bluuebunny/crossref_metadata_2025_split huggingface dataset:
Serial Imaging of Luciferase Positive Bone Marrow Stromal Cell Migration to Form Radiation Pulmonary Fibrosis
Authors: Goff Julie P. , Dixon Tracy M. , Epperly Michael W. , Sprachman Melissa , Wipf Peter , Zhang Xichen , Greenberger Joel S. 
Published: October 2019

Abstract
Abstract 4734
Thoracic irradiation of C57BL/6 mice leads to an acute reaction phase in the lungs characterized by increased cytokine production and inflammation days 1–14 post irradiation. This is followed by a latent period where inflammation, histologic appearance and cytokine response returns to control levels. The late reaction phase occurs 100+ days post irradiation and is characterized by organizing alveolitis/fibrosis and involves migration of marrow origin macrophages and proliferating mesenchymal stem cells (fibroblasts), a subpopulation which migrates from marrow to the lungs. To quantitate migration in real time, thoracic irradiated mice were either made chimeric for luciferase positive (luc+) whole marrow or were injected with cells from a positive luc+ bone marrow stromal cell line and serially imaged at day 7, 60 or 120 using an IVIS.. 200 Optical Imaging System. As a control for migration to the lung, another group of mice received 20Gy to the right hind leg and 1.5 ×106 luc+ bone marrow stromal cells i.p. Imaging of chimeric mice revealed luc+ cell lung migration only after day 120. C57BL/6NTac female mice that received 20Gy thoracic irradiation followed by an i.p. injection 1.5 × 106 luc+ positive bone marrow stromal cells revealed no migration of luc+ cells to the lungs at day 7 or day 60. Furthermore there was no migration to 20Gy irradiated leg at any timepoint. In marked contrast, at the time of the late reaction phase, at day 100, fibrosis was revealed as an increase in luc+ cell migration in lungs. The lung fibrosis model in C57BL/6 mice combined with live imaging allows sequential measurement of the effect of agents which may alter migration of bone marrow cells that contribute to radiation pulmonary fibrosis.

Disclosures:
No relevant conflicts of interest to declare.
Hematopoietic Stem Cells Have an Intrinsic Expansion Limit
Authors: Rojas-Sutterlin Shanti , Haman André , Hoang Trang 
Published: October 2019

Abstract
Abstract 4749
Hematopoietic stem cell (HSC) transplantation is the first successful cellular therapy and remains the only treatment providing long-term cure in acute myeloblastic leukemia. At the apex of the hematopoietic system, quiescent HSCs are spared by chemotherapeutic treatments that target proliferating cells and therefore can regenerate the entire blood system of a patient after drug exposure. Nevertheless, the consequence of repeated chemotherapy regimen on HSC function remains to be clarified. We previously showed that Scl/Tal1 gene dosage regulates HSC quiescence and functions when transplanted at limiting dilutions (Lacombe et al., 2010). In the present study, we investigate how massive expansion in vivo influences stem cell functions. To address this question, we optimized a protocol based on 5-fluorouracil (5-FU), an antimetabolite that has been used to treat colon, rectum, and head and neck cancers. In addition, we used Scl+/− mice to address the role of Scl in controlling HSCs expansion post-5-FU. We show that within 7 days following 5-FU treatment, HSCs exit quiescence and enter the cell cycle. To deplete cycling HSCs, we injected a second dose of 5-FU and showed that the stem cell pool was disseminated. Nonetheless, the remaining HSCs proliferated extensively to re-establish the HSC pool, which was twice larger than that of untreated mice. At this point, most HSCs have exited the cell cycle and were back to quiescence. Despite a near normal stem cell pool size and a quiescent status, HSCs from these 5-FU treated mice could not compete against untreated cells to regenerate the host in transplantation assays. Furthermore, we show that this extensive proliferation in vivo severely impaired the clonal expansion of individual HSC as measured by the mean activity of stem cell (MAS). Our results demonstrate that HSCs lose their competitive potential after two 5-FU treatments, suggesting that HSCs have an intrinsic expansion limit beyond which their regenerative potential is impaired. In addition, Scl is haplodeficient for cell cycle entry and cell division but Scl gene dosage does not affect this expansion limit. Therefore, our data dissociate the control of HSC expansion under extensive proliferative stress from cell cycle control during steady state. We surmise that chemotherapy regimen based on repeated administration of 5-FU or other antimetabolites are likely to severely impair long-term stem cell functions.

