bluuebunny/crossref_metadata_2025_split huggingface dataset:
Bone Marrow Cells From Low-Risk Myelodysplastic Syndrome Patients Show Features of Enhanced Autophagy
Authors: Wang Yuan-yuan , Chen Zi-Xing , Cen Jian-nong , Shen Hong-jie , Qi Xiao-fei , Yao Li 
Published: October 2019

Abstract
Abstract 4961
Myelodysplastic syndrome (MDS) is a preneoplastic condition that frequently develops into overt acute myeloid leukemia (AML). Beclin 1, a gene plays an important role in autophagy, has been recognized as a tumour suppressor. The aberrant expression of Beclin 1 has been correlated to the development and progression of cancer. However, The function and expression of Beclin 1 in MDS is largely unexplored.
The present study aimed to investigate Beclin 1 expression and its correlation with IPSS in bone marrow cells from Patients with Low-Risk MDS. We have analyzed the expression of Beclin 1 in BM mononuclear cells (BMMNCs) from patients of Low-Risk MDS (31 cases), 22 AML(22 cases) and healthy control (9 cases) by quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR), immunofluorescence and western blot analysis.
The expression level of Beclin 1 mRNA is significantly higher in BMMNCs in patients with Low-Risk MDS(mean± SD=6.26±4.87) when compared to healthy controls (mean± SD= 1.52±1.37), and the IPSS score was negatively correlated with the percentage of Beclin 1 positive cells. A markedly decreased expression of Beclin 1 was observed in AML (mean± SD=1.12±1.04) compared with healthy controls. The amount of Beclin 1 protein determined by either western blotting or immunofluorescence was markedly increased in MDS compared with that in controls (P<0.05). These results indicated that the cells from Patients with Low-Risk Myelodysplasia show features of enhanced autophagy, suggesting that the biological phenotype of autophagy, in addition to apoptosis and senescence, may also participate in the development of MDS and its progression to AML.

Disclosures:
No relevant conflicts of interest to declare.
Expression of Differentiation Antigens on Bone Marrow Myeloid Cells of the Patients with Myelodysplastic Syndromes and it's Clinical Significances
Authors: Shao Zonghong , Li Lijuan , Fu Rong , Wang Huaquan , Yue Lanzhu , Liu Hui , Wang Jun , Wang Honglei , Ruan Erbao , Qu Wen , Liang Yong , Wang Guojin , Wang Xiaoming , Liu Hong , Song Jia , Wu Yuhong , Xing Limin , Guan Jing 
Published: October 2019

Abstract
Abstract 4967

Objective
To study the abnormal differentiation of bone marrow myeloid cells in myelodysplastic syndromes (MDS) and its correlation with the prognosis of MDS patients.

Methods
Quantitative assessment of CD11b, CD13, CD16 and HLA-DR expression on the membrane of bone marrow granulocytes, and CD71 and glycophorin A on erythroblasts of 12 MDS patients in low-risk, 22 in high-risk and 31 normal controls was conducted with flow cytometry. The correlation between the abnormality of these antigen expression and the prognosis of MDS cases were analyzed.

Results
The granulocytic differentiation was analyzed with the combinations of CD13/CD11b, CD13/CD16 and CD11b/CD16. The “right hook”, “sickle” and “retroflex 7” shape expressions were found in normal controls while there were various changes in MDS groups. The ratios of CD11b-/CD11b+(0.39±0.34)and CD16-/CD16+(1.33 ±0.77)of high-risk MDS group were significantly higher than those of control group (0.07±0.05 and 0.39 ±0.31 respectively) (P<0.05). The MFI (mean fluorescence index) of SSC (side scatter) in the granulocyte gate of MDS groups was lower while their MFI of CD13 was higher. The mean percentages of CD11b-HLA-DR+ (3.88%±3.07%), CD11b- HLA-DR- (16.23%±15.59%), CD16-HLA-DR- (41.12%±24.53%), CD11b+CD16- (33.53%±17.26%) and CD13+CD16- (44.51%±21.99%) granulocytes of high-risk MDS group were significantly higher than those of low-risk and control groups (P<0.05). The erythroid cell lineage differentiation was analyzed with CD71/glycophorin A combination. Double antigen positive expression was found in all controls, but asynchronous expression of CD71/glycophorin A was found in some MDS cases. The mean percentage of double antigen positive cells in CD45- and glycophorin A+ cell population was significantly lower in low-risk and high-risk MDS groups. The abnormal numbers and patterns of the antigen expression of MDS cases correlated directly with their IPSS (international prognostic scoring system) (r=0.690, P=0.000) and WPSS (WHO adapted prognostic scoring system) (r=0.651, P=0.000) scores.

