bluuebunny/crossref_metadata_2025_split huggingface dataset:
Selective HDAC6 Inhibition and Antileukemic Activity of the Novel HDAC Inhibitor ST80 in Myeloid Leukemia Cell Lines.
Authors: Hackanson Bjorn W. , Rimmele Leander , Jung Manfred , Lübbert Michael 
Published: October 2019

Abstract
Abstract 4808
The antileukemic activity of histone deacetylase inhibitors (HDACi) has driven the search for epigenetic drugs with higher substrate specificity. Most of the currently used HDACi target class I, II and IV, with some bearing a class preference but only very few being selective in inhibiting specific HDACs, such as the HDAC6-selective inhibitor tubacin. HDAC6 inhibition leads to acetylation of non-histone proteins such as Hsp90 and alpha-tubulin. As it was recently demonstrated that HDAC6 is overexpressed in AML cells, we sought to investigate the effect of selective HDAC6 inhibition by the novel hydroxamic acid derivate ST80 in myeloid cell lines.

Methods
ST80 (selective HDAC6 inhibitor, 30-fold higher enzyme inhibition as compared to HDAC1) and the non-selective novel hydroxamic acid derivates ST13 (pan-HDACi) and ST34 (pan-HDACi with preference for class I) had previously been tested in enzymatic assays for their HDAC inhibitory potential (Scott et al. Mol Cancer Res. 6:1250-8, 2008). Cell lines HL60, Kasumi-1, NB4, THP1, U937 and K562 were treated with 10 nM to 30 μM of these three drugs. Viability and growth inhibition were determined using trypan blue staining. Acetylation of histone H3, H4 and alpha-tubulin and HDAC6 expression were determined by Western blot and quantified by densitometry. Tubulin-selective acetylation was calculated as the ratio of tubulin acetylation vs. H4 acetylation (ac-tubulin:ac-H4 quotient).

Results
At 1 μm, ST80, ST13 and ST34 all acetylated tubulin (8-, 8- and 2-fold in NB4, 11-, 14- and 3.4-fold in HL60, respectively, after a 12 h treatment). However, the calculated ac-tubulin:ac-H4 ratio of ST80 was 15- and 8-fold higher in NB4 and 9-fold higher in HL60 when compared to ST13 and ST34. The inhibitory concentration (IC) 50 (cell growth) of ST80 in the six myeloid cell lines ranged from 2.8 μM (NB4) to 5.1 μM (Kasumi-1) after 48 h treatment. Median cell viability of all 6 cell lines at 48 h was 93.7 % (range: 87.0 - 96.8 %) at 1 μm and 90.3 % (65.7 - 95.7 %) at 5 μm of ST80. HDAC6 protein levels were strongly variable between cell lines; however, growth inhibition by ST80 was independent of HDAC6 expression.

Conclusion
The novel hydroxamic acid derivate ST80 shows antileukemic activity in myeloid cell lines at low micromolar concentrations, which affect cell viability only modestly. The degree of relative tubulin acetylation by ST80 indicates a selective HDAC6 inhibitory activity in myeloid leukemias. The favorable ratio of ST80 growth inhibition vs. cytotoxicity warrants combination studies of this drug with other compounds.

Disclosures:
No relevant conflicts of interest to declare.
Recipient T and B Lymphocytes Dictate the Severity of Antibody-Mediated Transfusion-Related Acute Lung Injury (TRALI).
Authors: Fung Yoke Lin , Kim Michael , Speck Edwin R , Freedman John , Semple John W. 
Published: October 2019