Disclosures:
No relevant conflicts of interest to declare.
Primary CNS Lymphoma in a HIV Negative Population: A Review of Clinicopathologic Characteristics, Therapy, and Outcomes of 59 Patients
Authors: Jaglal Michael V. , Shah Bijal D. , Cultrera Jennifer L. , Sotomayor Eduardo M. , Tomblyn Michael B , Bello Celeste M. , Sokol Lubomir 
Published: October 2019

Abstract
Abstract 4858

Background
Primary central nervous system lymphoma (PCNSL) is a rare aggressive variant of diffuse large B cell lymphoma (DLBCL) with a poor prognosis. Optimal therapeutic strategies have not been defined yet in primary CNS lymphoma. High dose Methotrexate (HD MTX) is an effective chemotherapeutic agent with superior outcomes compared to historical studies using whole brain radiation therapy (WBRT). The combination of HD MTX with WBRT showed improved response rates compared to chemotherapy (CT) alone, but was associated with greater risk of neurotoxicity in patients >60 years old.

Purpose
To review clinicopathologic characteristics, therapy and outcomes of 59 patients with primary CNS DLBCL without HIV.

Methods
This was a single center retrospective review of pts with confirmed diagnosis of primary CNS DLBCL from 1999 to 2012. Data was extracted from the Moffitt Cancer Center (MCC) electronic records. Baseline demographics, clinical, pathological and treatment data were collected and analyzed. Patients were stratified according to their treatment regimens including HD MTX (3g/m2) alone or in combinations and WBRT alone or in combination with CT.
Descriptive statistical analyses were utilized. Chi square analysis and t- test were performed to compare categorical and continuous variables. Kaplan-Meier method was used to estimate OS and log rank test was used to compare the groups. All data was analyzed using SPSS version 19.0 statistical software.

Results
59 patients who underwent CT and/or WBRT for PCNSL between 1999 and 2012 were identified. The age range at diagnosis was 17–85 years with median age of 64. 35 of 59 patients (59%) were ≥ 60 years old. Male to female ratio was 1.27:1 (33:26). The median ECOG PS was 1. A majority of patients presented with motor deficits, 29 out of 59. The most common location of lymphoma was in the cerebral hemispheres. The median survival of the entire cohort was 37 months. 18 of 59 pts (25%) survived ≥ 60 months. In the cohort of pts that survived ≥ 60 months, a majority 16 of 18 (89%) received HD MTX. Patients treated with initial WBRT and chemotherapy revealed inferior overall survival (OS) compare to patients treated with induction chemotherapy alone (OS 37 months vs. 66 months) (p=0.011). Patients over the age of 60 had worse outcomes compared to patients who were less than the age of 60 (OS 33 months versus 70 months) (p=0.023).

Conclusions
HD MTX was the most frequently utilized CT regimen in the cohort of patients surviving > 60 months. Administering WBRT combined with chemotherapy was associated with worse outcomes in this retrospective analysis. Patients with primary CNS lymphoma who are older than 60 have worse outcomes in this retrospective analysis compared to patients younger than 60.

Disclosures:
Sokol: Celgene: Honoraria, Speakers Bureau.
An RNAi-Therapeutic Targeting Tmprss6, in Conjunction With Oral Chelator Therapy, Ameliorates Anemia and Additively Diminishes Secondary Iron Overload In a Mouse Model Of β-Thalassemia Intermedia
Authors: Schmidt Paul J , Racie Tim , Butler Jim S , Fitzgerald Kevin , Fleming Mark D 
Published: October 2019

