR prevents this and robustly immortalizes murine bone marrow from every mouse strain tested so far. Senescence-associated upregulation of the cell-cycle regulators p21 and p19 was efficiently blocked by mPR expression. In mouse cells, mPR exhibits higher potency in disrupting the PML-associated Daxx/ATRX complex than hPR. Knockdown of ATRX, but not Daxx ameliorated ATRA-induced growth suppression and p21 upregulation in the human APL model cell line NB4. These data suggest, that PML-RAR promotes leukemogenesis by disrupting the Daxx/ATRX complex, which assembles at PML nuclear bodies during the onset of senescence.

Disclosures:
No relevant conflicts of interest to declare.
Dual PI3K/Akt/mTOR Pathway Inhibitor Potentiate Cytarabine Cytotoxicity For Acute Myeloid Leukemia
Authors: Kim Eun Kyung , Jin Kang Hyoung , Kim Hyung Joon , Lee Ji Won , Kim Hyery , Park Kyung Duk , Shin Hee Young , Ahn Hyo Seop 
Published: October 2019

Abstract
Phosphatidylinositol 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signal transduction pathway integrating signals from multiple receptor tyrosine kinases has been firmly established as a major determinant for cell growth, proliferation, and survival in a wide array of solid cancers. PI3K/Akt/mTOR pathway is frequently activated in acute myeloid leukemia (AML) cells and contributes to survival and drug-resistance of AML through various mechanisms. BEZ235 is one of the most promising dual inhibitor of PI3K and mTOR currently under clinical development in solid tumor area. In this study, the potential of BEZ235 was investigated as antileukemic agent using alone or with cytarabinearabinoside (AraC) as combination regimen.
AML cell lines KG-1, MV 4-11, THP-1 and HL60 were treated with AraC, BEZ235 and combination regimen with various mixed ratio. BEZ235 effectively inhibited leukemic cell growth with similar range of half maximal inhibitory concentration (IC50) values among different cell lines. Apoptosis was induced gradually as BEZ235 concentration increased, but significant level of apoptosis was not shown even at higher concentration beyond IC50 value. Then, AraC-resistant MV4-11 and THP-1 cell lines were chosen to investigate interaction between BEZ235 and AraC. Using CalcuSyn software based on Chou and Talalay analysis, Combination Index (CI) value was calculated in each combination regimen. Moderate to strong synergism was shown and it was well maintained as combination ratio of AraC versus BEZ235 gradually decreased from 20:1 to 1,000:1. BEZ235 reduced resistance to AraC when it was added as combination regimen, and significance of combination effect changed according to AraC concentration. When antileukemic effect was compared among combination regimens with different schedules, synergism was maximized when BEZ235 was pretreated before AraC administration. This means BEZ235 sensitizes leukemic cells to apoptotic effect of AraC.
Genetic alteration in PI3K/Akt/mTOR pathway is an attractive target to investigate and dual pathway inhibitor BEZ235 has potential to maximize AML treatment through sensitization of leukemic cells to conventional drug. The results of this experiment suggest the possibility of designing early clinical trials for the therapy - AraC administration after dual inhibitor BEZ administration. An effective control of a signal transmission system using innovate new drugs along with an accurate understanding of a signal transmission system which is the target of therapy will establish a foothold for the development of AML therapy.

Disclosures:
No relevant conflicts of interest to declare.