Abstract
Abstract 641
Transfusion-related acute lung injury (TRALI) is a serious complication of transfusion and has been ranked as the leading cause of transfusion-related fatalities. The majority (approx. 80%) of TRALI reactions are associated with and probably initiated by donor alloantibodies recognizing recipient granulocytes and/or human leukocytes antigens (HLA). Nonetheless, many details of the immunopathogenesis of TRALI are unknown. Previous studies have shown that a murine anti-MHC (H-2Kd) class I antibody (clone 34-1-2s) can induce TRALI in mice (Looney et al J Clin Invest. 116:1615,2006) and we utilized this model in an attempt to understand the role that recipient lymphocytes might play in TRALI reactions. BALB/c (H-2d) mice were injected iv with titrations of 34-1-2s and body temperature, morbidity/mortality, pulmonary granulocyte accumulation and serum levels of MIP-2 (the murine analog of human neutrophil chemokine IL8) were measured at various time points. Results showed that when BALB/c mice were administered 34-1-2s, a significant drop (N=20) in rectal body temperature indicating shock occurred within 30 min post-infusion, with evidence of recovery beginning at 1 hour post-infusion. Visible signs of breathing difficulty were apparent but there was no mortality observed. A significant granulocyte accumulation (N=20) within the lungs was also observed by 30 min post-infusion, which continued on to the end of the experiment (2 hours post-infusion). Serum MIP-2 levels were also significantly elevated concurrently with the granulocyte accumulation. To determine the role of recipient lymphocytes on these responses, BALB/c mice with severe combined immunodeficiency (SCID; lacking T and B lymphocytes) were infused with 34-1-2s. Compared with the BALB/c recipients, the decreases in rectal temperatures in the SCID mice were significantly greater (N=18) and there was a 66% mortality rate (N=18) with symptoms of severe respiratory distress and tracheal edema with 30 minutes after infusion of 34-1-2s. In addition, there was a significantly greater accumulation of pulmonary granulocytes in the SCID mice at lower doses of 34-1-2s and the antibody stimulated the production of significantly higher serum levels of MIP-2. These findings were also seen in 34-1-2s-infused SCID mice that were first depleted of natural killer cells suggesting that NK cells play no role in the enhanced severity of the antibody-mediated TRALI reaction. Taken together, these results suggest that recipient T and B lymphocytes have a protective role in suppressing antibody-mediated TRALI reactions perhaps by modulating recipient chemokine production. They identify a potentially new recipient mechanism that controls the severity of antibody-mediated TRALI.

Disclosures:
No relevant conflicts of interest to declare.
Reducing the Risk for Transplant Related Mortality After Allogeneic Hematopoietic Cell Transplantation: How Much Progress Has Been Made?.
Authors: Horan John , Logan Brent R. , Agovi Manza-A. , Lazarus Hillard M , Bacigalupo Andrea , Ballen Karen , Martino Rodrigo , Juckett Mark B. , Khoury Hanna J. , Bredeson Christopher , Gupta Vikas , Smith Franklin O. , Hale Gregory , Carabasi Matthew , McCarthy Philip L. , Rizzo J. Douglas , Pasquini Marcelo C. 
Published: October 2019

Abstract
Abstract 649

TRM is among the major challenges for the success of HCT. Over the past two decades advances in the prevention and treatment of the major sources of TRM; regimen related toxicity, graft-versus-host disease (GVHD) and infections, have been made. To estimate the combined effect of these advances over time, we assessed changes in the incidence of TRM from 1985 through 2004 in 5,972 patients younger than 50 years, who received bone marrow (BM) or peripheral blood (PB) HCT with myeloablative conditioning for AML in first (CR1) or second (CR2) complete remission reported to the Center for International Blood and Marrow Transplant Research (CIBMTR). The incidence of TRM was determined for four consecutive five-year periods for HLA-matched sibling donors (MRD) and the later three for unrelated donor (URD) separately by donor type and disease status at transplant. Cox proportional hazard regression models of TRM and overall survival outcomes were constructed with time periods as the main effect. Adjustments for patient and disease characteristics including age, performance score, coexistent diseases and cytogenetics were made in all multivariate models. Subgroup analyses were performed to account for the influence of major changes in transplant characteristics over time, i.e. GVHD prophylaxis, graft source and HLA matching. We observed a steady drop in the risk of TRM over time among patients in CR1 and CR2 receiving MRD transplants, which was associated with a significant reduction in risk of death (table below). Among URD recipients, TRM also improved with lower RR in 2000-2004 compared to earlier periods. No improvements in long term OS was observed in URD CR1 group. For patients in CR2, the RR for overall mortality was 0.74 (0.6-0.9, p=0.03) for 2000-2004 compared to 1990-1994. Subgroup analyses restricted to recipients of BM grafts, cyclosporine/methotrexate for all transplants and partially HLA-matched grafts for URD resulted in similar trends, suggesting that improvements in TRM were not solely related to utilization of PB, newer GVHD prophylaxis or better HLA matching. In conclusion, our results demonstrate lower risk for TRM over time in patients receiving HCT from MRD and URD for AML in CR1 and CR2. These reductions in risk of TRM have been accompanied by reduced risk of overall mortality in most groups of patients studied.