Abstract
β-Thalassemias are a group of inherited blood disorders caused by loss of β-globin synthesis and are characterized by anemia, extramedullary hematopoiesis and ineffective erythropoiesis leading to secondary iron overload. Increased iron absorption is due to inappropriately low levels of the liver hormone, hepcidin (HAMP). The membrane serine protease Matriptase-2 (TMPRSS6) attenuates BMP-mediated HAMP induction by cleaving the BMP co-receptor, hemojuvelin (HJV). Previously, we demonstrated that an RNAi-therapeutic targeting Tmprss6 elevates hepcidin expression and reduces disease severity in the Hbbth3/+ mouse model of β-Thalassemia intermedia (Blood. 2013; 14;121(7):1200-8). To further interrogate the efficacy of this therapeutic approach, Hbbth3/+ animals were treated with a siRNA directed against Tmprss6 on a replete 50ppm iron diet, a low iron diet (3-5ppm iron) or a 50ppm iron diet containing deferiprone.
Systemic administration of an siRNA directed against Tmprss6 in the three diet conditions leads to significant inhibition of Tmprss6 mRNA in the livers of Hbbth3/+ mice with concomitant elevation in hepcidin expression. In correspondence with earlier studies, we demonstrate here that Tmprss6 silencing in animals under each of the three diet regimens leads to a significant improvement in the anemia of Hbbth3/+ mice as evidenced by increased total hemoglobin. Furthermore, hallmarks of ineffective erythropoiesis, including splenomegaly and reticulocytosis, were decreased in all Tmprss6 silenced Hbbth3/+ animals. If untreated, excessive iron loading in humans with β-Thalassemia leads to tissue iron deposition and eventual organ damage and failure. Importantly, here we demonstrate that the total body iron burden of Hbbth3/+ mice, as assessed by non-heme liver iron, is decreased by almost 30% in animals chelated with oral deferiprone and treated with Tmprss6 siRNA. A similar diminution of iron deposition is not evident in animals on a low iron diet or in mice fed deferiprone alone. Taken together, this data suggest that siRNA suppression of Tmprss6, in conjunction with chelation therapy, may provide an improved modality for treatment of the anemia and secondary iron loading seen in hemoglobinopathies such as β-Thalassemia.

Disclosures:
Racie: Alnylam Pharmaceutical, Inc: Employment. Butler:Alnylam Pharmaceutical, Inc: Employment. Fitzgerald:Alnylam: Employment. Fleming:Alnylam Pharmaceutical, Inc: Research Funding.
Novel NGS-Based Platforms For Molecular Diagnosis Of Severe Congenital Neutropenia
Authors: Racek Tomas , Puchalka Jacek , Kohistani Naschla , Klein Christoph 
Published: October 2019

Abstract
Congenital neutropenia (CN) is a heterogeneous disorder. More than 30 distinct genetic defects have been discovered in patients with genetic diseases associated with decreased numbers of peripheral neutrophil granulocytes. Currently, most molecular diagnostic laboratories use Sanger-based sequencing techniques to define disease-causing mutations in patients with CN. In approximately 50% of patients no known genetic disorder can be found.
To identify novel genes that can be causative for unexplained CN cases we embarked on next-generation whole-exome sequencing using SOLiD 5500™ and Ion Proton™ sequencers. Up to date we sequenced whole exomes of 49 families, in which children were diagnosed with CN. The fragment libraries were constructed using the SureSelect™ V4+UTRs System (Agilent) allowing us to target whole coding sequence and the majority of UTRs of human genome (approx. 71 Mb). The vast majority of the families were analysed in the “Trio” approach and suitable homozygous or compound heterozygous rare variations (frequency below 1%) in protein coding regions or in splice sites were chosen for further validations.
In seven cases mutations previously described as causative for neutropenia were identified including G6PC3, HAX1, and ELANE. Four other rare variants are currently being analysed for their potential to cause CN. In 35 patients, no plausible candidate could be identified so far.
When we assessed variants within the genes related to CN, our data revealed unequal coverage pattern over these genes. Around 10% of the exons were insufficiently covered (coverage of less than 10) to allow for reliable variant and genotype call. These facts limit the power of whole exome sequencing as a diagnostic tool, as mutations at the non-covered positions cannot be ruled out, and demonstrate the need of an alternative comprehensive approach.
We are currently assessing sensitivity and specificity of a robust, rapid, and cost-effective approach that comprehensively analyses the sequence of 34 CN-relevant genes. Our approach is based on enrichment of specific exon regions by amplification using custom made AmpliSeq™ (Life Technologies) panel. For 25 genes we are able to sequence coding region as well as both UTR sequences, for 9 genes sequencing is limited to coding regions. This approach will provide a reliable, quick, and inexpensive diagnostic strategy for CN patients which will be offered free-of-charge to patients worldwide, independent of ethnic, national, or financial considerations.

Disclosures:
No relevant conflicts of interest to declare.
AGM-Derived Endothelial Cells and Notch Ligands Provide Embryonic Hematopoietic Stem Cell-Supportive Niches In Vitro
Authors: Hadland Brandon K , Varnum-Finney Barbara , Moon Randall T , Butler Jason M , Rafii Shahin , Bernstein Irwin D. 
Published: October 2019

