bluuebunny/crossref_metadata_2025_split huggingface dataset:
Pregnancy-Induced Thrombocytopenia in Pseudo-Von Willebrand Disease
Authors: George Simi , Donahue Laura L. , Funk Dorothy M (Adcock) , Allen Steven L. 
Published: October 2019

Abstract
Abstract 4658
Platelet-type von Willebrand disease, or pseudo-VWD, is a rare congenital platelet disorder due to a mutation in the glycoprotein 1b alpha receptor. This results in increased binding of normal VWF to platelets, leading to platelet aggregation and intermittent thrombocytopenia. Although the mechanism is similar to type IIb VWD, this gain of function mutation affects the platelet, rather than plasma VWF.
We report a patient with pseudo-VWD who developed progressive thrombocytopenia during pregnancy. She was initially seen at 10 weeks gestation, when the platelet count was 143 × 109 /L (figure 1). As the pregnancy progressed, the platelet count decreased, accompanied by an inverse rise in VWF antigen and activity, factor VIII activity, and an increase in high molecular weight (HMW) multimers (figure 2). The VWF activity to antigen ratio remained abnormal at all times. At the time of delivery, the platelet count reached a nadir of 36 × 109/L. Post-partum, the platelet count normalized to 185 × 109/L with a concomitant fall in VWF antigen and activity levels. VWF multimer analysis after delivery showed an absence of high molecular weight multimers.FIGURE 1FIGURE 1. FIGURE 2Weeks GestationPlatelet count (×109 /L)VWF antigenVWF activityFactor VIIIPlatelet aggregationVWF multimers1014381%35%88%↑ ristocetin 0.6normal3654144%57%205%not donenormal+15 post-partum18555%19%49%not doneHMW multimers absent
The likely etiology of the progressive decrease in the platelet count is secondary to the physiologically stimulated rise in VWF levels during pregnancy. With rising VWF levels, the abnormal platelet GP1b alpha receptor binds VWF, resulting in platelet-VWF aggregates with subsequent clearance. Transfusion of normal platelets would be the only way to correct the abnormality. Our patient, a Jehovah's Witness, would not accept platelet transfusion if it became necessary. She underwent an uncomplicated cesarean section for obstetric indications.
Our patient's family was the subject of the landmark article initially describing this condition (Weiss HJ, Meyer D, Rabinowitz R, et al. Pseudo- von Willebrand's disease: An intrinsic platelet defect with aggregation by unmodified human factor VIII/von Willebrand factor and enhanced adsorption of its high-molecular-weight multimers. N Engl J Med 1982; 306:326-333). This patient is the 2-year old who was not tested when the family was initially evaluated. To our knowledge, this is the first reported case of pregnancy-associated thrombocytopenia in a patient with pseudo-VWD.

Disclosures:
No relevant conflicts of interest to declare.
Anti-Leukemic Activity Is Increased with Low Dose Busulfan and Irradiation: A Preclinical Model
Authors: Patel Pritesh R. , Mahmud Dolores , Zhang Wei , Zhou Tong , Mahmud Nadim , Weichselbaum Ralph R , Garcia Joe GN , Rondelli Damiano 
Published: October 2019

Abstract
Abstract 4696
Combinations of busulfan (Bu) and low dose total body irradiation are commonly used in conditioning regimens for allogeneic stem cell transplant in acute myeloid leukemia (AML). In this study we tested the in vitro effect of physiologic doses of Bu and ionizing radiation (IR) to address the question whether additive or synergistic antileukemic activity can be demonstrated in two leukemic cell lines, one sensitive (HL60) and one resistant (K562) to radiation.
Cells were treated for 24 hours with Bu at 0, 12.5, 25, 50, 100 or 200 μ g/ml, or IR at 1.5, 3 or 6 Gy, or a combination of both with 12.5 or 25 μ g/ml for 24 hours, followed by IR at 1.5 or 3Gy. Cells were then tested for proliferation by thymidine uptake, apoptosis by surface expression of annexin V and caspase 3, and in-vitro colony formation assays. Exposure to Bu or IR for 24, 48 and 72 hours induced a dose dependent inhibition of proliferation and colony formation in both cell lines. However, 3 or 6 Gy of IR were both very effective against HL60 cells whereas a significant increase in cytotoxicity in K562 cells was observed only by using 6Gy (p=0.008).
In-vitro treatment with either Bu or IR caused apoptotic cell death mainly in HL60 cells, but in a very limited amount in K562 cells. To test the effect of the combination of Bu and IR we utilized a low dose of Bu for 24 hours with low doses of radiation. HL60 cells were strongly inhibited by both agents when separate or combined. In contrast, treatment of the relatively radio-resistant K562 cells with low dose IR (3Gy) alone inhibited colony formation by 28%. The inhibitory effect increased to 84% when combined with low dose Bu.
In order to identify genes associated with a response to Bu, we formed a linear regression model controlled for cancer type and also leukemic cell lines using GI50 and Stanford cDNA array data from the NCI-60. Among 7 genes (p<0.001 and FDR ∼50%) that were identified for all cancers, CLK-2 was also shown to correlate with response to Bu in leukemic cell lines (p=0.047). We then tested how these significant genes performed in the Affymetric U133 plus 2.0 platform. Six other genes (ERC2, HCLS1, CD74, KCNH2, HLADQB2, CD53) were significantly associated with response to Bu (p<0.05).
In conclusion, we demonstrated an additive effect of Bu and low dose IR even on radioresistant leukemic cells. These findings prompted the design of a clinical trial with IV Bu and marrow irradiation in the conditioning regimen prior to allogeneic stem cell transplant for acute leukemia. Expression of genes correlating with response to Bu may represent a novel biomarker to predict the outcome of leukemic patients undergoing stem cell transplant and could lead to a personalized anti-leukemic therapy with Bu.