Disclosures:
No relevant conflicts of interest to declare.
Phase I Clinical Trial of the Anti-CD22 Immunotoxin CAT-8015 (HA22) for Pediatric Acute Lymphoblastic Leukemia (ALL).
Authors: Wayne Alan S. , Bhojwani Deepa , Jeha Sima , Stetler-Stevenson Maryalice , Pui Ching-Hon , McDevitt Jennifer , FitzGerald David J. , Kreitman Robert J , Kaucic Karen , Pastan Ira 
Published: October 2019

Abstract
Abstract 839
Although most children with ALL are cured, treatment is associated with multiple toxicities and outcome after relapse is poor. New therapies are needed to overcome drug resistance and reduce non-specific toxicities of chemotherapy. CD22 is a B-lineage differentiation antigen expressed on most B-lineage ALL blasts. The anti-CD22 immunotoxin RFB4(dsFv)-PE38 CAT-3888 (BL22) was recently shown to have clinical activity with an acceptable safety profile in children with ALL (Blood 2007;110:262a). We undertook a Phase I trial of a modified agent with higher CD22 binding affinity (CAT-8015 or HA22).

Methods:
Patients 6 months to 24 years of age with relapsed or refractory CD22 + B-lineage ALL or non-Hodgkin lymphoma were eligible for enrollment into this Phase I trial. CAT-8015 was administered at doses of 5, 10, 20, or 30 mcg/kg every-other-day for 6 doses every 21 days for up to 6 cycles. One patient was enrolled at each of the first 3 dose levels (5, 10, 20 mcg/kg) with standard 3+3 dose escalation commencing at 30 mcg/kg. All patients received acetaminophen, ranitidine and diphenhydramine to mitigate infusion-related symptoms, and prophylaxis for central-nervous-system leukemia with intrathecal hydrocortisone, cytarabine and methotrexate. Patients at high risk for tumor lysis syndrome received standard prophylaxis.

Results:
Seven patients with ALL (6 precursor-B, 1 mature B-cell) 5 to 17 years of age (median, 10) were treated on the clinical trial. All patients had been heavily pre-treated and had baseline cytopenias due to active malignancy and thus were not evaluable for hematologic toxicities. The most common adverse events observed to date have been hyperbilirubinemia, transaminase elevations, hypoalbuminemia, elevated creatinine, febrile neutropenia, abdominal pain, pyrexia, hypertension, microscopic proteinuria, hemoglobinuria, hypoxia and pleural effusion. Two of 4 patients treated at 30 mcg/kg experienced Grade 3 or greater toxicity consistent with capillary leak: 1 with Grade 3 pleural effusion and hypoxia and 1 with Grade 4 vascular leak syndrome. All toxicities attributed to CAT-8015 were reversible. Clinical activity was demonstrated in 4 of 7 subjects. One patient treated at 10 mcg/kg had a complete remission by morphology and flow cytometry. Three patients met the protocol definition for hematologic activity (blood count improvement). One of these patients developed high-titer neutralizing antibodies. Two patients met the protocol definition for stable disease. The patient treated at the lowest dose level had progressive disease.

Conclusions:
CAT-8015 appears to be active against chemotherapy-refractory ALL. Strategies to predict and/or prevent vascular leak syndrome are currently being developed.

Disclosures:
No relevant conflicts of interest to declare.
Recognition of B2GPI by the Lipoprotein Receptors.
Authors: Beglova Natalia , Lee Chang-Jin , De Biasio Alfredo 
Published: October 2019

Abstract
Abstract 851
Lipoprotein receptors of the LDLR family serve as clearance receptors for beta2-glycoprotein-I (B2GPI) and as signaling receptors for the B2GPI/antibody complexes in patients with antiphospholipid syndrome (APS). B2GPI binds lipoprotein receptors through its domain V (B2GPI-DV) and the lipoprotein receptors use the LA modules to interact with B2GPI. The LA modules, which are about 40 residues each, are arranged in sequence to form the ligand-binding domains of the receptors. The LA modules have little sequence homology both within a single receptor and between different receptors, but adopt a very similar three dimensional structure. We have investigated the structural features of the contact interface in the B2GPI-DV/LA complexes and the characteristic features of the LA modules required for efficient binding to B2GPI. Using solution NMR spectroscopy, we have demonstrated that three different LA modules bind to the same site on B2GPI-DV and that the binding site for B2GPI-DV on these LA modules is centered on the residues involved in coordination of the calcium ion. The dissociation constants measured for different LA complexes are in the range from 1 mM to approximately 100 mM. The difference in binding affinity is attributed to the auxiliary contacts that augment the interactions of the residues at the core of the binding interface. We have solved the structure of the B2GPI-DV/LA4 complex by molecular docking guided by NMR-derived restraints and have validated the structure by four independent experimental observations based on mutagenesis, solution NMR spectroscopy, fluorescence polarization and intrinsic tryptophan fluorescence measurements. The structure identified three lysine residues of B2GPI-DV, Lys308, Lys317 and Lys282, as critical for binding the LA modules. Our data shows that not all the LA modules can bind B2GPI-DV and that those LA modules capable of binding B2GPI-DV will interact with it in a similar fashion as observed in the B2GPI-DV/LA4 complex. Whether an LA module is capable of binding B2GPI could be distinguished based on the primary sequence in the vicinity of the calcium-coordinating residues. According to the current hypothetical model, the binding of B2GPI to anionic phospholipids in the presence of APS-related antibodies is the first event leading to the pathology of APS. However, we have shown that the LA modules interfere with binding of B2GPI-DV to cardiolipin indicating that B2GPI-DV cannot simultaneously bind to anionic phospholipids and lipoprotein receptors.