Abstract
Greater knowledge of embryonic niche signals regulating the establishment, maintenance, and expansion of hematopoietic stem cells (HSC) during development will be essential in deriving therapeutically useful HSC from pluripotent stem cells (PSC). To this end, we have used the murine embryo model to dissect components of embryonic hematopoietic microenvironments which are sufficient to support nascent HSC and their precursors in vitro. We demonstrate that Akt pathway-activated endothelial cells (ECs) derived from the AGM (aorta-gonad-mesonephros) region, a critical site of HSC emergence during development, can substantially increase short and long-term multilineage engraftment potential from isolated embryonic day 11 (E11) VE-Cadherin+/CD45+ AGM-derived hematopoietic cells by co-culture in vitro. Furthermore, preliminary experiments show that co-culture with AGM-ECs also promotes high level, multilineage engraftment capacity from VE-cadherin+/c-kit+ precursors isolated from younger embryos (E9-E10). These results suggest that endothelial cells from an embryonic HSC-producing niche provide signals sufficient to promote maturation of HSC from embryonic precursors and subsequently support early HSC expansion in vitro. Further dissection of required signals for embryonic HSC expansion identified a unique combination of Notch activation by immobilized Notch ligands, cytokines, and small molecule inhibition of the TGF-β pathway, which is sufficient to inhibit differentiation and enhance self-renewal of embryonic, definitive-stage hematopoietic precursors in vitro. Notably, these conditions significantly increased short and long-term, multilineage repopulating HSC from E11 VE-Cadherin+/CD45+, but not E9-10 VE-Cadherin+/c-kit+ AGM-derived hematopoietic cells, indicating AGM-ECs provide additional, yet to be identified, signals for HSC maturation from developmental precursors. These findings have important implications for dissecting critical niche signals for HSC formation and expansion that will be essential for addressing the elusive goal of deriving HSC from pluripotent precursors.

Disclosures:
Rafii: Angiocrine Bioscience: Founder Other.
Long Non-Coding RNAs As Components Of The MYC Network In B Cell Lymphoma
Authors: Kluiver Joost , Winkle Melanie , Tayari Mina , Terpstra Martijn , Kortman Gertrud , de Jong Debora , Sietzema Jantine , Kroesen Bart-Jan , Kok Klaas , van den Berg Anke 
Published: October 2019

Abstract
MYC is an important oncogenic transcription factor in B-cell lymphoma and high MYC expression is associated with aggressive behavior and poor clinical outcome. A large number of genes are regulated by MYC, several of which are shown to contribute to the MYC induced phenotype. Long non-coding (lnc)RNAs have recently emerged as a novel class of regulatory RNAs acting at the epigenetic, transcriptional or posttranscriptional level. Aberrant expression of several lncRNAs has already been implicated in various aspects of tumorigenesis. It is currently unknown to what extend MYC can regulate lncRNA expression and whether these lncRNAs contribute to the pathogenesis of B-cell lymphoma.
Using an inducible MYC B cell lymphoma model and a custom microarray we investigated the expression >10,000 lncRNA loci and identified 1,820 lncRNA probes that show a MYC regulated expression pattern. Of these, 355 responded already after 4h, indicating direct MYC regulation. To identify transcripts relevant to lymphoma pathogenesis, we determined if these 355 lncRNAs were differentially expressed between primary lymphoma cases with high and low MYC expression and in addition also between MYC-high lymphoma cell lines and normal germinal center B cells. This revealed an overlap of 176 lncRNAs that were MYC regulated, aberrantly expressed in B cell lymphomas and differentially expressed between MYC-high and MYC-low lymphomas. Differential expression patterns were validated by qRT-PCR. As a first indication for lncRNA function, we isolated RNA from nuclear and cytoplasmic fractions of B cell lymphoma cell lines and determined enrichment fold in comparison to RNA isolated from the total cell lysates. Approximately 40% of all lncRNA transcripts showed specific subcellular localization, 80% nuclear and 20% cytoplasmic enriched. 31 of the 176 candidate lncRNAs were enriched in a specific cellular fraction. Furthermore, we analyzed which lncRNAs are enriched in Argonaute 2 containing complexes as an indication for lncRNA-miRNA interaction. For ∼5% of all expressed lncRNAs we found evidence for miRNA-lncRNA interactions, including 8 of the 176 differentially expressed MYC-induced lncRNAs.
This study identified 176 MYC responsive lncRNAs that are deregulated in B cell lymphoma. To establish a definitive role in B cell lymphoma pathogenesis a further characterization is warranted.