Disclosures:
No relevant conflicts of interest to declare.
Examining Knowledge and Information Seeking Behaviors towards Blood Transfusion Among Individuals with Cancer
Authors: Naim Ahmad B , Walls Danielle , Gollins Jan , Reynolds Chuck 
Published: October 2019

Abstract
Abstract 4762

Objective:
Examine knowledge and information seeking behaviors towards blood transfusions among individuals with metastatic or recurrent cancer.

Methods:
An online survey was conducted from a nationally representative patient panel in 1Q2011. All respondents were ≥18 years and diagnosed with cancer by a physician. Participants were asked about their blood transfusion history, information seeking behaviors, and knowledge about blood transfusion.

Results:
206 individuals responded to survey. 65% (n=133) were female and 25% (n=52) were over 55 years. 55% (n=114) were anemic and 45% (n=92) not anemic. 62% (n=128) had received blood transfusion, whereas, 38% (n=78) had no transfusions. Top two sources of information are doctor (85%) and Internet (78%). Among those previously transfused, 74% received right amount of information, whereas, 11% received too little information, and 15% received too much information. More than 90% of transfused indicated they knew the reasons for and benefits of getting a blood transfusion. Less than two-thirds received information about the costs. Over 65% of not transfused said that it is extremely important to know risks of infections, right blood type and screening techniques. Among previously transfused, about 70% agreed that they made an informed choice about receiving blood transfusions. Among the previously transfused, 78% agree that they knew the benefits compared with 68% not transfused. About 68% of transfused and not transfused agree they knew the risks of blood transfusion.

Conclusions:
Doctor's office and internet are primary sources of information about blood transfusions. Gaps in knowledge exist about benefits, risks, and costs of blood transfusions. A significant number feel that they need more information about blood transfusion to make an informed choice. Providers should consider adopting shared-decision making with their patients.

Disclosures:
Naim: Janssen Scientific Affairs, LLC: Employment. Walls:Janssen Scientific Affairs, LLC: Consultancy. Gollins:Janssen Scientific Affairs, LLC: Consultancy. Reynolds:Janssen Scientific Affairs, LLC: Consultancy.
EYA3 May Be Required for Globin Gene Expression in Erythroid Differentiation
Authors: Branco Diana Santos , Brugnerotto Ana Flavia , Lanaro Carolina , Fertrin Kleber Yotsumoto , Cunha Anderson Ferreira , Costa Fernando Ferreira 
Published: October 2019

Abstract
Abstract 4797

Background:
Extensive studies have led to a considerable understanding of the cellular and molecular control of haemoglobin production during red blood cell differentiation, however, identification of the genes expressed as part of the erythroid differentiation programme remains an important goal because of the insights that these data will bring to erythrocyte biology and disease. Previous results using SAGE identified 93 differentially-expressed genes during erythroid development. One of these genes, EYA3, a homologue gene of Eyes Absent 3 in Drosophila, is a transcription cofactor with intrinsic phosphatase activity and its expression was observed to be high at the end of CD34+ cell differentiation and in human bone marrow.

Aim:
To evaluate globin gene, fetal hemoglobin (HbF) expressions and apoptosis levels in the erythroleukemic K562 cell line after EYA3 gene silencing and induction with hemin.

