bluuebunny/crossref_metadata_2025_split huggingface dataset:
Expression of Kindlins in Acute Myeloid Leukemia
Authors: Mi Yingchang , Wu Wenbin , Zhang Qing , Li Yan , Li Xiaoyan , Tian Zheng , Wang Min 
Published: October 2019

Abstract
Abstract 4910
The Kindlin family of intracellular proteins has recently emerged as key regulators of cellular functions and cell-matrix interactions. They comprise of three evolutionarily conserved members, kindlin-1, kindlin-2 and kindlin-3, they share considerable sequence and structural similarities. A few of study revealed that Kindlin-2 influences solid tumor cell invasion and resistance. With regard to AML, the influence of Kindlins is still unknown. To evaluate the clinical significance of Kindlin-2 in acute myeloid leukemia (AML), we investigated the expression of Kindlin-2, kindling-3 in AML cells.

1. Materials and methods
K562, KG-1a, HL60, U937, Jurkat cell lines were cultured in RPMI 1640 medium, supplemented with 10% fetal bovine serum (FBS, GIBCO) at 37°C in a humidified atmosphere of 5% CO2. Bone marrow (BM) samples were obtained from 88 patients with de novo AML from Blood Diseases Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College (CAMS & PUMC). Samples of 9 normal donors and ITP were used as the control group.
Bone marrow mononuclear cells (BMMCs) were prepared by Ficoll-Hypaque density gradient centrifugation. Expressions of Kindlin-2, Kindlin-3 were detected by RQ-PCR. The following primers for real-time PCR were used: (a) Kindlin-2 sense primer, 5'-CCGCTCGAGCTATGCGTATCCCCGTAG-3'; (b) Kindlin-2 antisense primer, 5'-CGACGCGTCTAGCGAGGGGTTGTC-3'; (c) Kindlin-3 sense primer, 5'-CCGCTCGAGCTATGCGTATCCCCGTAG-3'; (d) Kindlin-3 antisense primer, 5'-CGACGCGTCTAGCGAGGGGTTGTC-3'; (e) GAPDH sense primer, 5'-GAAGGTGAAGGTCGGAGTC-3'; (f) GAPDH antisense primer, 5'-GAAGATGGTGATGGGATTTC-3'. Analysis was performed using ABI 7500 Sequence Detection software (Applied Biosystems). The expression of Kindlin-2 and Kindlin-3 were showed as RQ value calculated through ΔΔCt method [ΔΔCt = (CtKindlin □ CtGAPDH)sample □ (CtKindlin □ CtGAPDH)calibrator]. The ΔCt (CtKindlin □ CtGAPDH) of K562 was defined as calibrator, and the RQ of calibrator was 1.000. Relationships between Kindlin-2, Kindlin-3 and the patients' clinical data were analyzed.

2. Results

Expression of Kindlins in newly diagnosis AML
The level of Kindlin-2 in AML (0.163±1.665) was significantly lower than that in non-AML (1.683±1.395) controls (p=0.010). No significant difference was found between the AML and controls in levels of Kindlin-3 (p=0.216). Out of the 79 patients who accepted treatment, 61 patients achieved complete remission (CR) and 18 patients were NR. Patients with higher expression of Kindlin-2 had a higher CR rate (86.8% vs 68.3%) (p=0.050). Expression of kindling 3 was unrelated to CR rate. Both of kindling-2 and kindling-3 increased after CR. This finding implicates Kindlin-2 as a potential prognostic factor of AML.

