bluuebunny/crossref_metadata_2025_split huggingface dataset:
Non-Canonical NF-Kb Signaling Regulates Hematopoietic Stem Cell Self-Renewal and Microenvironment Interactions
Authors: Zhao Chen , Xiu Yan , Ashton John M , Xing Lianping , Morita Yoshikazu , Jordan Craig T. , Boyce Brenda 
Published: October 2019

Abstract
Abstract 859
RelB and NF-kB2 are the main effectors of NF-kB non-canonical signaling and play critical roles in many physiological processes. However, their role in hematopoietic stem/progenitor cell (HSPC) maintenance has not been characterized. To investigate this, we generated RelB/NF-kB2 double-knockout (dKO) mice and found that dKO HSPCs have profoundly impaired engraftment and self-renewal activity after transplantation into wild-type recipients. Transplantation of wild-type bone marrow cells into dKO mice to assess the role of the dKO microenvironment showed that wild-type HSPCs cycled more rapidly, were more abundant, and had developmental aberrancies: increased myeloid and decreased lymphoid lineages, similar to dKO HSPCs. Notably, when these wild-type cells were returned to normal hosts, these phenotypic changes were reversed, indicating a potent but transient phenotype conferred by the dKO microenvironment. However, dKO bone marrow stromal cell numbers were reduced, and bone-lining niche cells supported less HSPC expansion than controls. Further, increased dKO HSPC proliferation was associated with impaired expression of niche adhesion molecules by bone-lining cells and increased inflammatory cytokine expression by bone marrow cells. Thus, RelB/NF-kB2 signaling positively and intrinsically regulates HSPC self-renewal and maintains stromal/osteoblastic niches and negatively and extrinsically regulates HSPC expansion and lineage commitment through the marrow microenvironment.

Disclosures:
No relevant conflicts of interest to declare.
A Monoclonal Antibody Specifically Targeting CD44 Inhibits B-Cell Chronic Lymphocytic Leukemia Cell Survival In Vitro and In Vivo
Authors: Zhang Suping , Wu Christina C.N. , Fecteau Jessie-Farah , Cui Bing , Wu Rongrong , Rassenti Laura Z. , Lao Fitzgerald S. , Tawatao Rommel , Weigland Stefan , Prussak Charles , Carson Dennis A. , Kipps Thomas J. 
Published: October 2019

Abstract
Abstract 927
B-cell chronic lymphocytic leukemia (CLL) with a highly variable clinical course is characterized by the clonal expansion of CD5+ B cells in blood, secondary lymphoid tissues, and marrow. Survival of CLL cells are supported by cells within the tissue microenvironment and by signals from the extracellular matrix, which in part are mediated via interactions with CD44, a surface glycoprotein that is expressed at high levels by CLL B cells. While monoclonal antibody (mAb) therapy targeting CD20 has improved the survival of patients with CLL, a large number of CLL patients do not achieve durable responses or become refractory altogether to therapy with anti-CD20 mAb. In this study we evaluated the cytotoxic activity against CLL cells of a newly developed, humanized anti-CD44 mAb (RO5429083, Roche). We found that sub-microgram concentrations of this anti-CD44 mAb were directly cytotoxic for CLL cells of different patients (n = 32), but had little or no effect on the viability of lymphocytes isolated from the blood of healthy donors (n = 4). The levels of cytotoxicity induced by this anti-CD44 mAb were significantly associated with the levels of CD44 expressed by each of the CLL samples (Spearman R-0.5785, p<0.001). Furthermore, survival and downstream signaling events of CLL cells induced by hyaluronic acid (HA), the principal ligand of CD44, were inhibited by this CD44 mAb. Of note, this CD44 mAb also was equally cytotoxic for CLL cells co-cultured with mesenchymal stromal cells (MSC), which otherwise can support CLL-cell survival in vitro. We also examined whether this anti-CD44 mAb could induce clearance of human CLL cells engrafted into immune deficient RAG-2−/−/γc−/− mice. Treatment of such xenografted animals with as little as 1 mg/kg of this mAb resulted in the complete clearance of engrafted CLL cells, an effect not observed in control treated animals. Based on these pre-clinical studies, we consider this anti-CD44 mAb has high potential for providing effective treatment for patients with this disease.