Disclosures:
No relevant conflicts of interest to declare.
The Receptor Tyrosine Kinase RON Is Constitutively Activated in Acute Myeloid Leukemia and May Represent a New Therapeutic Target.
Authors: Fialin Camille , De Mas Veronique , Manenti Stephane , Payrastre Bernard , Roche Serge , Recher Christian 
Published: October 2019

Abstract
Abstract 994
Poster Board I-16
Recepteur d'Origine Nantais (RON) is a receptor tyrosine kinase (RTK) expressed mostly on epithelial cells but also in normal hematopoietic stem cells and macrophages. RON is closely related to c-Met in terms of homology and function. Its oncogenic properties have been documented in solid tumors leading to the clinical development of small molecules inhibitors. However, the role of RON in haematological malignancies and specifically in AML, has received little attention. Interestingly, a truncated form of RON lacking most of the RON receptor extracellular domain but retaining the whole transmembrane and intracellular domains has been described in the leukemic cell line KG1 (Bardella C et al, Cancer Res 64, 5154–5161, 2004). Since RTK and TK play a crucial role in leukemogenesis, we have assessed the expression and the role of RON in both AML cell lines and patient samples.
The expression of both full length (fl) and short form (sf) of RON was assessed in 71 AML samples by RT-PCR. fl-RON and sf-RON were found in 48/71 pts (68%) and 26/71 (37%) respectively, whereas both forms were undetectable in 17 samples (24%). RON expression was further confirmed at the protein level by western blot analysis in 20 samples. Conversely, c-Met was not expressed at the protein level in 10 cases tested. In leukemic cell lines, fl-RON and sf-RON were expressed both at the mRNA and protein levels in KG1 and KG1a but not in HL60 and U937. Immunoprecipitation analysis showed that fl-RON and sf-RON were constitutively phosphorylated on tyrosine in KG1/KG1a cells. This phosphorylation was fully inhibited by the dual c-Met/RON inhibitor, SU-11274. Cell signaling induced by RON has also been explored. Specific down regulation by si-RNA to RON induced a significant decrease of Lyn phosphorylation. Conversely, AKT phosphorylation was not influenced by RON down regulation. We then assessed the activity of SU-11274 on the proliferation and survival of KG1 (RON+) and HL60 (RON-). The proliferation of KG1 (IC50=3.5μM) but not HL60 (IC50 not reached at 10μM) was strongly inhibited in a time and dose dependant manner. This inhibition of proliferation was mostly due to apoptosis induction. Accordingly using clonogenic and cytotoxic assays, we show that only RON positive samples from AML patients responded to SU-11274 (n=8). Moreover, specific down regulation of RON by si-RNA inhibited the clonogenic properties of KG1 cells.
Altogether, these data demonstrate that the tyrosine kinase RON is aberrantly deregulated in AML cells, control cell proliferation and could represent a new target for the treatment of AML.

Disclosures:
No relevant conflicts of interest to declare.
Ribosome Dysfunction and Ineffective Hematopoiesis.
Authors: Ebert Benjamin L. 
Published: October 2019