Disclosures:
No relevant conflicts of interest to declare.
Hyper Immunoglobulin E Syndrome (HIES): Report of a New Case Treated with Omalizumab and Dexametasone
Authors: Cortázar-Benítez Luis Fernando , Rojas Sotelo R.M. , Vargas-Viveros Pablo , Hurtado-Monroy Rafael , Romo Aguirre Catalina , Valenzuela Tamariz Jorge , Zárate Alejandra , Masse Sergio 
Published: October 2019

Abstract
Abstract 4934
The classical triad of eczema, recurrent skin and lung infections and high serum IgE, other connective tissue, cardiac and brain abnormalities are the accepted diagnostic criteria for HIES. The treatment of HIES is not established due to the scarce number of published cases, however is limited to skin local measures and supportive care. Herein we inform a severe case with response to Omalizumab, a monoclonal anti IgE antibody in combination with Dexametasone.
A 36 year old male with family history of asthma and atopic dermatitis (AD). Since child, the patient had allergies and AD. At 8 years he presented asthma and recurrent upper airway infections at 2 to 3 times per year and 7 dental pieces removal. In the last 3 years he noted important increase of AD with very intense and disabling pruritus without response to treatment and progression to generalized erithrodermia, nodes in legs and arms, and skin trasudate, axilar and inguinal lymph nodes (6×4 cm). Laboratory positive test were eosinophils 6%, serum IgE 29,280 IU/mL. Skin biopsy showed psoriasis-like dermatitis, micro abscesses and dermatopathic lymphadenopathy. We started treatment with trimetoprim/Sulfamethoxazole 80/400 mg twice a day, oral Dexametasone 20 mg weekly and Omalizumab 200 mg subcutaneously every two weeks. After 15 days of treatment there was a rapid improvement of skin derangement and IgE levels dropped to 2000 IU/mL after 16 weeks of treatment. Patient continues on treatment with no side effects and pruritus disappears.
Omalizumab is a monoclonal antibody against IgE recently introduced to the treatment of HIES in a few patients reported since 2008 with excellent response, along with intensive care of skin lesions, prompt antibiotic and antimycotic treatment for infections are the mainstay of HIES management.

Disclosures:
No relevant conflicts of interest to declare.
Generation of Melanoma Antigen Specific T Cells From Human Ips Cells Using a Feeder Free System.
Authors: Vizcardo Raul , Masuda Kyoko , Yamada Daisuke , Ikawa Tomokatsu , Shimizu Kanako , Fujii Shin-ichiro , Koseki Haruhiko , Kawamoto Hiroshi 
Published: October 2019

Abstract
Abstract 2292
iPS cells can be induced from various types of somatic cells by reprogramming them using Yamanaka factors (Oct4, Sox2, Klf4, and Myc). They are reported to be very similar to ES cells in many respects, such as gene expression pattern and multipotency. They are expected to be used as a cell source for production of various types of cells to be used in regeneration medicine. In this study, we planned to use iPS cells as a progenitor source for immune cell therapy. For this purpose, iPS cells derived from a lymphocyte that shows a certain antigen-specificity are preferable, because antigen specificity is inherited to iPS cells made from the lymphocyte. For example, if iPS cell are produced from cytotoxic T cells specific to a tumor antigen, T cells generated from these T-iPS can be used in cell therapy for patients bearing cancer.
We have succeeded in establishing iPS cells from human mature T cells (T-iPS cells), namely from whole CD3+ cells or CD4−CD8+ cells of cord blood as well of adult peripheral blood. These T-iPS cells were confirmed to bear productively rearranged TCRβ chain gene. These T-iPS cells were differentiated to CD4+CD8+ double positive T cells and eventually into CD8+ single positive T cells expressing αβTCR in an in vitro co-culture system using OP9-DL1 stromal cells. Such T cell induction occurred more efficiently from T-iPS cells than from human ES cells or from iPS cells derived from other cell types. Sequence analysis of their TCRβ chain suggests that these T cells preserved their original antigen specificity. To confirm that T-iPS cells can generate antigen specific T cells we established T-iPS cells from mature cytotoxic T cells specific to MART-1 (Melanoma antigen recognized by T cells 1) epitope. These MART-1 T-iPS cells were differentiated in vitro until CD4+CD8+ double positive T cells. By stimulation with anti-CD3 antibody, DP cells generated in vitro from MART1-T-iPS cells turned into a large number of CD8+ T cells specific to MART-1, which produced a substantial amount of IFNg upon TCR stimulation. The present study thus provides a novel method for cloning and expanding CD8+ T cells specific to a given antigen, which can be potentially applied for cell therapy against cancer.