Disclosures:
Weigland: Roche Diagnostics GmbH: Employment. Carson:Wintherix: Equity Ownership. Kipps:Gilead Sciences: Consultancy, Research Funding; GSK: Research Funding; Genentech: Research Funding; Abbott Industries: Research Funding; Celgene: Consultancy, Research Funding; Igenica: Equity Ownership, Membership on an entity's Board of Directors or advisory committees; Amgen: Research Funding.
Dimension of Residual CT Scan Mass in Hodgkin's Lymphoma (HL) Is a Negative Prognostic Factor in Patients with PET Negative After Chemo+/− Radiotherapy
Authors: Magagnoli Massimo , Marzo Katia , Balzarotti Monica , Rodari Marcello , Mazza Rita , Giordano Laura , Gandolfi Sara , Bramanti Stefania , Anastasia Antonella , Lutman Fabio Romano , Chiti Arturo , Santoro Armando 
Published: October 2019

Abstract
Abstract 93

BACKGROUND.
Positron emission tomography (PET) with 2-[18F]fluoro-2-deoxy-D-glucose (18F-FDG) currently represents the mainstay for response assessment in HL, as defined by revised response criteria (Cheson, 2007). PET negativity is mandatory to define complete remission (CR), independently from the persistence of residual masses at computed tomography (CT scan). Nevertheless, some reports suggest a slightly worse prognosis among patients (pts) with CT scan residual masses. The aim of this study was to evaluate the unfavorable predictive value of residual CT scan masses in HL pts with PET negative at the end of treatment.

MATERIAL AND METHODS.
The present analysis was retrospectively conducted in 105 pts with negative PET at the end of first or second line treatment, at our institution, from February 2004 to February 2009. All pts had disease evaluation performed also with CT scan.

RESULTS.
Main clinical characteristics: median age 58 years, males 62, B-symptoms 25, bulky disease 41, prior radiotherapy 57. Seventy-four pts were evaluated after first line treatment program, while 31 pts after salvage therapy program. In 76 pts, residual CT scan mass (PET-/CT scan +) of at least 2.0 cm in the largest diameter was assessed. Fifty-seven had only one site residual mass, while 19 pts had more than one site. Considering the whole series, with a median follow-up of 45 months, 23 pts relapsed, nine patients died and 94 are alive without disease.
The five year disease-free survival (DSF) for PET-/CT scan- vs PET-/CT scan+ pts was 89.4% and 68.7% respectively (P=0.053). The prognostic impact of residual mass at CT scan had a correlation with the dimension of the residual itself in a continuous fashion: the larger is the mass, the lower the DFS: HR 1.03 (1.01; 1.05) p 0.007. This difference is even more pronounced when a cut-off of 4 cm in the largest diameter of the residual mass is applied: in patients with a mass diameter ≥ vs < 4 cm, DFS is 50% vs 82% respectively (HR 3.25: CI 1.5; 7.04, p 0,029 – figure 1). Among all the other prognostic factors analyzed (number of masses, first vs salvage treatment program, sex, bulky disease, B-symptoms), no correlation with DSF or overall survival (OS) emerged.

CONCLUSIONS.
In our study we observe a significant difference in DFS among PET negative pts with or without CT scan residual masses after therapy for HL. This difference is more significant when the residual mass is larger than 4 cm. Thus, despite PET is the main tool in respons-e definition, CT scan maintains an important role and can not yet be abandoned. The role of consolidation radiotherapy in these cases should be focused.

Disclosures:
No relevant conflicts of interest to declare.
L-Leucine Improves the Anemia of the 5q- Syndrome Via Activation of the mTOR (mammalian target of rapamycin) Pathway
Authors: Virgilio Maria , Payne Elspeth , Narla Anupama , Sun Hong , Paw Barry H. , Look Thomas , Ebert Benjamin L. , Berliner Nancy , Khanna-Gupta Arati 
Published: October 2019

