bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dietary ω-3 Fatty Acid Supplementation As a Potential New Therapy for Vasculopathy in Sickle Cell Disease: Proof of Concept in a Transgenic Mouse Model
Authors: Kalish Brian T , Matte Alessandro , Andolfo Immacolata , Iolascon Achille , Weinberg Olga , Siciliano Angela , Federti Enrica , Puder Mark , Brugnara Carlo , De Franceschi Lucia 
Published: October 2019

Abstract
Sickle cell disease (SCD) is a genetic red cell disorder characterized by hemolytic anemia in association with acute and chronic life-threatening clinical complications mainly related to inflammatory vasculopathy. Promising, but limited, human studies have shown potential therapeutic effects of ω-3 fatty acid supplementation in SCD. Here, we sought to compare the effects of 6 weeks ω-6 (soybean oil diet, SD) vs ω-3 (fish oil diet, FD) diet on vasculopathy in SCD mice (Hbatm1(HBA)Tow Hbbtm2(HBG1,HBB*)Tow). We treated SCD and control healthy mice (AA, Hbatm1(HBA)Tow Hbbtm3(HBG1,HBB)Tow) (n=6-7 animals in each group). In SCD mice, FD was associated with a decreased the ratio of ω-6 to ω-3 fatty acids in red cell membranes, as well as a decrease in neutrophil count compared to SD. We then evaluated the effects of FD on cardiovascular system of SCD mice. We found that FD resulted in significantly (i) lower diastolic blood pressure; (ii) reduced cardiac output; (iii) down-regulation of VCAM-1, ET-1 and HO-1 expression in isolated aorta compared to SD, reaching levels similar to those observed in AA mice. In the lung of SCD mice, FD significantly reduced ET-1 expression compared to SD. FD was also associated with reduced bronchoalveolar lavage leukocyte and protein contents compared to SD, suggesting protection against pulmonary leak. We then studied the effect of FD during vaso-occusive crises (VOCs), mimicked by exposing SCD mice to hypoxia/reoxygenation (H/R) stress. In SCD mice, FD significantly (i) ameliorated the H/R-induced reduction in hematocrit and hemoglobin levels; (ii) reduced the fraction of dense red cells and the H/R-induced increase in neutrophil count. Histologic analysis revealed that H/R stress recapitulates the SCD acute organ damage. In the lung of H/R SCD mice, FD reduced inflammatory cell infiltrates, bronchial mucus, thrombosis, and ET-1 and ET1-1R expression. In isolated aorta from SCD mice exposed to H/R, FD prevented the H/R-induced ET-1 and VCAM-1 expression and the H/R-induced up-regulation of anti-oxidant systems compared to SD. These findings provide new insight into the mechanism of action of ω-3 supplementation on the pathogenesis of SCD and strengthen the rationale for ω-3 dietary supplementation as a simple therapeutic intervention to reduce vascular dysfunction in SCD.

Disclosures
No relevant conflicts of interest to declare.
The G-CSF Induced MiR-143 Targets MAPK-Family Proteins and Is a Prognostic Factor for RIC-Transplanted AML Patients
Authors: Hartmann Jens-Uwe , Braeuer-Hartmann Daniela , Gerloff Dennis , Katzerke Christiane , Wurm Alexander Arthur , Müller-Tidow Carsten , Schwind Sebastian , Tenen Daniel G. , Niederwieser Dietger , Behre Gerhard 
Published: October 2019

Abstract
Acute myeloid leukemia (AML) is a heterogeneous clonal disorder of defective hematopoiesis of white blood cells and often characterized by limited treatment options and a poor prognosis. MicroRNAs, a class of small non-coding RNAs, have been reported over the last years to play an important role in the regulation of “normal” and pathological processes. By binding to the 3`UTR of their target mRNAs, microRNAs lead to the translational repression of their target gene. Deregulation of a certain microRNA thereby may lead to disrupted signal pathways, such as MAPK-signaling, and to tumorigenesis. However, the role of microRNAs in hematopoietic differentiation and blood cancer development remains largely unknown. In AML only few miRNAs are functionally characterized, and the clinical relevance of these miRNAs still remains to be determined. In this study we identified microRNA-143 (miR-143) to be highly upregulated in G-CSF stimulated CD34+ cells. We could also show the granulopoietic association of miR-143 in several hematopoietic cell line models. In addition, we show that miR-143 is upregulated in APL patients after ATRA treatment. QPCR analysis of pri-miR-143 expression in RIC (reduced intensity conditioning) transplanted AML patients reveal that high expression of miR-143 is associated with high event-free and overall survival. In functional studies, stable overexpression of miR-143 in K562-C/EBPα-p42 cells enhances the C/EBPalpha induced granulocytic differentiation significantly. In contrast, stable knock down of miR-143 reduced the ability of K562-C/EBPα-p42 cells to undergo granulocytic differentiation upon β-estradiol treatment. Moreover, chromatinimmunoprecipitation (ChIP) analysis show the direct binding of C/EBPα to the promoter region of miR-143. By in silico prediction, we found MAPK protein family members (MAPK1, MAPK3 and MAPK7) as predicted targets of miR-143. Western blot analysis of AML patient samples and G-CSF stimulated CD34+ cells clearly show an inverse correlation of miR-143 and MAPK7 (ERK5) protein expression. By transient overexpression of miR-143 we could show a strong downregulation of MAPK protein family member expression in NB4 cells. These studies suggest miR-143 as an essential factor in granulocytic differentiation driven by G-CSF. Deregulation of miR-143 may play an important role in the development of AML. Furthermore, we propose miR-143 as prognostic marker for AML patients undergoing RIC transplantation therapy.