Disclosures:
No relevant conflicts of interest to declare.
Hematopoietic Lineage Selective Roles for Trib Pseudokinases Post Bone Marrow Transplantation.
Authors: Verma Rakesh , Dev Arvind , Wojchowski Don M. 
Published: October 2019

Abstract
Abstract 2298
Trib's 1–3 are a homologous set of pseudokinases that (through their molecular actions as adaptors for E3 ubiquitin ligases, Akt and/or Mek/Erk) can exert diverse tissue and cellular effects. Upon ectopic expression, Trib2 promotes myeloid leukemia, and is elevated in its expression in melanoma. Trib3 can modulate insulin action, adipogenesis, and cardiomyopathy. Trib1 is implicated in inflammatory cytokine responses, and megakaryocytic leukemia. This illustrates the range of Trib functions, and also points to important regulatory roles for Tribs during hematopoiesis. To date the majority of Trib action studies have applied gain-of-function approaches. To gain new insight into hematopoietic roles of Trib2 and Trib3, we have generated novel knockout mouse models, and have employed a bone marrow transplant approach to assess possible effects of Trib2 or Trib3- deficiency on blood cell development. This approach also brings to bear the frequent requirement of physiological stress for the engagement of Trib pseudokinases. We assessed blood lineage reconstitution via hematological and flow cytometric analyses in irradiated (2 × 450 RAD) and bone marrow transplanted congenic recipient C57BL/B6.Ptprca, Ly-5.1 mice (5 × 105 Trib2-KO, Trib3-KO or wild-type donor cells). In all experiments, full representation of donor derived Ly5.2 hematopoietic cells was confirmed in blood, bone marrow and spleen. Trib2-KO and Trib3-KO transplanted mice each exhibited skewed hematopoietic reconstitution profiles (as compared directly to wild-type BM transplanted controls). Trib2-KO transplanted mice exhibited persistently elevated lymphoid and WBC levels at week 6, 10 and 36 post BMT, and peripheral blood CD19+/B220+/Ly5.2+ B-cells also were elevated. In contrast, Trib3-KO transplanted mice exhibited sustained increases in reticulocyte production, together with heightened splenic erythropoiesis. Within a bone marrow transplantation context, findings reveal novel lineage- specific roles for Trib2 during lymphopoiesis, and for Trib3 during stress erythropoiesis.

Disclosures:
No relevant conflicts of interest to declare.
Activation of Notch Signaling Mediates Ex Vivo Expansion of Multilineage, Engrafting Murine Hematopoietic Progenitors From Embryonic Sources.
Authors: Hadland Brandon K , Varnum-Finney Barbara , Bernstein Irwin D. 
Published: October 2019

Abstract
Abstract 2321
An important goal in the application of pluripotent stem cells (PSC) for therapeutic purposes is the derivation of hematopoietic stem and progenitor cells (HSPC) capable of efficient engraftment in vivo. Fundamental to achieving this goal is improved understanding of key signal pathways required to establish, maintain and expand HSPCs from embryonic sources. Ex vivo activation of Notch signaling in mouse bone marrow and human cord blood-derived HSC can facilitate expansion of rapidly engrafting multilineage progenitors, which has recently been translated for therapeutic purposes. In contrast, similar expansion of engrafting progenitors has not been successful from PSC. This prompted us to evaluate whether embryonic-derived HSPC have capacity to respond to ligand-induced Notch signaling ex vivo, and whether Notch activation could promote expansion of engrafting progenitors from these embryonic sources. We have examined the effects of ex vivo activation of Notch receptors by immobilized, exogenous Notch ligands on highly enriched populations of embryonic HSC and HSC precursors (pre-HSC) at various developmental stages. We find that activation of Notch by the ligand Delta1 within HSC/pre-HSC isolated from embryonic aorta-gonad-mesonephros (AGM) promotes expansion of progenitors with erythromyeloid colony forming potential and T/B-lymphoid potential in vitro, with concurrent expression of surface phenotypes resembling fetal liver-stage HSC. Furthermore, Notch activation in embryonic HSPC also mediates expansion of progenitors with rapidly engrafting myeloid and lymphoid capacity in irradiated mouse models. Our results demonstrate that embryonic stage HSPC have capacity to expand in response to Notch activation, and thus further studies comparing AGM- and PSC-derived hematopoietic precursors are needed to elucidate differences that may account for failure to expand HSPC from PSC.

