bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Novel Dimeric Inhibitor That Targets Dimeric ß2GPI In ß2GPI/Anti-ß2GPI Antibody Complexes.
Authors: Beglova Natalia , Kolyada Alexey , Lee Chang-Jin , De Biasio Alfredo 
Published: October 2019

Abstract
Abstract 1142
ß2GPI is a major antigen for autoantibodies associated with antiphospholipid syndrome, an autoimmune disease characterized by thrombosis and recurrent pregnancy loss. Only the dimeric form of ß2GPI generated by anti-ß2GPI antibodies is pathologically important, in contrast to monomeric ß2GPI which is abundant in plasma. Several cell-surface molecules including anionic phospholipids and ApoER2, a member of the lipoprotein receptor family, are implicated in the pathology of antiphospholipid syndrome.
We created a dimeric inhibitor, A1-A1, to selectively target ß2GPI in ß2GPI/antibody complexes. This inhibitor consists of two ligand-binding A1 modules from ApoER2 connected by a flexible linker. A1-A1 disrupts the binding of ß2GPI/antibody complexes with anionic phospholipids and lipoprotein receptors. We compared the potency of A1-A1 to monomeric A1 for inhibition of the binding of ß2GPI/antibody complexes to anionic phospholipids. We tested the inhibition of ß2GPI present in human serum, ß2GPI purified from human plasma and domain V of ß2GPI. We demonstrated that when ß2GPI/antibody complexes are formed, A1-A1 is more effective than A1 in inhibition of the ß2GPI binding to cardiolipin, regardless of the source of ß2GPI. Half maximum inhibition of ß2GPI in human serum in the presence of anti-ß2GPI antibodies was achieved at 9 μM of A1-A1 and 218 μM of A1. Similarly, ß2GPI purified from plasma was inhibited to 50% by 26 μM of A1-A1 and 191 μM of A1. Also, A1-A1 strongly inhibited the binding of dimerized domain V of ß2GPI to cardiolipin compared to monomeric A1. Concentration of A1-A1 and A1 at half maximum inhibition of the binding of ß2GPI domain V to cardiolipin in the presence of dimerization antibodies was 29 μM and 204 μM, respectively. In the absence of anti-ß2GPI antibodies, A1-A1 and A1 were equally inefficient in the inhibition of the binding of monomeric ß2GPI to cardiolipin. The concentration of inhibitor at half-maximum inhibition of the binding of ß2GPI in human serum to cardiolipin was 189 μM for A1-A1 and 176 μM for A1.
A novel inhibitor A1-A1 may be a starting point in the development of an effective therapeutic for antiphospholipid syndrome.

Disclosures:
No relevant conflicts of interest to declare.
Effects of Rabbit Anti-Mouse Thymocyte Globulin Treatment in a Splenocyte-Induced Model of Aplastic Anemia
Authors: Ruzek Melanie C , Phillips Kathleen , Richards Susan , Mamlouk Khalid , Williams John , Garman Richard 
Published: October 2019

Abstract
Abstract 1172
Acquired aplastic anemia is an immune-mediated disease where destruction of hematopoietic stem cells (HSCs) in the bone marrow results in severe and life-threatening pancytopenia. Thymoglobulin® is often used as immunosuppressive therapy in this disease with up to 80 percent of patients responding to a combination of Thymoglobulin and cyclosporine. In an effort to better understand the activities and mechanism of action of Thymoglobulin we developed a mouse model of immune-mediated aplastic anemia and evaluated a murine surrogate of Thymoglobulin®, rabbit anti-mouse thymocyte globulin (mATG) in this model. We modified a graft-versus-host (GVH)-induced model described in the literature (Bloom, et al., 2004) utilizing HSC-depleted spleen cells transferred from C57BL/6 into CByB6F1 mice instead of lymph node cell transfer. Our modified model shows a cell dose-dependent increase in pancytopenia and lethality. Mice receiving a high dose (100×106) of HSC-depleted splenocytes experienced severe pancytopenia and rapid death occurring around day 21 whereas mice receiving lower doses (70×106, 35×106 and 17×106) of cells showed progressively less pancytopenia and lethality as the dose of cells decreased. Histopathology also showed marked loss of hematopoietic progenitor cells in the bone marrow with little evidence of GVHD in other tissues. Prophylactic administration of mATG (25mg/kg, 2x, day 0 and 3) to mice given high doses of HSC-depleted splenocytes (100×106) resulted in a significant improvement in pancytopenia and survival (70%) in this model. Interestingly, therapeutic administration of mATG was more effective when given later relative to disease induction. Delivery of mATG (25mg/kg, 2x, three days apart) starting on day 3 showed some delay in disease progression (day 30 vs day 21) and mATG started on day 6 slightly increased survival (40%). However, mice receiving mATG starting on days 10 or 14 showed a much greater overall survival of 100% and 60%, respectively, with full rebound of hematopoietic cells in the blood to normal levels. The complete response observed with later mATG administration (day 10 or day 14) mimics the treatment and response of patients given Thymoglobulin®. In summary, we have established a novel model of HSC-depleted splenocyte induction of bone marrow failure in mice that is responsive to therapeutic ATG administration. Studies in this model will aid in further understanding the mechanism of ATG in aplastic anemia and may contribute to the development of potential new therapies.