Methods:
Four different cultures from human K562 cells (1×105cells/mL in DMEM, 10% FBS, penicillin/streptomycin, 5% CO2, 37°C) were transfected with control or EYA3 knockdown lentiviruses (MOI=2.5). After proliferation and selection of successfully transfected cells with puromicin (2.0 ug/mL), cells were treated with 30μM hemin and collected after 0, 24, 48, 72 and 96h for gene expression and flow cytometry analyses. EYA3, LXN, α, and g-gene expression was measured by qRT-PCR and normalized using the Genorm program. HbF expression and apoptosis were evaluated by flow cytometry.

Results:
Analysis of globin gene expression showed that α-globin gene was downregulated in EYA3 silenced K562 culture cells compared with the control culture in 72h after hemin addition (1.791±0.1735; 0.7404±0.1709, respectively, **P<0.001, n=4). g-globin gene expression was found to be downregulated in K562 EYA3 silenced culture after 24h (1.350±0.1296; 0.5285±0.1736, respectively, *P<0.05, n=4) and 72h (1.554±0.1042; 0.6889±0.1535, respectively, **P<0.001, n=4). HbF expression was found to be downregulated in the same culture compared to the control culture at 72h after hemin addition (3568±41.00; 1947±206.50, respectively, *P<0.05, n=4). Apoptosis levels were found increased in EYA3 silenced K562 culture cells compared with the control culture in 72h after hemin addition (4.7±0.10; 8.55±0.55, respectively, *P<0.001, n=4).

Conclusions:
Our results show that silencing EYA causes modifications in the expression pattern of α- and g-globin gene expression as well as in HbF expression pattern and apoptosis levels in a model of erythroid differentiation. Further studies should be performed in primary erythroid cell cultures using siRNA-based gene silencing and overexpression of these genes to determine how these genes are involved in the mechanisms of globin gene regulation.
Support by FAPESP, CNPq and INCTS

Disclosures:
No relevant conflicts of interest to declare.
Cell Fate Modulation by Microvesicles: Transcriptionally-Mediated and Long Term in Nature
Authors: Quesenberry Peter J 
Published: October 2019

Abstract
Abstract 4801
Cell-derived membrane enclosed vesicles containing mRNA, protein, microRNA, and DNA, can enter cells and effect a phenotype change. We have shown that lung-derived microvesicles enter marrow cells inducing them to express pulmonary epithelial cell-specific protein and mRNA, a variety of microRNA and to enhance their capacity to engraft in irradiated mice and express the phenotype of type II pneumocytes (Aliotta et al, Exp Hematol 38,2010). In the present studies using rat/mouse hybrid cultures and measuring species-specific mRNA, we have shown that immediately after co-culture of rat lung across from mouse marrow, mouse marrow cells express both rat and mouse specific surfactants B and C mRNA. However, when these cells are cultured in steel factor supported long-term culture, rat-specific mRNA disappears rapidly, while mouse-specific mRNA persists out to 12 weeks in liquid culture. Identical studies with rat liver cultured across from mouse marrow have shown early expression in mouse marrow of both rat and mouse albumin mRNA, but in long term in vitro culture, expression of albumin mRNA was mouse-specific. Thus, the major long-term persistent event is an alteration of transcription in the target marrow cells. In a similar fashion, marrow modulated by lung microvesicles in vitro and engrafted into lethally irradiated (950 cGy split dose) mice, evidences expression of pulmonary epithelial cell-specific mRNA or protein (surfactants) in host lung, marrow, thymus, spleen and liver 6 weeks after engraftment – the furthest time tested. These results indicate that microvesicle cell fate modulation is biologically meaningful and represents an important new mechanism for cell phenotype determination.

Disclosures:
No relevant conflicts of interest to declare.
A Wharton's Jelly Mesenchymal Stromal Cell Derived 3D Osteogenic Niche Allows for Cord Blood Stem Cell Expansion Using Cytokine-Free Culture Media
Authors: Aljitawi Omar S. , Keefe Peggy , Henderson Christianna , Akel Salem , Abhyankar Sunil , McGuirk Joseph , Detamore Michael , Garimella Rama 
Published: October 2019

Abstract
Abstract 4832

Introduction:
In prior experiments, a 3 dimensional (3D) osteogenic niche was developed using Wharton's jelly mesenchymal stromal cells (WJMSC) that allowed for successful attachment of CD34 + umbilical cord blood (UCB) stem cells. The 3D osteogenic scaffold resulted in 10 times expansion of total nucleated cells (TNCs), however, most of the expanded cells were CD34 negative possibly secondary to their differentiation attributed to the cytokine-rich media. Accordingly, it was proposed to use a cytokine-free media in expansion of CD34+UCB using the same 3D osteogenic structure.