Conclusion
There were abnormal expressions of differentiation antigens on bone marrow myeloid cells of MDS patients. And the severity of these abnormal expressions was correlated with their prognosis. The abnormal differentiation of myeloid cells is probably involved in the pathogenesis of MDS. So the examination of these antigenic expressions with flow cytometry might be helpful for diagnosis and prognosis of MDS.

Disclosures:
No relevant conflicts of interest to declare.
Apoptosis on U266 Cells Induced by Brucine by Death-Receptor Pathway
Authors: Ma Yanping , Qiao Zhenhua 
Published: October 2019

Abstract
Abstract 5022

Objective
Multiple myeloma (MM) is an incurable hematological malignancy with high incidence in the elderly. The currently chemotherapeutic drugs show severe side effects, dose-limiting toxicity and development of resistance. The study in order to observe the Anti-proliferative of brucine which is a novel plant anti-cancer agents on multiple myeloma cell line-U266 and find the mechanism of apoptosis.

Method
U266 cells was cultured in 90% RPMI-1640 medium supplemented with 10% fetal bovine serum (FBS), 100 IU/ml penicillin and 100 μg/ml streptomycin, and maintained in a humidified atmosphere with 5% CO2 at 37°C. Cells (5×104) were plated in the presence or absence of the brucine (0, 0.05, 0.1, 0.2, 0.4 mg/ml) in 96 well culture plates for 24–72 h. The anti-proliferative response of brucine was assessed by MTT assay. Nuclear morphology of the treated cells was observed using Fluorescence microscope. The mitochondrial membrane potential of treated cell were measured by flow cytometry. The expression change of caspase-3 was detected using RT-PCR.

Results
The apoptotic effect of brucine show a dose and time dependent manner (p<0.05). The 50% inhibiting concentration of brucine was 0.16 mg/ml. Brucine treated cells show significant feature associated with apoptosis by Fluorescence microscope. The mitochondrial membrane potential using flow cytometry revealed no significant decline at different concentration. In addition, Brucine induced U266 cell apoptosis was caspase dependent, with caspase-3 activated by caspase-9 (p<0.05).

Conclusions
At 0.4mg/ml concentration range brucine can induce apoptosis in U266 cells. Brucine induces the apoptosis of U266 cell via death-receptor pathway.

Disclosures:
No relevant conflicts of interest to declare.
Heparin-Induced Thrombocytopenia Antibodies Inhibit PF4-Dependent Enhancement of Activated Protein C Formation by Binding to Antigenic Complexes Formed with the Chondroitin Sulfate Side-Chain of Thrombomodulin
Authors: Kowalska M. Anna , Rauova Lubica , Hayes Vincent , Cines Douglas B. , Bougie Daniel W. , Aster Richard H. , Krishnaswamy Sriram , Poncz Mortimer 
Published: October 2019

