bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Survival of Transplant Ineligible Multiple Myeloma Patients Treated with Bortezomib-Based Chemotherapy Is Affected By the Patient Fitness
Authors: Lee Ji Hyun , Kim Sung-Hyun , Huh Seok Jae , Jo Jae-Cheol , Shin Ho-Jin , Lee Won-Sik , Lee Yoojin , Lee Ho Sup 
Published: October 2019

Abstract
Background
Frail patients who are in a poor perfomance status and have serious comorbidities or abnormal laboratory results comprise about one-third of the transplant ineligible multiple myeloma (MM). These patients, most of the cases, are excluded from the clinical studies. By far, the outcome of the real world data according to the patients' fitness in transplant ineligible MM patients is limited.
Method
Four hundred and fifteen patients with MM who have been treated with bortezomib, melphalan and prednisone (VMP) as a first-line treatment were retrospectively analyzed. 4 factors included into the frailty scores were Age, ECOG PS (Eastern cooperative group performance status), eGFR (estimated glomerular filtration rate), and number of comorbidities (heart disease, lung disease, liver disease, cerebrovascular disease, diabetes mellitus). These factors were di- or trichotomized by the reference of previously published papers and the meaningful cut-offs in our data. The points given to each variables were after the following criteria: Age < 70 (point 0), 70-79 (point 1), ≥ 80 (point 2); ECOG PS 0-2 (point 0), 3-4 (point 2); eGFR ≥ 60 mL/min/1.73 m2 (point 0), < 60 mL/min/1.73 m2 (point 1); comorbidity 0-1 (point 0) and ≥ 2 (point 1). Patients were further classified by frailty scores: fit (points 0-2), unfit (points 3-5) and frail (point 6).
Result
The median follow-up duration was 26.72 months (range 0.37 - 180.20 months). All of the factors including the Age ≥ 80, ECOG PS 3-4, eGFR < 60 mL/min/1.73 m2, and comorbidity ≥ 2 statistically significant for overall survival (OS) by univariate analysis. In multivariate analysis, ECOG PS 3-4 and eGFR < 60 mL/min/1.73 m2 remained statistically significant for OS and Age ≥ 80 was marginally significant for OS (Table 1). The response rates by the fit, unfit, and frail were as follows: 77.5% (148/191), 71.4% (55/77), and 0% (0/2) (P= 0.028). The 3-year overall survival rates by the frailty were 66%, 41%, and 0% in the fit, unfit or frail patients respectively and showed statistically significant difference (Figure 1 and Table 2, P= 0.000). The comparison of the frailty score with international staging system (ISS) and revised ISS (R-ISS) revealed statistical significance of all of the prognostic systems affecting OS by the univariate analyses but R-ISS and frailty score remained significantly affecting OS.
Conclusion
Unfit and frail patients showed a significant decrease in OS compared to fit patients. But there was no difference between the unfit and fit patients in terms of response rate in patients with transplant ineligible MM patients who have received bortezomib-containing regimen. Treatment of the unfit and frail patients needs more caution on toxicity management and appropriate dosing schedules to improve the survival outcome.

Disclosures
No relevant conflicts of interest to declare.
Domain-Focused CRISPR-Cas9 Screen Identifies the E3 Ubiquitin Ligase Substrate Adaptor Protein SPOP as a Novel Repressor of Fetal Hemoglobin
Authors: Lan Xianjiang , Khandros Eugene , Grevet Jeremy D. , Peslak Scott A. , Bhardwaj Saurabh , Keller Cheryl A. , Giardine Belinda , Garcia Ben A. , Hardison Ross C. , Shi Junwei , Blobel Gerd A. 
Published: October 2019