Disclosures:
Bernstein: Seattle Genetics, Inc.: Consultancy.
Incorporation of a Histone Demethylase, JARID1B, in a Novel ETS-E Box DNA-Binding Complex in Murine Erythroid Progenitors.
Authors: Cai Ying , Nagarajan Lalitha , Curtis David J. , Brandt Stephen J. 
Published: October 2019

Abstract
Abstract 2335
The TAL1/SCL gene, originally discovered from its involvement by a recurrent chromosomal translocation in T-cell acute lymphoblastic leukemia, is important for hematopoietic stem cell and progenitor function and is essential for hematopoietic and vascular development. A member of the basic helix-loop-helix family of transcription factors, TAL1 binds a DNA sequence motif, CANNTG, termed the E box. We and others recently identified a novel TAL1-containing DNA-binding complex that includes an ETS protein, ELF2 (also known as NERF) in erythroid cells, and recognizes a bipartite sequence element containing an adjacent ETS binding site and E box. Our work showed this complex also contains proteins common to other TAL1 DNA-binding complexes described, including a LIM domain protein, LMO2 in erythroid cells, the LIM domain binding protein Ldb1, and putative single-stranded DNA-binding proteins SSBP2 and SSBP3. As both ELF2 and histone demethylase JARID1A, and later the related JARID1B, were identified using the same methodology (yeast two-hybrid analysis) to interact with LMO2, and multiple peptides derived from Jarid1b were identified by mass spectrometry analysis of highly purified Tal1-containing complexes from murine erythroleukemia (MEL) cells, we investigated whether JARID1B was present in the TAL1- and ELF2-containing complex. First, co-immunoprecipitation analysis identified Jarid1b in Tal1- or Elf2-containing immune precipitates and Tal1 in Jarid1b-containing immune precipitates from MEL cell extracts. Further, chromatin immunoprecipitation (ChIP) and re-ChIP analysis showed that Elf2 and Jarid1b co-occupied a region in the fourth intron of the Ssbp3 gene in MEL cells, previously demonstrated to be a physiologic target of the Tal1/Elf2 complex. Finally, knockdown of Elf2 and Jarid1b in MEL cells produced the same phenotype, decreased cellular proliferation. These results suggest a role for the JARID type histone demethylase JARID1B, and by inference histone demethylation, in the function of the ETS-E box DNA-binding complex and in proliferation of late-stage murine erythroid progenitors.

Disclosures:
No relevant conflicts of interest to declare.
Secreted Mediators of Self-Renewal of Hematopoietic Stem Cells Identified Using Bio-Informatic Analysis of Co-Cultures of HSC and Stromal Cells.
Authors: Vilne Baiba , Istvanffy Rouzanna , Eckl Christina , Bock Franziska , da Costa Olivia Prazeres , Stümpflen Volker , Peschel Christian , Mewes Hans-Werner , Oostendorp Robert A.J. 
Published: October 2019