Abstract
Abstract 721
Previous studies have shown that platelet factor 4 (PF4) increases activated protein C (aPC) generation both in vitro and in vivo. PF4 increase of aPC generation by thrombin (IIa) and thrombomodulin (TM) complex followed a bell-shaped curve when tested in solution, on human TM expressing HEK293 and on endothelial cells. PF4 failed to enhance aPC in the presence of chondroitin sulfate (CS)-free TM. These results were consistent with PF4 binding to the CS on the TM glycosaminoglycan (GAG) domain and forming complexes that are similar to PF4/GAG antigenic complexes seen in heparin-induced thrombocytopenia (HIT). We tested the hypothesis that PF4 forms a HIT-like antigenic complex with the TM-CS using the HIT-like monoclonal antibody KKO. KKO abolished the potentiating effects of PF4 on aPC formation measured with TM in solution or with a TM-expressing cell line. To further address the nature of complexes formed between PF4 and TM, we used a mutant of PF4, PF4T38Q, which forms complexes with GAGs that are not recognized by KKO and a subgroup of HIT antibodies. Similar to PF4, PF4T38Q potentiated TM-dependent aPC generation in a bell-shaped manner, but this potentiation was not blocked by KKO. Moreover, KKO did not have any effect when PF4 was replaced with protamine sulfate (PS), which can also form macromolecular complexes with heparin/GAGs and can also enhance aPC generation. We also tested HIT antibodies isolated from patients that developed HIT with thrombocytopenia and thromboembolism developing >4 days after the last exposure to heparin. Patient IgGs specific for PF4/GAG complex were purified using PF4 bound to heparin columns. Specific binding of antibodies to PF4/heparin complexes was checked by ELISA. Complex-specific antibodies were then tested in an aPC generation assay in the presence IIa and TM and near peak concentration of PF4 and compared to a control human IgG. Three of four patient‘s antibodies significantly inhibited the increase in aPC generation in the presence of PF4. These studies provide evidence that HIT-like PF4/GAG complexes develop naturally in vivo. In this case, the ability of HIT or HIT-like antibodies to specifically inhibit the PF4-dependent increase in aPC formation may contribute to the prothrombotic state in HIT.

Disclosures:
No relevant conflicts of interest to declare.
Telomerase Gene Screening and Telomere Overhang Detection in Chinese Patients with Acute Myeloid Leukemia
Authors: Yan Siyi , Han Bing , Wu Yongji , Zhou Daobin , Zhao Yongqiang 
Published: October 2019

Abstract
Abstract 1467
Telomeres are complex structures capping the ends of all eukaryotic cell chromosomes. Loss-of-function mutations in telomerase complex genes reduce telomerase activity and shorten overall telomere length in leucocytes, and they can clinically manifest as aplastic anaemia and dyskeratosis congenita, which predispose to acute myeloid leukemia. Telomeres are constituted of double-stranded tandem TTAGGG repeats followed by a 3' G-rich single-stranded overhang, a crucial telomeric structural component responsible for the t-loop formation. Loss of telomerase function also leads to short telometric overhangs, potentially resulting in chromosome instability.
In this study, we screened bone marrow samples from 69 Chinese patients with acute myeloid leukemia (AML, excluding AML-M3) for variants in telomerase reverse transcriptase(TERT) and telomerase RNA component(TERC) gene. We also investigated the length of telomeric overhangs in those patients and 46 healthy individuals, using Southern blot analysis. Cytogenetic status, disease severity and short time survival rate in patients with AML were evaluated. Our cohort included 2 M1,48 M2, 5 M4, 5 M5 and 9 secondary AML. Age was 47(13–77),medium follow-up time was 4 months (2–10 months). 3 TERT mutations (m896G>A, m1079C>G and m1451G>A) but no TERC mutation was identified in patients with AML. Patients carrying TERT mutations had very short telomeres, critical short overhangs and poor response to the induction chemotherapy and died in their follow-up period. In contrast to overall telomere length, which shortened with ageing, telomeric overhang lengths were constant overtime and much shorter in patients with AML compared to those of normal controls(P<0.001). In AML cohort, those who have shorter overhangs did worse than those with normal ones. Multivariant analysis showed telomeric overhang length, as well as unfavorable chromosome abnormalities, served as an independent prognostic marker for AML patients(P<0.0001). In conclusion, we report 3 mutations in TERT gene patients with AML, which may cause short telomere and overhang and account for poor prognosis in those patients. Short overhang length may be an independent factor for poor response and shorter survival in patients with AML. These findings would have to be confirmed in large, prospective studies.