Abstract
Abstract 970
The “5q- syndrome” is a subtype of myelodysplastic syndrome (MDS) characterized by a profound macrocytic anemia that is thought to arise from the heterozygous loss of the RPS14 gene on chromosome 5q. The 5q- syndrome shares many characteristics with another ribosomopathy, Diamond Blackfan anemia (DBA), wherein half the patients have a heterozygous loss of a ribosomal protein gene, with RPS19 being the most frequently mutated. Both RPS19 and RPS14 are components of the 40S small ribosomal subunit. Heterozygous loss of both genes is thought to contribute to the pathophysiology of these ribosomopathies by impairing ribosome function and thereby decreasing mRNA translational efficiency. L-Leucine, a branched-chain amino acid has been shown to be a potent stimulator of mRNA translation. In a case report, administration of L-Leucine to a DBA patient resulted in complete recovery of the anemia and improvement in appetite and growth parameters. We therefore hypothesized that L-Leucine could improve the anemia associated with heterozygous loss of RPS14. To test this hypotheisis, we used zebrafish as an in vivo model for del(5q) MDS. Using antisense morpholinos, we knocked down Rps14 expression to haploinsufficient levels and observed anemia in the resulting morphants. Treatment of these morphants with L-Leucine, but not D-Leuicne, resulted in a dramatic increase in hemoglobinization as well as an increase in total erythroid cells. This observation was further validated in vitro using human CD34+ hematopoietic progenitor cells infected with RPS14 shRNA. Flow cytometric analysis demonstrated that treatment with L-Leucine increased the total number of erythroid cells (glycophorin-A/CD71 expressing cells) compared to untreated cells. In previous studies, L-leucine has been shown to upregulate mRNA translation by activating the mTOR (mammalian target of rapamycin) pathway and its downstream targets S6Kinase (S6K1) and 4E-Binding proteins (4E-BPs). We demonstrated increased levels of phospho-S6K1 as a result of L-Leucine treatment in the rps14 zebrafish morphant embryos. This increased phosphorylation of S6K1 was inhibited by rapamycin, suggesting specificity of mTOR activation in bringing about the L-Leucine effect. In summary, we have successfully demonstrated, in both a zebrafish model and in human primary hematopoietic stem cells, that L-Leucine alleviates the anemia associated with del(5q) MDS. This effect is likely mediated by activation of the mTOR pathway resulting in increased mRNA translation and an improvement in the anemia associated with loss of RPS14. Our studies support the further evaluation of L-Leucine as a potential therapeutic agent in the treatment of the 5q- syndrome.

Disclosures:
No relevant conflicts of interest to declare.
Thrombin Control Mechanisms and Thrombin Targets in Cancer Biology
Authors: Degen Jay L. , Palumbo Joseph S. 
Published: October 2019

Abstract SCI-17
A link between hemostasis and cancer has been recognized for more than a century, but the last decade has seen substantial strides in understanding the mechanisms by which hemostatic system components actively contribute to the malignant phenotype. The expression of procoagulants, such as tissue factor (TF), by tumor cells has been shown to be a poor prognostic factor in clinical studies and a crucial determinant of metastasis in animal models. While TF expressed by tumor cells likely plays a multifaceted role in cancer biology, a substantial body of evidence indicates that tumor cell-associated and circulating hemostatic system components (e.g., prothrombin, fibrinogen, platelets) play a cooperative role in supporting metastasis. The capacity of tumor cells to generate thrombin has been proposed to support metastasis through several mechanisms, including tumor cell proliferation, stable adhesion, regulation of apoptosis, and escape from innate immune surveillance mechanisms. More recently, the fundamental importance of endothelial regulators of thrombin activity in metastasis was established through studies of tumor dissemination in mice expressing mutant forms of thrombomodulin (TM). Mice expressing a TM derivative with reduced thrombin affinity (TMPro) exhibited a profoundly prometastatic phenotype relative to wild-type (WT) mice. The TMPro mutation was shown to support metastasis by promoting the survival of tumor cell emboli newly localized to the lung. The impact of the TMPro mutation on metastasis was dependent on tumor cell-associated tissue factor, prothrombin, thrombin function, and platelets. In contrast, mice expressing a mutant form of TM lacking the lectin-like domain (TMLed) that were shown previously to have altered immune function but normal thrombin affinity, exhibited metastatic potential comparable to wild-type mice. These studies further highlight the importance of the hemostatic system in metastasis and demonstrate that apart from tumor cell-associated and circulating procoagulants, TM-mediated regulation of hemostatic function strongly influences tumor cell metastatic success. In addition, recent studies of inflammation-driven cancer have revealed that the role of hemostatic factors in tumor biology is not limited to later phases of malignant progression, such as metastasis. Fibrin(ogen)-mediated regulation of leukocyte function was shown to support tumor development and tumor proliferation in a murine model of inflammation-driven colon cancer. Recent advances in our understanding of the role of hemostatic factors in cancer biology demonstrate that this system of proteins can be important in multiple phases of malignant progression, and underscore the potential utility of targeting selected coagulation factors as a novel adjunct therapy in the treatment of cancer.