Disclosures:
No relevant conflicts of interest to declare.
Dietary Determinants Of The White Blood Cell Count
Authors: Balkaransingh Pauline D , Wheeler David , Ning Yi , Helou Marieka A. , Massey Gita 
Published: October 2019

Abstract

Introduction
White blood cells are key components of the immune system. They defend the body against a host of diseases and infections. Various studies have explored the possible influence of dietary nutrients on the white blood cell (WBC) count. If it can be demonstrated that nutrients affect the WBC count, they may offer an inexpensive way of modulating the immune system and in turn, the body's ability to fight against disease and infection. The sample sizes of previous studies, however, have been relatively small. The results have therefore been variable and conflicting. The purpose of this study is to continue to explore the dietary determinants of the WBC count, using a database that allows for a larger sample size, the 2005-2006 National Health and Nutrition Health Survey (NHANES).

Methods
The main outcome variable, WBC count, was recoded to the normal range for ages ≥ 18 years, of 4-11(±2). This was done in order to account for extreme values; particularly high and low values secondary to disease or infection. The population means and 95% confidence intervals were obtained using the statistical weights for the key variables of interest (Table 1). Multiple linear regression and backward elimination were used to predict the final model (Table 2). Variables with p < 0.1 were kept in the model. SAS 9.3 was used for all statistical analyses.

Results
Associations for age, gender, race and Body Mass Index (BMI) and WBC count, were consistent with previous studies and were all statistically significant (p< 0.0001). It has been shown that more intense physical activity usually leads to an increase in the WBC count. In this study, however, more vigorous forms of activity were associated with a lower WBC count (p < 0.0001). Increases in dietary copper and iron were associated with a subsequent decrease in the WBC count (Table 2, p < 0.05). An increase in vitamin K was also associated with a decrease in WBC count, however, it was not statistically significant (Table 2, p > 0.05). Increases in vitamin B1 were associated with an increase in WBC count (p < 0.05).

Conclusion
The results of this study suggest that further prospective studies are needed to investigate the role of these nutrients as determinants of the WBC count.

Disclosures:
No relevant conflicts of interest to declare.
Survival In Advanced Diffuse Large B-Cell Lymphoma In Pre- and Post-Rituximab Eras In The United States: A Population Based Study
Authors: Bista Amir , Shafii Bahman , Shah Binay K. 
Published: October 2019

Abstract

Background
Rituximab was approved by FDA as first line agent for treatment of advanced diffuse large B-cell lymphoma (DLBCL) on February 2006. We conducted this population based study to determine if the results from clinical trials have translated into survival benefit in general population.

Methods
We selected patients with advanced diffuse large B cell lymphoma (Distant stage based on LRD Summary Staging 1998+ in SEER*Stat) from Surveillance, Epidemiology, and End Results (SEER) 18 database (1973-2010), and calculated relative survival rates for patients diagnosed from 2002-2005(pre-rituximab) and March 2006 to 2009 (post-rituximab). We used Z-test in the SEER*Stat to compare the relative survival rates in cohorts categorized by race (white, black and other), gender, and age groups (<60 and 60+). The “Cansurv” software, and specifically, Cox proportional hazard function were used to investigate the influence of age (<60 ; 60+ years), sex (male; female), race (white; black; others) and marital status (married; single; separated/divorced/widowed) on the relative survival.

Results
A total of 15627 patients with advanced stage DLBCL as the only primary cancer were identified. Patients were predominantly white of non-Hispanic, non-Spanish/non-Latino origin, males, belonging to the older age category and married. The median age at the diagnosis was 65 years. One-year relative survival in ‘Whites’ and ‘Others’ improved significantly in post-rituximab era compared to pre-rituximab era (64.80 ± 0.6% vs 61.3 ± 0.6%; p= 0.0002 and 64.5 ± 1.9% vs 54.9 ± 2.2%; p= 0.0011, respectively). Also, three year relative survival improved significantly in ‘Whites’ and ‘Others’ in post-rituximab era compared to pre-rituximab era (53.7 ± 0.7% vs 50.3 ± 0.7%; p= 0.0001 and 52.0 ± 2.3% vs 40.8 ± 2.3%; p= 0.0002, respectively). However, no significant improvement were observed in one-year and three-year relative survival in blacks, during post rituximab era comapared to pre-rituximab era. Interestingly, in pre-rituximab era, relative survival in blacks was comparable to that of whites, but it was significantly better than that of others (p<.05).
Although there was significant improvement in one- and three-year relative survival in young females, old males and old females, no significant improvement (p>.05) in survival was observed in the young males. Factors such as young age, female sex, non-Hispanic origin, white race, married status and post-rituximab era were associated with significantly better survival (p<.05).

Conclusion
The relative survival rates among young males and black patients with advanced diffuse large B cell lymphoma has not improved during post-rituximab era.

Disclosures:
No relevant conflicts of interest to declare.