Disclosures:
No relevant conflicts of interest to declare.
Pomalidomide Therapy for Myelofibrosis: Analysis of Results From Three Consecutive Clinical Trials
Authors: Begna Kebede , Pardanani Animesh , Mesa Ruben A. , Hogan William J , Litzow Mark R. , Tefferi Ayalew 
Published: October 2019

Abstract
Abstract 1759

Background:
Pomalidomide is a second generation IMiDsÒ immunomodulatory drug that has been shown to be active in the treatment of myelofibrosis (MF)-associated anemia (J Clin Oncol 2009; 27: 4563; Leukemia 2011;25: 301). In the current sponsor-independent analysis, we report long-term follow up data on patients from the Mayo Clinic who had participated in three consecutive clinical trials using single agent pomalidomide for MF.

Methods:
From May 2007 to January 2010, 94 patients with MF (including primary and post-polycythemia/essential thrombocythemia) were enrolled in three consecutive phase I and II clinical trials involving pomalidomide with or without prednisone. Eight two patients who received only single agent pomalidomide constitute the study population for the current study. Follow up information was updated in July, 2011.

Results:
Among the 82 study patients, 19 were included in a phase I dose escalation study (2.5-3.5 mg/day), 58 in a phase II low-dose single agent pomalidomide study (0.5 mg/day), and 5 in a phase II randomized study (2 mg/day). Median age was 67 years and 63 (77 %) patients were red blood cell transfusion dependent at study entry. Forty-five (55%), 24 (29%), 7 (9%), and 2 (2%) patients remained on pomalidomide therapy for at least 6, 12, 24, and 36 months, respectively (Table 1). The overall anemia response rate per IWG-MRT criteria was 27% (22/82). Response occurred in the first 6 months in 21 (96%) of the 22 responders. The median time for response was 2.3 months (range, 1–10). The median response duration was 16.5 months (2–40). The anemia response rate in patients with spleen size palpable at less than 10 cm below the costal margin was 44 % (17/39) vs. 10 % (4/39) in those whose spleen was palpable at or above 10 cm (p=0.002). The anemia response rate was twice as likely in JAK2 mutated versus non-mutated cases (30% vs 15% p=0.2). Anemia response rate was also higher in patients with at least 50% increase in absolute basophil count during the first month of therapy: 39% (19/49) vs 6% (2/32) p=0.001). The anemia response rate was not significantly affected by karyotype, transfusion need or leukocyte count. Among the anemia responders (n=22), 3 (14%) relapsed in the first 12 months; relapse was more likely in the presence of baseline leukocytosis (p=0.006). Among the 24 patients who took pomalidomide for at lease 12 months, grade 1 sensory neuropathy developed in 4 (16%) patients.

Conclusion:
Anemia response to pomalidomide therapy in myelofibrosis often occurs in the first 6 months of treatment and is more likely to occur in the presence of JAK2V617F and absence of marked splenomegaly. Long-term treatment with pomalidomide might be associated with sensory peripheral neuropathy in a subset of treated patients.