Abstract
Abstract SCI-11
The 5q- syndrome and Diamond Blackfan Anemia are related on a molecular level by ribosome dysfunction. The 5q- syndrome is a distinct subtype of myelodysplastic syndrome (MDS) associated with isolated, interstitial deletions of the long arm of Chromosome 5. Diamond Blackfan Anemia is a rare congenital disorder associated with bone marrow failure, craniofacial abnormalities, and limb bud defects. The hematologic phenotype of both diseases includes a severe refractory anemia, macrocytosis, and a deficiency of erythroid precursors. Recent evidence indicates that this erythroid defect is caused by RPS14 deletion in the 5q- syndrome, and by mutation of RPS19 or other ribosomal genes in at least 50% of patients with Diamond Blackfan Anemia. In both diseases, deletion or mutation of one allele of a ribosomal protein leads to defects in pre-rRNA processing and defective production of mature ribosomes. In murine and zebrafish models, haploinsufficiency for ribosomal genes phenocopies the erythroid failure characteristic of the human disorders. While the mechanistic consequences of ribosomal dysfunction have not been fully elucidated, the p53 pathway appears to play a central role. MDM2, an E3 ubiquitin ligase that promotes the degradation of p53, binds to several ribosomal proteins including RPL11. Deficiency of RPS6, and perhaps other ribosomal proteins, causes an accumulation of free RPL11, that binds to MDM2, preventing MDM2 from interacting with p53, thereby leading to an accumulation of p53. Several other genes that are mutated in hematologic disorders are involved in ribosome biogenesis and function, including SBDS, mutated in Shwachman Diamond syndrome; DKC1, mutated in some cases of dyskaratosis congenita; and NPM1, mutated in acute myeloid leukemia and deleted in some cases of MDS. The manner in which each of these genes disrupt ribosome function and cause distinct clinical phenotypes is currently under investigation.

Disclosures
Ebert: GlaxoSmithKline: Research Funding.
Bone Spared Myeloma.
Authors: Mundy Gregory R. 
Published: October 2019

Abstract
Abstract SCI-6
After many years, the precise role of the bone microenvironment in the initiation and progression of myeloma is still quite poorly understood. In part, this may be because it has such a complex but important role in the natural history of myeloma. Myeloma cells have dramatic effects on bone, which are well known. These effects are mediated by the effects of myeloma cells on the cells responsible for the normal bone remodeling cycle. What is less well appreciated is the effects of bone and its cellular constituents on the myeloma cells. It is apparent that myeloma cells interact closely with osteoclasts, osteoblasts and also marrow stromal cells in the bone microenvironment and these interactions probably lead to a change in the phenotype of the myeloma cells, and the subsequent production of factors that are associated with the bone destruction which is so common in myeloma. It is also becoming more appreciated that the actual quality of bone and its rigidity and integrity may have important effects on the myeloma cells. The bone loss associated with myeloma and the interaction of myeloma cells with host cells in the bone microenvironment seem to be local effects rather than generalized effects. The bone manifestations of myeloma, even at the clinical level, are still not well understood. For example, it is extremely likely that generalized osteoporosis is far more common in myeloma than is appreciated and occurs even in the presence of lytic bone lesions, and that bone mass may continue to decline in patients in remission. In experimental preclinical models of myeloma, it is quite apparent that modulation of the bone microenvironment has quite dramatic effects on the behavior of the myeloma cells and so the notion if the bone disease of myeloma could be abrogated or removed, the natural history of the neoplastic disease may be very different. Experiments are underway in the preclinical models to investigate this intriguing possibility.

Disclosures
No relevant conflicts of interest to declare.
Regulation of White Cell Development.
Authors: Newburger Peter E. , Weissman Sherman M. 
Published: October 2019

Abstract
Abstract SCI-9
During hematopoiesis, determination of lineage and maturation to functional leukocytes depend upon cytokine-mediated changes in the transcriptional programs of progenitor and precursor cells. The classic binary branching tree of hematopoiesis now appears to be a more subtle series of gradual changes in differentiation probabilities, with competitive promotion and inhibition of lineage pathways by regulatory transcription factors such as (among others) PU.1, C/EBPα, GFI-1, EGR1/2, and NAB2 for the myeloid lineages and RUNX1, Notch-1, E2A, GATA-3, EBF, and PAX5 for lymphoid cell development. In addition, the recent discovery of regulatory non-coding RNAs (ncRNAs) has revealed another, important layer of control of hematopoiesis. The best studied members of this diverse group are the microRNAs, which often down-regulate multiple target transcripts. miRNAs involved in the regulation of myeloid development and function include miR-155, miR-223, and miR-17-19 cluster members. In addition, miR-9, miR-146a, miR-155, and miR-181a regulate the responses of immunocytes of the innate and acquired immune systems. Most recently, increasing numbers of long ncRNAs have been identified and found to regulate expression of other genes, both in cis and in trans. EGO (eosinophil granule ontogeny), a 500 base pair spliced, polyadenylated transcript regulates eosinophil granule protein gene expression. HOTAIRM1 (Hox antisense intergenic RNA, myeloid-1), a ∼500 base pair spliced polyadenylated ncRNA, affects neutrophil expression of both contiguous and distant HoxA cluster genes, as well as transcripts for CD18 integrin. Thus the control of white cell development depends not simply on a small number of key transcription factors, but rather on a complex network of interacting protein and ncRNA regulators of the transcriptional and translational programs of cell differentiation and function.

Disclosures
No relevant conflicts of interest to declare.