Disclosures
No relevant conflicts of interest to declare.
Identification of the Lymphoma-Initiating Cell Population in a T Cell Lymphoma Mouse Model Resembling Human Anaplastic Large Cell Lymphoma
Authors: Klingeberg Cathrin , Kreutmair Stefanie , Miething Cornelius , Follo Marie , Peschel Christian , Illert Anna Lena , Duyster Justus 
Published: October 2019

Abstract
In 60% of anaplastic large cell lymphoma (ALCL) patients a translocation t(2;5) (p23;q35) is found, which results in NPM-ALK fusion gene expression and constitutive activation of the ALK tyrosine kinase. Immunophenotypic characterization of human ALCLs revealed highly CD30-positive cells of T- or Null-cell-origin. However, the origin of the lymphoma initiating cell population as well as NPM-ALK signal transduction in course of the disease remains unclear.
In this regard, we established a retroviral murine bone marrow transplantation model resembling human ALCL. Therefore we use an inducible Cre/loxP system where NPM-ALK expression is restricted to early T cells. We infected bone marrow of Lck-Cre transgenic mice with our MSCV-Stop-NPM-ALK-IRES-EGFP vector and transplanted it into lethally irradiated recipient mice. With a latency of 4-5 months, these mice developed Thy1.2-positive lymphomas and died from neoplastic T cell infiltration of bone marrow and lymphatic organs. Immunophenotypic analysis confirmed T cell origin of the lymphomas with the characteristic high CD30 expression. Staining of the T cell subpopulations demonstrated high NPM-ALK expression in immature CD4-/CD8- double negative T cells and undifferentiated CD4+/CD8+ double positive T cells. Interestingly, FACS-staining for the proliferation marker Ki-67 as well as the activation marker CD30 revealed highest expression in the CD4-/CD8- double negative T cells. Therefore we hypothesized that the lymphoma-initiating cell must be within this early T cell population.
To substantiate our hypothesis we performed secondary transplantations with sorted T cell subpopulations and indeed, only the CD4-/CD8- double negative population was able to initiate T cell lymphoma in the recipient mice. Immunophenotypic characterization of the lymphoma population of these secondary transplanted mice revealed undifferentiated T cells of all CD4/CD8 subtypes, which argues for the existence of a lymphoma initiating cell population, which can still partly differentiate. Interestingly the CD4-/CD8- double negative lymphoma population aberrantly expressed the T cell receptor alpha/beta chain, which may allow these early T cells to establish a systemic lymphoma. Further analysis of the lymphoma population showed lymphatic precursors (CLP) as well as multipotent progenitors (MMP) and haematopoetic stem cells (LSK), which suggests early bone marrow or thymic progenitor cells as the pool of the lymphoma-initiating cell population.
We therefore were able to prove the existence of lymphoma initiating stem cells in a highly relevant NPM-ALK positive CD30 expressing mouse model of ALCL. Further analysis will give insides into eradication of the identified lymphoma stem cell population by clinical relevant NPM-ALK inhibitors and CD30 immunotoxins.