Disclosures:
No relevant conflicts of interest to declare.
Phenotypic Analysis and Single-Cell Gene Profiling of Human Regulatory T Cell Subsets in Human Graft-Versus-Host Disease
Authors: Xhaard Alienor , Dong Shen , Maiella Sylvie , Pang Yuanyu , Benecke Arndt , Bianchi Elisabetta , Socie Gerard , Rogge Lars 
Published: October 2019

Abstract
Abstract 1975
Acute graft versus host disease (aGVHD) is an important cause of morbidity and mortality after allogeneic hematopoietic stem cell transplantation (alloHSCT). CD4+FOXP3+ regulatory T cells (Treg) are a population with potent immunosuppressive properties and may offer a new way to prevent or treat aGVHD. Treg therapy has demonstrated high efficacy in mouse models; however, translation to human has been hampered by the identification of different subsets of human Treg and doubt about in vivo conversion of Treg into effector Th17 cells. Based on the expression of CD45RA and HLA-DR, we identified 3 different subsets of human FOXP3+ Treg in healthy subjects' peripheral blood as well as in cord blood. All 3 subsets were suppressive in vitro. Gene expression profiling combined with global pathway analysis revealed clearly distinct immune signatures for each subset, which were validated by analysis at the single-cell level. Single-cell gene profiling also uncovered a striking heterogeneity of gene expression within these Treg subpopulations and revealed that cytokine-expressing Treg did not downregulate FOXP3 and other Treg markers.
We prospectively studied Treg subsets in 18 consecutive alloHSCT recipients' peripheral blood. Median age was 47 years (range: 26 to 65). Analysis was performed before steroid initiation in patients with aGVHD (n=7) and at hematopoietic recovery in the control group (n=11). First sample was analyzed a median of 20 days after alloHSCT (range: 11 to 36) with no difference between the 2 groups. Percentages of FOXP3+ cells in CD4+ cells were not significantly different in aGVHD patients and in the control group (10.4 and 12.6%, p=0.53). However, we observed in the aGVHD group a strong alteration of Treg subsets compared to the control group, with a pronounced bias towards an activated phenotype. RA-DR+ cells were significantly more represented among FOXP3+ T cells in aGVHD patients than in the control group (80.8 versus 53%, p=0.003). Conversely, RA-DR- and RA+DR- cells were more frequent in the control group than in patients with aGVHD (26.8 versus 10.6% and 13.5 versus 2.1%, p=0.014 and p=0.0012, respectively).
Our data suggest that frequencies of specific Treg subpopulations, rather than the frequency of the total pool of CD4+FOXP3+ Treg, is altered in aGVHD and may serve as a biomarker for this condition.
Current work addresses the molecular and functional characteristics of Treg subsets in aGVHD patients. Since Treg have been suggested to convert into effector T cells in pro-inflammatory and lymphopenic conditions, we will also assess the potential “plasticity” of the three Treg subsets.

Disclosures:
No relevant conflicts of interest to declare.
Absence of the Wnt Antagonist Sclerostin Adversely Affects B Cell Development in the Bone Marrow Niche
Authors: Cain Corey J , Rueda Randell , McLelland Bryce T , Collette Nicole , Loots Gabriela , Manilay Jennifer O 
Published: October 2019