Disclosures:
Ruzek: Genzyme: Employment. Phillips:Genzyme: Employment. Richards:Genzyme: Employment. Mamlouk:Genzyme: Employment. Williams:Genzyme: Employment. Garman:Genzyme: Employment.
Laser Transmyocardial Revascularization (TMLR) Combined with Autologous Bone Marrow Cells (ABMC) for Diffuse Coronary Disease (DCD). Results In 19 Cases
Authors: Alegre Adrian , Aguado Beatriz , Alfonso M. Perez , Muñoz C. , de Landázuri M. Ortiz , Sanchez F. Madrid , Cámara C. , García-Fernández N. , Bascones L. , Martinez S. , García-León N. , Roig M. , Duarte J. , Reyes G. 
Published: October 2019

Abstract
Abstract 1181

Introduction:
Stem cells (SC) show a potential of cellular therapy in diffuse coronary disease with controverted results. We present here an analysis of a group of patients treated with TMLR plus ABMC. (Caiber Program Advanced Therapy Proyect FIS 2009).

Patients and methods:
19 patients with DCD were included. Age 66 (range: 52–78). ABMC were obtained by aspiration and processed using HARVEST-TM system -TALEX (Harvest Technologies Inc. GMBH, Munich) to obtain 20 ml. Surgery consisted of anterior thoracotomy incision at the fourth and fifth intercostal space. Between 15–30 transmyocardial channels were realized with TLMR PHOENIX-TM (Cardiogenesis, Irvine, CA, USA) and CrystalFlex 1 mm optical fibre. 0.5–2 cc of MO were administered in each channel. Total number CMN were determined by automatic cell counter and flow cytometry and were typified specific cell populations such as CD34+ and CD133+.

Results:
The duration of stem cell collection was less than 30 minutes. All the procedure was performed on surgery in 1 step. Any complications were observed with this technique. Correlation between the cellular count and demographic data was not observed. After a follow up of 22 months (5-36) only one patient has died (20m) of DCD. Initial clinical results are satisfactory with a significant reduction in the functional grade of angina.

Comments and conclusions:
Our data show that TMLR plus ABMC is a safe method with clinical efficacy in patients with DCD that are not candidate to other therapy options. The described method offers efficiency and simplicity to obtain cells. More cases and follow-up are required, as well as later functional evaluations.

Disclosures:
Alegre: Celgene: Honoraria. Off Label Use: Lenalidomide is not approved for the treatment of smoldering multiple myeloma.
Isolation of G-CSF Mobilized Human Mesenchymal Stem Cells by Elutriation.
Authors: Jiang Yajuan , Leary Elizabeth , Yavanian Gregory , Anbazhagan Rajesh , Salerno Anthony , Marasco Wayne , Rodgerson Denis 
Published: October 2019