Abstract
Abstract 2353
Hematopoiesis is maintained throughout life by the constant production of mature blood cells from hematopoietic stem cells (HSC). One mechanism by which the number of HSC is maintained is self-renewal, a cell division in which at least one of the daughter cells is a cell with the same functional potential as the mother cell. The mechanisms of this process are largely unknown. We have described cell lines that maintain self-renewal in culture. To study possible mechanisms and mediators involved in self-renewal, we performed co-cultures of HSC model cells: Lineage-negative Sca-1+ c-Kit+ (LSK) cells and HSC maintaining UG26–1B6 stromal cells. Microarray analyses were performed on cells prior to co-culture and cells sorted from the cultures. STEM clustering analysis of the data revealed that most changes in gene expression were due to early cell activation. Functional enrichment analysis revealed dynamic changes in focal adhesion and mTOR signaling, as well as changes in epigenetic regulators, such as HDAC in stromal cells. In LSK cells, genes whose products are involved in inflammation, Oxygen homeostasis and metabolism were differentially expressed after the co-culture. In addition, genes involved in the regulaton of H3K27 methylation were also affected. Interestingly, connective tissue growth factor (CTGF), which is involved in TGF-b, BMP and Wnt signaling, was upregulated in both stromal and LSK cells in the first day of co-culture. To study a possible extrinsic role of CTGF as a stromal mediator, we co-cultured siCTGF knockdown stromal cells with wild-type LSK cells. Since self-renewal requires cell division, we focused on cell cycle regulation of LSK cells. We found that knockdown of CTGF in stromal cells downregulates CTGF in LSK cells. In addition, knockdown of stromal CTGF downregulated Ccnd1, Cdk2, Cdkn1a (p21), Ep300 and Fos. On the other hand, decreased CTGF in stromal cells upregulates Cdkn1b (p27) and phosphorylation of Smad2/3. These results show that stromal CTGF regulates the cell cycle of LSK cells. On a functional level, we found that decreased stromal CTGF results in an increased production of MPP and myeloid colony-forming cells in 1-week co-cultures. We will present data showing whether and how a decrease in CTGF in stromal cells affects the maintenance of transplantable HSC. In summary, our current results indicate that reduced expression of CTGF in stromal cells regulates mediators of cell cycle and Smad2/3-mediated signaling in LSK cells, resulting in an increased production of myeloid progenitors.

Disclosures:
No relevant conflicts of interest to declare.
Maintenance Therapies for Multiple Myeloma (MM): A Network Meta-Analysis
Authors: Mahony Helen , Kumar Ambuj , Mhaskar Rahul , Miladinovic Branko , Wheatley Keith , Djulbegovic Benjamin 
Published: October 2019

Abstract
Abstract 236

Background:
There is little consensus on which maintenance therapy clinicians should choose for their patients. Since 1999, the three novel agents of bortezomib, lenalidomide, and thalidomide have been approved for use among patients with MM. These agents have been increasingly used as maintenance therapy. To date, only two randomized controlled trials of maintenance therapy have examined the efficacy of these novel agents head-to-head. Here, we conduct a network meta-analysis of bortezomib, lenalidomide, and thalidomide to determine which of these novel agents could potentially increase overall survival (OS) and progression-free survival (PFS).

Methods:
A comprehensive literature search of MEDLINE (PubMed), the Cochrane Central Register of Controlled Trials (CENTRAL), and meetings abstracts from American Society of Hematology, American Society of Clinical Oncology, European Society for Medical Oncology and European Hematology Association was undertaken to identify all phase III randomized controlled trials (RCTs) of maintenance therapy published until July 2012. We applied the Bayesian mixed treatment comparison (MTC) method under the random-effects model. The indirect comparisons were constructed from trials that have one treatment in common. For each included RCT, we calculated the hazard ratio (HR) and its corresponding standard error and used this to calculate the indirect estimates of HR and corresponding credible intervals (CrI). We also ranked the treatments according to the probability of best treatment and calculated the surface underneath the cumulative ranking curve (SUCRA). All analyses were conducted in WinBUGS 1.4.3 and Stata 11.2.