Disclosures
No relevant conflicts of interest to declare.
Targeting Polo-like Kinase in Acute Myeloid Leukemia
Authors: Cher Chae Yin , Man Cheuk Him , Lam Stephen S.Y. , Ho Eric S.K. , Ng Nelson K.L. , Hospital Marie-Anne , Tamburini Jerome , Leung Anskar Y.H. 
Published: October 2019

Abstract
Acute myeloid leukemia (AML) carrying fms-like tyrosine kinase 3 (FLT3)-internal tandem duplication (ITD) is associated with poor prognosis when treated with conventional chemotherapy and allogeneic hematopoietic stem cell transplantation (HSCT). Despite much interest in tyrosine kinase inhibitor (TKI) targeting FLT3 activation, drug resistance is invariable and acquisition of secondary point mutation in the tyrosine kinase domain (TKD) of FLT3 was frequently reported. We have shown that genes encoding cell cycle regulators including polo-like kinase 1 (PLK1), cell division cycle 25 homolog A (CDC25A), cyclin B2 (CCNB2), and cyclin E1 (CCNE1) were up-regulated in sorafenib-resistant primary AML samples. In particular, PLK1, a member of the polo-like kinase family, has been found to express at several checkpoints critical for cell cycle progression. PLK1 inhibitors have recently been exploited for the treatment of both solid organ and haematological cancers. We hypothesized that aberrant cell cycle progression mediated by increased PLK1 expression might confer survival advantage to drug resistant AML cells and be targetable by PLK1 inhibition. In vitro treatment with PLK1 inhibitors volasertib and BI 2536 significantly inhibited the growth of 8 AML cell lines (KG-1, ML2, MOLM-13, MV4-11, Kasumi-1, NB4, THP-1 and OCI-AML3) with IC50 (all in nM) ranging from 48.7 - 98.4 (volasertib) and 35.1 - 81.6 (BI 2536). The growth inhibitory effects of PLK1 inhibition on two FLT3-ITD+ cell lines, MOLM-13 and MV411, correlated with induction of apoptosis and cell cycle arrest at G2/M phase. Moreover, both PLK1 inhibitors significantly suppressed the growth of a sorafenib resistant MOLM-13R cell line and sorafenib naïve MOLM-13N cell line. Introduction of FLT3-ITD alone or FLT3-ITD and TKD double mutations into Ba/F3 cell lines sensitized them to the growth inhibitory effects of PLK1 inhibitors. Primary FLT3-ITD+ AML cells obtained from patients at TKI resistance were shown to be more sensitive to PLK1 inhibitors than those obtained before treatment. The results suggested that the TKI resistant clones could be effectively targeted by PLK1 inhibition, providing an insight to the design of combination treatment with FLT3 inhibitors in FLT3-ITD+ AML.

Disclosures
No relevant conflicts of interest to declare.
A Systematic Review of Definitions and Reporting of Bleeding Outcome Measures in Hemophilia
Authors: Chai-Adisaksopha Chatree , Hillis Christopher M. , Thabane Lehana , Iorio Alfonso 
Published: October 2019

Abstract
Background: Bleeding frequency is an important outcome commonly used in hemophilia studies. There is variation in practice in how bleeding is measured and defined. We therefore performed a systematic review of the studies reporting bleeding frequency in hemophilia patients.
Methods: MEDLINE, EMBASE, and the Cochrane Central Register of Control Trials were searched from January 1990 to January 2014. We included all published studies that included patients with hemophilia A and/or B and reported some measure of bleeding. Two reviewers independently performed title and abstract screening, full-text review and data abstraction of the identified studies.
Results: A total of 118 studies fulfilled the inclusion criteria. Study designs were randomized controlled trials [RCT] (16/118: 14%), cohort (80/118: 68%), cross-sectional (6/118: 5%) and others design (13/118: 11%). The median duration of follow-up (first quartile [Q1], third quartile [Q3]) was 20 (7.9, 50) months. Joint bleeding is the most common site of reported bleeding (66.9%). We found 10 different bleeding outcomes reported (absolute number of bleeding 60 (50.8%) studies, annualized bleeding rate 60 (50.8%) studies, bleed per month 10 (8.5%) studies and others (bleed per patient-year, bleed per week, bleed per season, etc.) 11 (9.3%) studies). Of these, 32 studies (27%) reported only mean or median without dispersion and 33 (28%) studies did not report any measures of central tendency (dispersion).
Conclusions: There is substantial variation in definitions and measures of bleeding outcomes in the hemophilia literature. This creates difficulty and limitations in comparing the outcomes between studies and in performing meta-analysis. The hemophilia research community needs to develop a consensus definition of bleeding and how to address the limitations associated with variations in measures of bleeding between centers and studies.
Table 1. Measures of bleeding frequency used in the studies Bleeding outcome measures n , (%) 1. Bleeding frequency# Absolute number of bleeds 60 (50.8) Annualized bleeding rate 60 (50.8) Bleed per month 10 (8.5) Bleed per 100 days 4 (3.4) Others (bleed per patient-year, bleed per week, bleed per 5 years, etc.) 11 (9.3) 2. Central tendency and dispersion# Mean with standard deviation 33 (28.0) Median with range or interquartile range 35 (27.0) Only mean or median 32 (27.1) Not reported 33 (28.0)
#Studies could include more than one level of characteristics.