Abstract
Abstract 1183
Mesenchymal stem cells (MSCs) are a population of adult stem cells that constitute a promising source for therapeutic applications. MSCs exist in various adult tissues, such as bone marrow and umbilical cord blood. The most common method for MSC isolation relies on plastic adherence, however, this isolation procedure is hampered by the unpredictable influence of other co-cultured adhering cells. Phenotypic characterization of MSCs is usually carried out by FACS analysis of cell surface marker expression, but no unique marker has yet been identified. To circumvent these limitations, using granulocyte-colony stimulating factor (G-CSF) mobilized adult human peripheral blood (PB), we explored the use of Elutra® Cell Separation System (Caridian BCT) to isolate MSC enriched fractions based on their size and density.
A group of 28 donors were studied after mobilization for two consecutive day of G-CSF injection (480μg/day subcutaneously). Total nucleated cells (TNC) were collected by apheresis on the third day. An average of 6-fold increase in MSCs in the post-mobilized sample was observed. At least 1 million TNC in pre- and post-mobilized PB were analyzed by FACS for the presence of MSCs by cell surface staining with antibodies to CD105, CD90, CD73, CD31, CD45 and CD34. Circa 3 × 1010 TNC were loaded onto the Elutra to fractionate the apheresis product into several distinct cell fractions. Typically, the MSCs were enriched in 2 fractions, and yielded about 5–10% of the initial loaded cell number. Following culturing and passage, cells from these 2 fractions were grown on plastic with low contamination by other cell types. The cultured cells showed spindle-shaped morphology. Cells were further characterized by FACS analysis and shown to stained positive for mesenchymal specific markers CD90 and CD105; and negative for hematopoietic markers of CD45 and CD34. Furthermore, after cells from all fractions were plated for colony forming unit-fibroblast assay (CFU-F), no colony formation were observed from other than the 2 fractions.
In summary, we demonstrate that MSCs are mobilized into PB by G-CSF and can be collected by apheresis. Size-based Elutra fractionation of apheresis product in a closed system could provide a novel strategy for the large-scale isolation and expansion of human MSCs for autologous cell therapy *Dr. Marasco is a paid consultant for NeoStem and an equity holder.

Disclosures:
No relevant conflicts of interest to declare.
Arsenic Sulfide Binds c-CBL to Promote BCR-ABL Ubiquitination and Degradation In Chronic Myelogenous Leukemia.
Authors: Mao Jian-Hua , Sun Xiao-Yan , Liu Jian-Xiang , Zhang Qun-Ye , Liu Ping , Huang Qiu-Hua , Li Ke-Qin , Zhang Xiao-Wei , Chen Quan , Chen Zhu , Chen Sai-Juan 
Published: October 2019

Abstract
Abstract 1200
Using immunoprecipitation (IP)-2D-nano-HPLC-MALDI-MS-MS, we identified c-CBL in association with BCR-ABL in a multi-protein complex in K562 cells. In vitro ubiquitination and mutagenesis analyses show that c-CBL serves as a specific E3 ligase for ubiquitination of BCR-ABL at K1517. Arsenic sulfide (As4S4) treatment results in increased c-CBL protein level, which promotes ubiquitination and subsequent degradation of BCR-ABL and apoptosis of K562 cells. Elevated c-CBL is necessary and sufficient to recapitulate the effect of As4S4. Interestingly, arsenic directly binds the RING finger domain of c-CBL, inhibiting its self-ubiquitination and degradation, thus leading to accumulation of c-CBL. However, this interaction between As4S4 and c-CBL does not interfere with its E3 ligase activity towards BCR-ABL. Increased c-CBL protein and BCR-ABL degradation are also observed in vivo after As4S4 administration in BCR-ABL leukemia mice. These findings provide insights into the molecular mechanisms of arsenic and its potential therapeutic applications in CML.

Disclosures:
No relevant conflicts of interest to declare.
Distinct Roles for the NF-κB Pathway In Myeloid and Lymphoid Transformation and Leukemogenesis by BCR-ABL.
Authors: Hsieh Mo-Ying , Van Etten Richard A. 
Published: October 2019