Results:
The network, number of trials for each comparison, and number of patients enrolled is shown in Figure 1. The network for OS was based on 12 RCTs enrolling 5542 patients and the network for PFS was constructed from 13 RCTs and 5784 patients. The MTC networks were consistent for both OS and PFS. For both OS and PFS, two comparisons were produced (Figure 2). For OS, the analysis showed that none of the treatments were superior. For PFS, lenalidomide was superior to thalidomide (HR = 0.58, 95% CrI [0.37, 0.94]). The estimates of SUCRA and rank probabilities (Figure 3) suggested that for OS bortezomib was best followed by lenalidomide and thalidomide. For PFS, lenalidomide was best followed by bortezomib and thalidomide.

Conclusion:
Using the MTC method, we found no evidence that any of the novel agents are superior to one another in terms of OS. Lenalidomide was the only novel agent which was superior to another active therapy (thalidomide). While these results provide preliminary evidence to which novel agent may be more beneficial as maintenance therapy, definitive conclusions cannot be reached until large, well designed RCTs evaluating these therapies head-to-head are conducted.

Disclosures:
No relevant conflicts of interest to declare.
Novel Bone Marrow Failure Syndrome Due to a Deletion of the EVI1/Mecom Gene.
Authors: van der Veken Lars T. , Bierings Marc , Maiburg Merel , Groenendaal Floris , Bloem Andries C , Knoers Nine V. , Buijs Arjan 
Published: October 2019

Abstract
Abstract 2368
We report on a patient born with severe congenital abnormalities including macrocephaly, brain MRI abnormalities, bilateral clubfeet, and bone marrow failure with pancytopenia presenting with none but T-lymphocytes in the peripheral blood.
A 824 kb to 868 kb heterozygous deletion on chromosome 3q26.2 was identified by array-CGH on peripheral blood using an Agilent 180k oligo array (Amadid 023363). The deletion truncated the 3' part of EVI1/MECOM and extended into microRNA 551b (MIR551B). The 3' MECOM deletion was confirmed by FISH using a MECOM specific probe on uncultured peripheral blood cells in 10 investigated metaphases and 200 interphase nuclei. To assess whether the deletion was constitutional, FISH analysis was performed on buccal swaps and cultured skin fibroblasts. Loss of one MECOM signal was observed in 200 interphases, and 10 metaphases of cells derived from buccal mucosa and skin fibroblasts, respectively, demonstrating the constitutional nature of the 3' MECOM deletion. Karyotyping and FISH analysis of both parents did not reveal a structural or numerical abnormality of 3q26.2 MECOM, indicating the deletion to be de novo.
EVI1/MECOM is a transcriptional regulator essential for maintaining embryonic and adult hematopoietic stem cells by directly regulating transcription of GATA2. In mouse models, homozygous disruption of MECOM results in embryonic lethality, with hypocellular bone marrow, reduced body size, small or absent limb buds, abnormal development of the nervous system and heart and massive hemorrhaging. Furthermore, MECOM heterozygosity leads to a marked impairment of the self-renewal capacity of hematopoietic stem cells.
In our case the heterozygous deletion results in 3' terminal truncated MECOM lacking the C-terminal acidic amino acid cluster domain (AD) encoding sequences. The AD of MECOM is important for activation of GATA2 transcription in vitro. This supports the notion that haploinsufficiency of MECOM causes bone marrow failure due to loss of critical MECOM transcriptional AD encoding sequences in this case, supposedly by deregulating GATA2 mediated control of hematopoietic stem cell homeostasis.
We will discuss our data in relation to one seemingly similar case deposited recently in the DECIPHER database. In conclusion, we report on a novel 3q26 EVI1/MECOM deletion syndrome with multiple severe congenital abnormalities and bone marrow failure due to EVI1/MECOM haploinsufficiency.

Disclosures:
No relevant conflicts of interest to declare.