Abstract
Abstract 220
Hematopoietic cell fate decisions are dependent on their localized microenvironmental niche. In the bone, endosteal osteoblasts have been shown to support hematopoietic stem cells (HSC) self-renewal, as demonstrated by transgenic and knockout mouse models in which osteoblast populations were increased or decreased. In addition, Wnt signaling and the Wnt antagonist Dkk-1 have been implicated in various aspects of hematopoiesis and HSC self-renewal. Sclerostin (Sost) is a secreted protein that is primarily expressed by fully mature osteocytes and acts on osteoblasts as a negative regulator of bone growth, by antagonizing Wnt signaling by its binding to the Wnt co-receptors Lrp4, Lrp5, and/or Lrp6. Here, we investigated the role of Sost on hematopoiesis in the bone marrow niche. Increased osteoblast activity in sclerostin-knockout (Sost−/−) mice results in hypermineralized bones with small bone marrow cavities. As such, Sost−/− mice contain markedly reduced numbers of CD45+hematopoietic cells in the bone marrow. Since hematopoietic stem cell activity is dependent on osteoblast function, we examined whether the hyperactive osteoblast activity in Sost−/− mice influences the numbers of hematopoietic stem cells, lymphoid progenitor cells and myeloid progenitor cells in the bone marrow. Surprisingly, no differences were observed in hematopoietic stem and progenitor cell frequency and cell number. However, we found the bone marrow of Sost−/− mice to be depleted of B cells, and this reduction can be attributed to premature apoptosis beginning at the pre-pro-B cell stage. Examination of Sost expression showed that no hematopoietic cells expressed Sost, however, pre-pro, immature and recirculating B cells expressed Lrp5 and Lrp6. These gene expression patterns suggested that the defect in B cell development in Sost−/− mice is non-cell autonomous and that absence of Sost could affect Wnt signaling in these populations. We observed that the expression of Wnt target genes CCND1 and Lef-1 were not affected by the absence of Sost, but c-Myc was significantly upregulated in recirculating B cells in the bone marrow. We also observed a significant decrease in CXCL12 expression in the bone marrow stroma in Sost−/− mice, consistent with their inability to adequately support B cell development. Taken together, our results indicate that the B cell developmental defects in Sost−/− mice are non-cell autonomous, and we are currently performing reciprocal bone marrow transplantation experiments to further support this hypothesis. Our studies demonstrate a novel role for Sost in the regulation of B cell development in the bone marrow, and demonstrate that distinct Wnt antagonists play specific roles in the regulation of hematopoiesis.

Disclosures:
No relevant conflicts of interest to declare.
Risk of Acute Myeloid Leukemia Among Solid Organ Transplant Recipients
Authors: Morton Lindsay M , Clarke Christina A , Chang Ellen T , Hall Erin C , Lynch Charles F , Pfieffer Ruth , Weisenburger Dennis D , Engels Eric A 
Published: October 2019

Abstract
Abstract 2559

Background.
Patients receiving solid organ transplants experience increased risk of subsequent hematologic malignancies, particularly post-transplant lymphoproliferative disorder and non-Hodgkin lymphoma, likely in relation to infection with oncogenic viruses and pharmacologic immunosuppression to prevent graft rejection. However, less is known about the risks for myeloid neoplasms such as acute myeloid leukemia (AML).

Methods.
We linked data from the US Scientific Registry of Transplant Recipients, a national database of solid organ transplantation, with 13 state and regional cancer registries to obtain cancer occurrence among 175,732 solid organ transplants during 1987–2008. The observed number of patients developing AML was compared to that expected in the general population of the 13 registry areas using standardized incidence ratios (SIRs).

Results.
AML was identified in 107 solid organ transplant recipients (13.8 cases/100,000 person-years), a rate that was nearly three times higher than expected in the general population [SIR=2.9, 95% confidence interval (CI) 2.4–3.5]. Excess risks were highest among children and young adults but remained significantly elevated among individuals receiving a solid organ transplant prior to age 65 years (SIR, 95%CI: <20 years=9.3, 3.4–20.2; 20–34 years=6.3, 3.2–11.3; 35–49 years=3.9, 2.6–5.6; 50–64 years=2.5, 1.9–3.3; 65+years=1.7, 0.9–2.9). Patients receiving liver or heart and/or lung transplants had higher risks than patients receiving kidney transplants (liver=4.1, 2.9–5.7; heart and/or lung=4.0, 2.7–5.7; kidney=2.0, 1.4–2.7). Risks also were higher among individuals receiving a second solid organ transplant than those receiving their first (second=6.8, 3.5–11.9; first=2.7, 2.2–3.3). Excess risks persisted throughout the 22-year period and showed no trend over time.

Conclusions.
In the largest population-based study of transplant-related malignancies conducted to date, we demonstrate that patients receiving solid organ transplants have substantially elevated risk for subsequent AML. Future investigations should evaluate the possible role of specific immunosuppressive medications in the etiology of post-transplant AML.

Disclosures:
No relevant conflicts of interest to declare.