Abstract
Abstract 1225
The BCR-ABL tyrosine kinase, product of the t(9;22) Ph chromosome, activates multiple signaling pathways in leukemic cells from patients with chronic myeloid leukemia (CML) and Ph+ B-cell acute lymphoblastic leukemia (B-ALL). Previous studies have shown that NF-κB is activated in BCR-ABL-expressing cell lines and contributes to transformation of primary B-lymphoid cells by BCR-ABL (Reuther et al., Genes Dev. 1998;12:968), but the mechanism of activation has not been defined (Kirchner et al., Exp. Hematol. 2003;31:504), and importance of NF-kB to myeloid and lymphoid leukemogenesis by BCR-ABL is unknown.
To interrogate the role of NF-κB in BCR-ABL-mediated transformation, we utilized a super-repressor mutant form of IκBα (IκBαSR), which has been used to block NF-κB nuclear localization and transactivation by constitutively sequestering NF-κB in the cytoplasm. Using retrovirus co-expressing BCR-ABL and IκBαSR, we found that IκBαSR blocked nuclear p65/RelA expression and inhibited the IL-3 independent growth of Ba/F3 cells and primary B-lymphoid cells transformed by BCR-ABL. The effect of NF-κB inhibition was primarily on proliferation rather than on cell survival, as there was no increase in apoptosis in cells expressing IκBαSR. When primary bone marrow cells were transduced and transplanted under conditions favoring induction of B-ALL or CML-like myeloproliferative neoplasm in recipient mice, co-expression of IκBαSR significantly attenuated disease development and prolonged survival of diseased mice. Molecular analysis of these leukemias demonstrated that NF-κB inhibition decreased the frequency of leukemia-initiating (“stem”) cells in the CML model, but not in the B-ALL model, and was associated with decreased expression of c-Myc, an NF-κB target. To clarify the mechanism of activation of NF-κB in BCR-ABL-expressing cells, we targeted two upstream kinases that negatively regulate IκBα, IKKα/IKK1 or IKKβ/IKK2. To accomplish this, we engineered retroviruses co-expressing BCR-ABL and kinase-inactive, dominant-negative mutants of IKK1 (IKK1KM) or IKK2 (IKK2KM). Co-expression of either IKK mutant inhibited both B-lymphoid transformation and leukemogenesis by BCR-ABL, as well as induction of CML-like MPN, with IKK1 inhibition more effective than IKK2.
Together, these results demonstrate that NF-κB is activated in part through the canonical IKK pathway in BCR-ABL-expressing leukemia cells, and that NF-κB signaling plays distinct roles in the pathogenesis of myeloid and lymphoid leukemias induced by BCR-ABL. In CML, NF-κB may play a role for in generation and/or maintenance of leukemic stem cells. These results validate IKKs as targets for therapy in Ph+ leukemias, and motivate the evaluation of small molecule IKK inhibitors in these diseases.

Disclosures:
No relevant conflicts of interest to declare.
Adherence to Treatment In Patients with Chronic Myelogenous Leukemia During a 10-Year Time Period: A Medical Record Review.
Authors: Yood Marianne Ulcickas , Oliveria Susan A , Hirji Ishan , Cziraky Mark , Davis Catherine 
Published: October 2019

Abstract
Abstract 1235

Introduction:
Over the past 10 years, oral treatment options for chronic myelogenous leukemia (CML) have provided patients autonomy over treatment administration. Low adherence to long-term maintenance therapy is a known challenge in a number of chronic diseases. The correlation between poor adherence and poor clinical outcomes has been previously demonstrated among CML patients in a clinical study setting. Few studies have evaluated adherence to CML treatments in a real-world setting and none have assessed treatment and adherence trends over time in a contemporary cohort of patients. Further, no studies have used medical record review to obtain comprehensive CML diagnosis and treatment information.

Method:
Medical and pharmacy claims from HealthCore Integrated Research Database™ (8.5 million covered US lives during study time period) were used to identify patients with CML (2001 – 2005). Review of medical records validated diagnosis and treatment exposure. Currently, additional data are being collected on this cohort to extend follow-up through 2010, and expand the number of CML patients to include data on the newer CML treatments. To measure adherence we used medication possession ratio (MPR) (number of days' supply of prescription divided by 365) and treatment interruptions (TI) (failure to refill prescription within 30 days of end of supply from previous prescription or clinician-directed discontinuation). For the current analysis, adherence to imatinib treatment was assessed for one year following treatment initiation.

Result:
In the initial study time frame (2001-2005), during which imatinib was indicated as first-line treatment, 216 CML patients treated with imatinib were identified. Mean age at imatinib initiation was 51 years and 42.6% were female. Fifty-one percent of patients had a MPR <85% for the one year period after imatinib initiation and 57% of patients experienced at least one TI during the follow up.

Conclusion:
Our preliminary findings indicate that adherence to imatinib treatment is low in CML patients for the time period studied. This is the first study to use medical record review to validate exposure and adherence by obtaining comprehensive imatinib treatment information on CML patients in a real-world setting and indicates that adherence may be even lower than previously reported. Adherence data from ongoing analyses which includes longer patient follow-up (up to 10 years) and second-generation tyrosine-kinase inhibitors will be presented.