Disclosures
No relevant conflicts of interest to declare.
The Stimulatory Role of CD300a in Diffuse Large B-Cell Lymphoma
Authors: Jiang Lei , Zhou Jeff Xiwu , Morse Herbert , Xu Yulian 
Published: October 2019

Abstract
CD300a is a type I transmembrane receptor protein which has shown inhibitory effect on B-cell receptor mediated signals. In an analysis of publicly available data on diffuse large B-cell lymphoma (DLBCL), however, we found that the expression levels of CD300A mRNA were inversely correlated with the overall survival of DLBCL patients. When analyzing the transcript levels of CD300a in human tissues, we found that CD300a mRNA levels were significantly greater in DLBCL tissues than benign lymphoid tissues (P<0.05). To decipher the role of CD300a in DLBCL, we used shRNA system to knock-down the expression levels of CD300a in DLBCL cell lines, and found that decreased levels of CD300a significantly inhibited cell proliferation of OCI-Ly1 cells, but not of VAL, OCI-Ly10 or SUDHL-8 cells. Mechanistically, reduced expression of CD300a resulted in a marked attenuation of Akt phosphorylation in OCI-Ly1 cells. Pharmacologic inhibition of PI3K by LY294002 displayed a similar inhibitory effect on cell proliferation, indicating the possible involvement of PI3K/Akt signaling pathway in CD300a’s effect. Furthermore, using a xenograft animal model, we found that decreasing expression levels of CD300a in OCI-Ly1 cells significantly inhibited tumor formation of these cells in vivo. Collectively, our results suggested a stimulatory role of CD300a in DLBCL which could serve as a potential biomarker and therapeutic target for this malignance.

Disclosures
No relevant conflicts of interest to declare.
Primary CLL Xenograft: A Model System to Study the Role of T-Cells in CLL Biology and Therapeutic Response
Authors: Oldreive Ceri E , Skowronska Anna , Agathanggelou Angelo , Parry Helen M , Krysov Sergey , Packham Graham K , Zbgniew Rudzki , Pratt Guy , Taylor A. Malcolm R , Moss Paul , Stankovic Tatjana 
Published: October 2019

Abstract
The interaction between chronic lymphocytic leukaemia (CLL) cells and T-cells is an important aspect of CLL biology. CLL cells require T-cell support for their proliferation and in addition induce proliferation of regulatory and cytotoxic (CD8+) T-cells. T-cell number and repertoire are both markedly affected by CLL therapy and there is considerable interest in how current treatments modulate the interaction between T-cells and the tumour clone. In this study we investigated whether this relationship was maintained in a xenotransplantation model.
CLL engraftment in NOG mice was facilitated by humanisation of the murine microenvironment by allogeneic CD34+ umbilical cord cells or CD14+ monocytes. Accelerated engraftment of both CLL and T-cell compartments was observed in xenografts derived from patients with progressive CLL, suggesting that the biological properties of both subsets are maintained in the murine model. Furthermore, the distribution of helper (CD4+), cytotoxic (CD8+) and regulatory (CD4+CD25+FoxP3+) T-cells was maintained within the xenografts, including retention of the CD4:CD8 ratio. Interestingly, the anergic PD-1+CD160+CD244+TIM3+ T-cell phenotype reported in CLL patients was also evident in T-cells expanded in xenograft models. Consistent with an anergic T-cell phenotype, T-cells from CLL xenografts lacked anti-tumour activity in vitro. Importantly, such anergic cells were observed when T-cells were reconstituted from allogeneic cord blood cells as well as autologous cells, suggesting that CLL cells have the ability to shape T-cell populations of different origin in diverse microenvironments. Finally, to investigate the interaction between specific T-cell subsets and engrafted CLL cells, CD4+, CD8+, and CD25+ T-cells were depleted prior to generation of xenografts. CD8+ T-cell depletion significantly prolonged CLL engraftment (p≤0.01) whereas neither depletion of CD4+ nor CD25+cells had a significant impact.
In summary, our results demonstrate that the relationship between CLL tumour cells and reactive T-cells is accurately maintained in a murine xenograft model. Such models will be of great value for investigation of aspects of T-cell function in CLL biology.