Disclosures:
Palumbo: Novo Nordisk Corporation: Research Funding.
Cellular Bioenergetics in Lymphoid Neoplasia
Authors: Heiden Matthew Vander 
Published: October 2019

Abstract
Abstract SCI-25
Many cancer cells metabolize glucose by aerobic glycolysis, a phenomenon characterized by increased glycolysis with lactate production and decreased oxidative phosphorylation. We have argued that alterations in cell metabolism associated with cancer may be selected by cancer cells to meet the distinct metabolic needs of proliferation. Unlike metabolism in differentiated cells, which is geared toward efficient ATP generation, the metabolism in cancer cells must be adapted to facilitate the accumulation of biomass. Cancer cells divert a larger fraction of their nutrient metabolism to pathways other than mitochondrial respiration regardless of oxygen availability. Nevertheless, oxygen levels still influence how nutrients are metabolized. We have found that the source of carbon used in various anabolic processes varies based on oxygen levels. Furthermore, the enzymes used to metabolize nutrients can also differ based on the cellular context. This includes regulation of isocitrate dehydrogenase, an enzyme that is mutated in some cancers. There is also strong selection for use of the M2 isoform of pyruvate kinase (PK-M2) to metabolize glucose in cancer cell lines. However, evidence from mouse models suggests that PK-M2 is dispensable for glucose metabolism by many tumors in vivo, suggesting an alternate pathway to convert phosphoenolpyruvate to pyruvate can be used to metabolize glucose. This regulation of pyruvate kinase also plays an important role in hematopoietic stem cell biology. Together, these findings argue that distinct metabolic phenotypes exist among proliferating cells, and both environmental and genetic factors influence how metabolism is regulated to support cell growth.

Disclosures:
Vander Heiden: Agios Pharmaceuticals: Consultancy, Equity Ownership.
Therapeutic Targeting of Cancer Cell Metabolism in Lymphoid Neoplasia
Authors: Van Dang Chi 
Published: October 2019

Abstract
Abstract SCI-27
The MYC oncogene plays a pivotal role in human lymphoid neoplasias, specifically in lymphomas and acute leukemias, which are characterized by altered glucose metabolism, termed the Warburg effect. The Warburg effect or elevated conversion glucose to lactate by cancer cells has been a prevailing model of cancer metabolism. Since the 1980’s, genetic alterations of oncogenes and tumor suppressors have provided insights into tumorigenesis. However, whether metabolism contributes to tumorigenesis was highly debated. In 1997, we reported that the MYC oncogene product, the Myc transcription factor, regulates the lactate dehydrogenase A (LDHA)gene. Myc also activates many glycolytic enzymes, mitochondrial biogenesis, and glutamine metabolism by inducing glutaminase (GLS) and glutamine transporters, thereby providing not only ATP through the TCA cycle but also anapleurotic building blocks. Myc also induces biomass accumulation by stimulating ribosomal biogenesis. It stimulates the cell cycle machinery and DNA replication. Deregulated MYC in cancer results in enforced biomass accumulation, such that cell death occurs when bioenergetic demands exceed nutrient availability. In this regard, we have exploited this conceptual framework and targeted LDHA and GLS with small molecular inhibitors as proof-of- concept that altered cancer metabolism could be targeted for cancer therapy. Specifically, we documented that a drug-like inhibitor of LDHA could decreased tumor xenograft growth, providing evidence that metabolic therapy is feasible. We further found in a human Burkitt lymphoma model that Myc induces a genetic program that drives glutamine metabolism both under aerobic and hypoxic conditions. Inhibition of glutaminase, which converts glutamine to glutamate for its catabolism by the TCA cycle, by a drug-like molecule also diminished lymphoma xenograft growth in vivo. These studies indicate that targeting cancer cell metabolism could constitute a novel strategy to treat lymphoid neoplasias.

Disclosures:
Dang: Agios Pharmaceuticals, Inc.: Consultancy, Honoraria.
Genome-Wide Programming of Myeloid Cell Fates
Authors: Göttgens Berthold 
Published: October 2019

Abstract
Abstract SCI-29
Hematopoiesis represents one of the most tractable models of adult stem cell development and differentiation. Transcription factor (TF) proteins have long been recognized as major regulators of blood stem cell development as well as the subsequent differentiation into the multiple mature hematopoietic lineages. Seminal studies in multiple vertebrate model systems have identified specific TFs that control cell fate choices during myeloid differentiation (1). It remains largely unknown, however, how individual TFs are integrated into wider transcriptional regulatory networks, and how combinatorial TF interactions within these networks drive lineage specific gene expression programs. We are addressing these issues using two complementary approaches. First, we use a combination of transgenic reporter assays and network modeling approaches to reconstruct core transcription factor networks operating in early myeloid differentiation. Second, we employ genome-scale analysis of transcription factor binding sites for key hematopoietic regulators in both stem/progenitor cells and mature lineages (2,3). Integrated analysis of genome-scale datasets reveals previously unrecognized combinatorial interactions within core hematopoietic regulatory networks, which can be validated using both biochemical and mouse knockout approaches. Moreover, our studies also pinpoint novel candidate hematopoietic regulators, several of which we have validated using high throughput loss-of-function assays in zebrafish.