Disclosures:
Hirji: Bristol-Myers Squibb: Employment. Davis:Bristol-Myers Squibb: Employment.
Involvement of Ets Factor Spi-B and Id2 In Waldenström's Macroglobulinemia
Authors: Zhou Yangsheng , Liu Xia , Xu Lian , Hunter Zachary , Sun Jenny , Yang Guang , Ciccarelli Bryan , Patterson Christopher , Treon Steven P. 
Published: October 2019

Abstract
Abstract 129
Waldenström's macroglobulinemia (WM) is an incurable disorder with a lymphoplasmacytic infiltrate in the bone marrow (BM) and IgM monoclonal gammopathy. WM tumor cells show variable differentiation, ranging from mature B-cells to plasma cells, which likely results from failure to fully undergo differentiation. Here we analyzed the expression of several genes involved in B cell differentiation by real time RT-PCR, such as Ets factors, the basic helix-loop-helix (bHLH) E proteins, as well as the inhibitors of DNA binding (Id) proteins which antagonize E protein activity. Comparison of bone marrow CD19+ B cells obtained from 12 untreated WM patients with 15 age-matched healthy donors showed that expression of the Ets factor Spi-B was increased four-fold, while Id2 was decreased three-fold. However, transcript levels of E proteins were similar between these two groups. Furthermore, along with differentiation of primary human CD19+ cells from peripheral blood into CD38+CD20− plasmablasts, Spi-B and Id2 expression levels were significantly decreased and increased, respectively. Ectopic expression of Spi-B in primary human CD19+ cells inhibited the plasma cell differentiation associated with decreased transcription levels of BLIMP1, XBP-1 spliced form, and IRF4. In addition, overexpression of Spi-B in BCWM.1 WM cells also resulted in repressed expression of BLIMP1, XBP-1 spliced form, and IRF4. Conversely, knocking down Spi-B in BCWM.1 WM cells increased IRF4 and Id2 expression. Importantly, in primary WM bone marrow CD19+ cells, knocking down of Spi-B induced CD38+CD20- plasma cell formation and increased expression of BLIMP1, XBP-1 spliced, IRF4, and Id2. Moreover, knocking down Spi-B in primary WM cells decreased Bcl-2 expression. Collectively, our results suggest that Spi-B overexpression plays an essential role in the pathogenesis of WM by repressing factors involved in plasma cell differentiation while promoting tumor cell survival through Bcl-2.

Disclosures:
No relevant conflicts of interest to declare.
Total Therapy 2 (TT2) for Multiple Myeloma (MM): Contributions to Survival Outcomes of Dosing of Thalidomide (T), Dexamethasone (D) and Interferon (I) Maintenance Components.
Authors: van Rhee Frits , Barlogie Bart , Szymonifka Jackie , Anaissie Elias J. , Nair Bijay , Waheed Sarah , Alsayed Yazan , Petty Nathan , Shaughnessy John , Hoering Antje , Crowley John 
Published: October 2019

Abstract
Abstract 1356

Background:
We have previously reported on the roles of VTD (bortezomib, thalidomide, dexamethasone) maintenance components in Total Therapy 3 (TT3), noting the important contributions of cumulative dosing of VTD components for overall survival (OS) and, especially, a detrimental effect of pre-mature drug discontinuation (PMDD) in the case of V (van Rhee et al: Blood 2010). Here we examine in a similar fashion the roles of T, D and I in TT2 toward OS.

Patients and Methods:
TT2 accrued a total of 668 patients, of whom 323 were randomized to the T arm and the remainder to the control arm. The cumulative dosing effects of T and D during induction, transplantation, consolidation and maintenance phases were evaluated while, in the case of I, its contribution in the maintenance phase only was examined. We also examined outcomes according to the number of induction and consolidation cycles applied prior to and after transplantation and according to the number and timeliness of administering both transplant interventions. In order to utilize the entire patient population, post-baseline treatment interventions and responses were treated as time-dependent variables. Multivariate analyses were performed to examine the collective impact on OS and EFS of baseline characteristics, treatment compliance and type of and time to salvage therapy.

Results:
Randomization to T prolonged both OS and EFS, as did timely completion of second transplant and first cycle of consolidation chemotherapy. During most protocol steps, cumulative dosing of both D and T benefited OS or EFS while pre-mature discontinuation of D shortened both OS and EFS; this was upheld in multivariate analyses, whether or not GEP data were included and also when time to salvage therapy was considered. D dosing prior to relapse was an independently favorable variable for post-relapse survival, which also was longer when disease progression did not occur within 2 years. D's dosing role was apparent only in the subset of patients presenting with lowest tertile gene expression levels of the glucocorticoid receptor gene, NR3C1.

Conclusion:
In addition to confirming well-established variables (beta-2-microgobulin, lactate dehydrogenase, cytogenetic abnormalities, gene expression profiling [GEP]-derived risk and molecular subgroups), randomization to T and timely applications of second transplantation consolidation and maintenance phases favorably affected outcomes. We conclude that D dosing, especially in low NR3C1-level myeloma, is an important treatment component that favorably affects global survival.

Disclosures:
No relevant conflicts of interest to declare.