Disclosures
No relevant conflicts of interest to declare.
Syk and Btk Transduce Survival and Proliferation-Inducing Signals By Activating Distinct Signaling Pathways and Transcriptional Programs in AML Cells
Authors: Oellerich Thomas , Mohr Sebastian , Beck Julia , Bohnenberger Hanibal , Doebele Carmen , Corso Jasmin , Bug Gesine , Schuetz Ekkehard , Urlaub Henning , Serve Hubert 
Published: October 2019

Abstract
Acute myeloid leukemia (AML) is a genetically heterogeneous disease where multiple driver mutations coincide in branching hematopoietic subclones leading to malignant transformation. One important class of mutations alters the function of signaling intermediates such as FLT3, thereby helping AML cells to overcome the physiological communication with their microenvironment. To investigate, if also non-mutated signaling molecules might be potential drug targets in AML, we performed an unbiased proteomic kinase activity profiling. Our (phospho)proteomic and functional analyses revealed spleen tyrosine kinase (SYK) and Bruton’s tyrosine kinase (BTK) as novel non-mutated drug targets in AML. By immunohistochemical analyses we identified expression of both kinases in more than 70% of analyzed AML cases. Oncogenic SYK signaling in AML cells turned out to be critically regulated by beta-integrins upon intercellular contact formation between AML cells and bone marrow stroma cells. BTK, however, turned out to act downstream of FLT3-ITD to activate MYC-dependent oncogenic transcriptional programs. FLT3-ITD expression induced activation of BTK in cell culture models and patient-derived AML cultures and sensitized them towards BTK inhibitors. This newly identified FLT3-ITD/BTK signaling axis can be therapeutically exploited by combining FLT3 and BTK inhibitors resulting in additive apoptosis-inducing effects. As in some cases also FLT3-ITD-negative AML cells showed response to BTK inhibition, we characterized the BTK interactome and its downstream signaling networks by quantitative proteomic techniques and identified the apoptosis sensor Toll-like receptor 9 (TLR9) as an upstream activator of BTK in FLT-ITD-negative cells. In the absence of FLT3-ITD, TLR9 activates BTK to induce cell survival through NFkB as revealed by transcriptome sequencing and cell biological studies. While TLR9 stimulation enhanced AML cell proliferation, knock-down of TLR9 induced a partial cell cycle arrest and apoptosis. Thus, we here describe a novel microenvironment-derived signaling axis that may represent an important mechanism of AML maintenance in various important clinical settings including efficient FLT3-ITD inhibition and cytotoxic therapy. Taken together, these data highlight the importance of interactions between AML cells and their microenvironment and unravel cooperative dependencies between mutant oncoproteins and wild-type kinases in AML pathogenesis. They also provide a rationale for the clinical evaluation of SYK- and BTK inhibitors in AML.

Disclosures
Beck: Chronic Biomedical: Employment. Schuetz:Chronix Biomedical: Employment.
Reversal Multidrug Resistance and Biodistribution of Targeted Nanoparticles Tf-Peg-PLL-PLGA Combined with Daunorubicin and Tetrandrine in Tumor-Bearing Nude Mice
Authors: Guo Liting , Liu Ping , Wang Fei , Xia Guohua , Wang Yonglu , Chen Wei , Yin Haixiang , Chen Baoan 
Published: October 2019

Abstract
The study was to investigate the reversal multidrug resistance treatment and biodistribution of nanoparticles polyethylene glycol (PEG)-poly L-lysine (PLL)-poly lactic-co-glycolic acid (PLGA) (PEG-PLL-PLGA) modified by transferrin (Tf) combined with daunorubicin (DNR) and tetrandrine (Tet) in leukemia bearing nude mice model. Anti-tumor effect of nanoparticles in nude mice with K562/A02 resistant leukemia cell lines xenografts was observed. Subsequently, transcription and protein expression levels of transferrin receptor (TfR), P-glycoprotein (P-gp), MRP and NF-κB were determined by quantitative real-time polymerase chain reaction (quantitative real-time PCR) and Western blot analysis. DNR concentration in plasma, tumor tissue and major organs was detected by high performance liquid chromatography (HPLC). DNR-Tet-Tf-PEG-PLL-PLGA nanoparticles group significantly inhibited tumor, while the immunohistochemistry of tumor tissue displayed that the nanoparticles can induce apoptosis of tumor cells. Furthermore, DNR-Tet-Tf-PEG-PLL-PLGA group markedly increased TfR expression, and noticeably decreased P-gp, MRP and NF-kB expression. DNR concentration of tumor tissues in DNR-Tet-Tf-PEG-PLL-PLGA group injected by the tail vein was significantly higher than DNR-Tet-PEG-PLL-PLGA group. DNR-Tet-Tf-PEG-PLL-PLGA nanoparticles have the good ability to active targeting and reverse drug resistance, therefore provide a new initiative targeted therapy for cancer.

Disclosures
No relevant conflicts of interest to declare.