Disclosures:
No relevant conflicts of interest to declare.
TET2: Mechanism and Functional Consequences of Hydroxymethylation
Authors: Rao Anjana , Ko Myunggon , Pastor William , Huang Yun 
Published: October 2019

Abstract
Abstract SCI-32
TET family enzymes convert 5-methylcytosine (5mC) to 5-hydroxymethylcytosine (5hmC) in DNA. Somatic TET2 mutations are frequently observed in myeloid neoplasms in humans. Bone marrow samples from patients with mutant TET2, as well as some patients with wild type TET2, display low levels of 5hmC in genomic DNA compared to healthy controls. Measurement of 5hmC levels in myeloid malignancies may prove valuable as a diagnostic and prognostic tool to tailor therapies and assess responses to anticancer drugs. We have developed novel and specific approaches to profile the genomic localization of 5hmC and will describe their application to profiling 5hmC in mouse hematopoietic progenitor cells that express or lack Tet2.

Disclosures:
No relevant conflicts of interest to declare.
Strategies for Genome Repair in Normal and Leukemic Stem Cells
Authors: Passegué Emmanuelle 
Published: October 2019

Abstract SCI-41
Blood development is organized hierarchically, starting with a rare but well-defined population of hematopoietic stem cells (HSC) that give rise to a series of committed progenitors and mature cells with exclusive functional and immunophenotypic properties. HSC are the only cells within the hematopoietic system that self-renew for life, whereas other hematopoietic cells are short-lived and committed to the transient production of mature blood cells. Under steady-state conditions, HSC are a largely quiescent, slowly cycling cell population that, in response to environmental cues, is capable of dramatic expansion and contraction to ensure proper homeostatic replacement of all blood cells. While considerable work has deciphered the molecular networks controlling HSC activity, still little is known about how these mechanisms are integrated at the cellular level to ensure life-long maintenance of a functional HSC compartment. HSC reside in hypoxic niches in the bone marrow microenvironment, and are mostly kept quiescent in order to minimize stress and the potential for damage associated with cellular respiration and cell division. Recently, we have shown that HSC can also engage specialized response mechanisms that protect them from the killing effect of environmental stresses such as ionizing radiation (IR). We demonstrated that long-lived HSC, in contrast to short-lived myeloid progenitors, have enhanced expression of pro-survival members of the bcl2 gene family and robust induction of p53-mediated DNA damage response, which ensures their specific survival and repair following IR exposure. We reasoned that HSC have other unique protective features, which allow them to contend with a variety of cellular insults and damaged cellular components while maintaining their lifelong functionality and genomic integrity. We will present some of our recent findings on the fundamental mechanisms of stress-response that preserve HSC fitness during periods of metabolic stress, and allow for survival and repair following environmental stress associated with DNA damaging agents. It is now clear that oncogenic insults in diseases such as myeloproliferative neoplasms (MPN) can transform HSC and dramatically alter their biological functions leading to the emergence of leukemia-initiating stem cells (LSC), which are left untouched by most current therapies and can thereby mediate disease relapse. We will also discuss how transformed HSC may take advantage of some deregulated features of these normal stress-response mechanisms to escape therapeutic killing.

Disclosures:
No relevant conflicts of interest to declare.
Th17 Cells and Associated Cytokines in Inflammation and Cancer
Authors: Dong Chen 
Published: October 2019

Abstract
Abstract SCI-5
CD4+ T cells, upon activation, differentiate into cytokine-producing effector helper T (TH) cells. In addition to TH1 and TH2 lineage cells, additional TH subsets, including TH17 and T follicular helper (Tfh) cells have been identified. TH17 cells produce IL-17, IL-17F, IL-21, and IL-22 and mediate tissue inflammation. TH17 cells play protective or pathogenic roles in cancer, depending on the context. On the other hand, Tfh cells produce IL-21 and regulate germinal center reactions. Tfh cells may play a role in some forms of lymphoma. I will discuss on the regulation and function of these two subsets of T cells in the context of cancer.

Disclosures:
Dong: Ono: Consultancy; Tempero: Consultancy; Genentech: Honoraria; GSK: Consultancy; AnaptysBio: Consultancy.