Aims:
To assess the ex vivo expansion of CD34+UCB stem cells in a 3D osteogenic niche using cytokine-free culture media. The expansion in 3D conditions was compared to 2D and liquid-culture conditions.

Methods:
CD34+ selected UCB stem cells were expanded in 3D osteogenic scaffold using cytokine-free media for 2 weeks. In 2D conditions, CD34+ UCB stem cells were expanded over a monolayer of osteogenic differentiated WJMSCs using the same cytokine-free media. In liquid culture conditions, the CD34+ UCB stem cells were expanded in culture flask. Pre- and post-expanded TNC was determined and post expansion CD34 expression was assessed using flow cytometry. Colony forming unit (CFU) assays were used to compare the expanded population in the three culture conditions.

Results:
TNCs were expanded 26X in 3D, 265X in 2D, and only 2X in liquid culture conditions. CD34 expressing cells increased 2.4X in 3D, 10X in 2D, and decreased in liquid cultures. Relative to 3D, CFU expansion in 2D was increased 2.6X. In 3D, CFUs were almost exclusively CFU-GM (>93%), compared to 2D (74%). Absence of CFU-E was noticed in the expanded population that remained attached to the 3D osteogenic scaffold. On the other hand, low CFU-E percentage was noticed in the population of expanded cells that dettached from the 3D osteogenic scaffold.

Conclusions:
The experimental conditions lead to a modest but significant expansion of CD34+UCB stem cells in 3D as well as 2D conditions. Though the majority of the expanded population did not express CD34, they retained their colonogenic potential. Contrary to the expanded cells that detached from the 3D osteogenic scaffold, the cells that remained attached did not form CFU-E.

Disclosures:
No relevant conflicts of interest to declare.
Early Events of Innate Immune Response to Influenza Virus in a Complex Cellular Environment Revisited with Multi-Parametric Flow Cytometry
Authors: Olive Daniel , gondois-Rey Françoise , Le Thi Kieu Suong , Herrera Diana , Hirsch Ivan 
Published: October 2019

Abstract
Abstract 4931
The development of an innate immune response against viruses involves multiple cellular interactions implying infected cells and the cells endowed with innate immune functions. These cells have to act in unison to develop a potent response. Although multiple interactions sustained the development of the innate response, much of the knowledge on the function of human innate immunity comes from investigations of isolated cells or dual partner coculture. Our aim was to investigate the early interactions of influenza virus with innate immune cells in a complex cellular environment in order to deepen our knowledge on the dynamics of the innate immune response. Polychromatic flow cytometry has been the appropriate method of exploration. We designed panels of 12 markers to explore PBMC stimulated with live-influenza virus or a mix of TLR agonists mimicking the interactions of influenza virus with TLR7/8 and TLR4. The DC populations were identified as lineage−HLA-DR+ cells and we took advantage of the polychromatic cytometry to split the lineage markers into separate channels to analyze NK, T cells and monocytes simultaneously to DC. Intra-cellular expression of IFN-a, TNF-a, IL-12, IL-6, IFN-g, and CD107 were analyzed in a time-dependent fashion. Our results demonstrate the benefits of using complex cell culture for the understanding of innate immune response to virus. We show a new field of application for multi-parametric flow cytometry as a powerful tool of investigation of innate immune responses in complex cell systems, with a sensibility to detect subtle modifications comparable to isolated cell systems. The multiple time-dependent results allowed to approach innate immune activation as correlated dynamic events. Dynamic studies lead to re-visit the relative roles of infection and TLR induced-DC response in the early specific features of Flu induced innate activation.

Disclosures:
No relevant conflicts of interest to declare.
A Study of Outcomes of Diffuse Large B Cell Lymphoma in the Elderly
Authors: Hoffmann Marc , Ahmadi Tahamtan , Cobain Erin , Loren Alison , Frey Noelle V , Porter David , Stadtmauer Edward A. , Schuster Stephen J. , Nasta Sunita D. 
Published: October 2019

Abstract
Abstract 4950

Introduction:
Despite improvements in outcome using chemoimmunotherapy, treating elderly patients with diffuse large B cell lymphoma (DLBCL) remains challenging. Reliable risk stratification beyond age is lacking and consequently elderly patients are often treated with reduced-dose therapeutic regimen with palliative intent. We retrospectively analyzed outcomes of DLBCL patients aged 65 and older treated at the University of Pennsylvania.

Patients and method:
We identified 41 patients (pts) with diagnosis of DLBCL and age >65 years. Median age was 74 years (range: 65–86). Eight pts (20%) were age 80 or older. There were no differences in IPI, elevated LDH, or bcl-2 expression, whereas bcl-6 expression was more common in patients >80 years with 75% (6/8) vs. 27% (24/33) (p=0.01).
Overall, 31 pts were treated with R-CHOP, 2 pts with R-HyperCVAD and one pt with R-CVP. Seven patients with a median age of 77 years were considered too frail for standard chemotherapy and were treated with a “split R-CHOP” regimen consisting of: rituximab 375 mg/m2 day 1, cyclophosphamide 375 mg/m2 day 1 and 15, adriamycin 25 mg/m2 day 1 and 15, vincristine 1 mg day 1 and 15 and Prednisone 50 mg day 1–5 and day 15–19.

Results:
Overall, the complete remission (CR) rate was 56% with seven treatment related deaths (17%). For patients between 65 to 80 years of age and deemed fit for standard chemotherapy (n=28), CR rate was 57% with four treatment related deaths (14%). Among patients over 80 and deemed fit for standard chemotherapy (n=6), the CR rate was 50% with 2 treatment related deaths (33%). Frail patients treated with the “split R-CHOP regimen” (n=7) had a CR rate of 57% with one (14%) treatment related death.
For all patients, the median progression-free survival (PFS) was 1 year with a median overall survival (OS) of 2 years. The median PFS for pts between 65 and 80 years of age treated with standard chemotherapy was 16 months. Median PFS in pts >80 years of age treated with standard chemotherapy was 7 months, whereas median PFS in frail pts treated with “split R-CHOP regimen” was 11.7 months.

Conclusions:
Our data reveal interesting findings about elderly pts treated for DLBCL. PFS and OS in general are poor as has been reported by others. In our data set, pts>80 years considered fit for standard chemotherapy, had a shorter PFS then fit patients between 65 to 80 years.
Intriguingly, PFS among frail elderly patients treated with a “split R-CHOP” regimen appears to be superior to that of elderly deemed more robust who were treated with standard dose RCHOP-21. Though limited by small numbers, our institutional data suggest that frail patients can tolerate a modified R-CHOP regimen although survival remains short.

Disclosures:
No relevant conflicts of interest to declare.
Primary Central Nervous System Lymphoma (PCNSL) in Immunocompetent Patients: An Analysis of Prognostic Factors and Treatment Strategies
Authors: McFarland Daniel C , Gregory Stephanie A. , Li Yan , Deng Youping , Venugopal Parameswaran 
Published: October 2019

Abstract
Abstract 4955

Background:
The incidence of primary CNS lymphoma in immunocompetent patients is increasing for unknown reasons. Survival estimates from academic centers show an improvement in overall survival (OS) while community epidemiological data estimate minimal difference in OS in the last thirty years. Age greater than 60, poor performance status, elevated initial LDH, high CSF protein and involvement of deep regions of the brain have been associated with worse survival. Optimal dosing strategies and delivery of chemotherapy as well as the role of radiation therapy (RT) have yet to be clearly defined given the limitations of treatment related toxicity in patients achieving remissions.

Purpose:
To characterize the PCNSL patient population treated at a single institution by studying prognostic factors and assessing OS according to treatment strategies.

Methods:
45 cases of confirmed CNS Lymphoma diagnosed at Rush University Medical Center between January 2000 and November 2010 were studied. Demographic, clinical and treatment data were collected and reviewed. Hospital records and the Social Security Death Index were used to determine death dates. Survival data were compared based on treatment groups.

Results:
23 of the 45 cases met all inclusion criteria and had no evidence of previous systemic lymphoma or pre-morbid immunocompromised conditions (i.e. HIV). The age range at diagnosis was from 21 to 81 years old with a median age of 68. One patient had ocular involvement. Three patients had malignant cells in the CSF at diagnosis and 4 patients had multifocal tumors. Biopsies revealed one case of a cerebral T-cell lymphoma and 22 were diffuse large B-cell lymphoma (DLBCL). Elevated CSF protein above the mean (96.1 mg/dl) was associated with worse OS (p=0.02). Elevated LDH above the mean (227 U/L) was associated with worse OS (p= 0.039). Age, performance status and location of tumor within the brain (deep vs. superficial) did not correlate with survival patterns. Comparative survival analysis between treatment groups revealed no statistically significant difference (p=0.134). In addition to systemic chemotherapy, 15 patients received intrathecal chemotherapy

Conclusions:
The description of PCNSL in immunocompetent patients outside of the clinical-trial setting is necessary to inform prospective trial designs. This study supports the use of LDH and CSF protein in determining prognosis and treatment strategy. This analysis of conventional treatment is ongoing. Collaboration on gathering descriptive data for PCNSL in immunocompetent patients should be pursued.

Disclosures:
Gregory: Genentech:.