Abstract
Elevated levels of fetal hemoglobin (HbF) are beneficial to patients with sickle cell disease (SCD) and b-thalassemia. The identification of pharmacologically controllable pathways that regulate HbF levels remains an important goal in the treatment of these disorders. BCL11A and LRF (ZBTB7A) are the two major transcriptional repressors of g-globin (fetal beta-type globin). However, since transcription factors are challenging to inhibit with small molecules, we aimed to identify novel HbF regulators that might be amenable to control by pharmacologic means.
Using an improved protein-domain based CRISPR-Cas9 screen recently developed in our laboratory (Grevet et al., Science 361:6399, 285-290), we screened for new HbF regulators using a gRNAs library targeting chromatin associated proteins including BTB domain containing factors. We identified SPOP, a substrate adaptor of the CUL3 ubiquitin ligase complex, as a novel inhibitor of g-globin expression. Depletion of SPOP strongly elevated the mRNA and protein levels of g-globin in both HUDEP-2 and primary human CD34+ derived erythroid cells. The effects of SPOP loss were remarkably selective as evidenced by RNA-seq and whole cell mass spectrometry. Importantly, SPOP depletion did not appear to impair cell growth or differentiation. Mechanistically, we demonstrated that the repressive role of SPOP on g-globin levels is dependent on the CUL3 ubiquitin ligase complex. Additionally, forced expression of a dominant negative form of SPOP (Y87N) unable to bind to its substrates strongly induced HbF production. We also found that SPOP depletion did not measurably alter LRF levels and modestly reduced BCL11A levels. Overexpression of BCL11A did not restore g-globin silencing in SPOP depleted cells, indicating that SPOP works largely independently of these two factors. Finally, we tested whether the effects of SPOP depletion can be augmented through the use of experimental pharmacologic HbF inducers, such as pomalidomide. In proof of principle experiments we found that SPOP depletion and pomalidomide treatment induced HbF production to levels higher than either treatment alone.
In sum, our work uncovered SPOP as a new regulator of HbF and suggests targeting SPOP alone or in combination with other HbF inducers may provide a novel therapeutic strategy for the treatment of hemoglobinopathies such as sickle cell disease and b-thalassemia.

Disclosures
No relevant conflicts of interest to declare.
Increased SERUM VON Willebrand Levels in Chronic Lymphocytic Leukemia (CLL) Patients Are Related to a Shorter Time to Treatment
Authors: Tryfou Thommais , Papaioannou Paraskevi , Repousis Panagiotis , Gkioka Annita Ioanna , Sachanas Sotirios , Bitsani Aikaterini , Dimou Maria , Maltezas Dimitrios , Koudouna Aspasia , Kopsaftopoulou Anastasia , Chamoulkou Konstantina , Grafakos Ioannis , Kokori Styliani , Tsantes Argyrios , Pangalis Gerasimos , Panayiotidis Panayiotis , Kyrtsonis Marie-Christine 
Published: October 2019

Abstract
CLL is a frequent and usually indolent disease that require only follow-up in about 2/3 of patients while the rest 1/3 of patients are symptomatic, have a worse outcome and need to be treated. The role of angiogenesis in CLL prognosis remains unclear. It has been shown that CLL plasma promotes von Willebrand factor (vWF) secretion, and expression in human umbilical vein endothelial cell cultures (HUVECs) [1]. Therefore, we investigated whether serum vWF levels in CLL patients at diagnosis could provide prognostic information.
Patients and methods: 33 CLL patients were studied of whom 55%, 32% and 13% were Binet staged A, B and C respectively. 45% never required treatment while 55% required treatment either at the time of diagnosis or during their follow-up. The time to first treatment (TFT) was 34 months. Serum vWF was measured by ELISA (R&D Quantiquine, duo Set) in frozen sera drawn at patients' diagnosis and in 10 healthy individuals (HI). Measurements were performed in duplicate according to the manufacturer's instructions. Statistical analysis was performed by conventional methods, using the SPSS v.22 software. P values below 0,05 were considered statistically significant.
Results: Serum vWF levels ranged from 178,5 to 14353,5 pg/ml (median 1028,5) in patients and from 374,6 to 1141,3 pg/ml (median 528,5) in HI, the difference being statistically significant (p< 0,05). Patients with Serum vWF levels above 528,5 pg/ml (median normal value) had a significantly shorter TFT than the others (p=0,01) as shown in figure. Otherwise, serum vWF levels correlated only with hypogammaglobulinemia (p=0,04).
Conclusion: Serum vWF levels were increased in CLL patients. Importantly, we showed for the first time, to our knowledge, that higher vWF levels correlated with a shorter TFT, suggesting that increased vWF levels reflect active neoangiogenesis that in turn, contributes to a more aggressive disease behavior.
[1] Shahidi M et al, Induction of endothelial cell proliferation and von Willebrand factor expression and secretion by leukemic plasma of patients with chronic lymphocytic leukemia before and after inhibition of NF-κB. Blood Coagul Fibrinolysis. 2016 Sep;27(6):711-6.
Figure. Figure.

Disclosures
No relevant conflicts of interest to declare.
Integrin Activity Reduces Immunogenic Cell Death By Inhibiting Cell Surface Presentation of ERp57 and Calreticulin
Authors: Liu Chi-Chao , Leclair Pascal , Pedari Foujan , Mahdis Monajemi , Sly Laura , Reid Gregor S. D. , Lim Chinten James 
Published: October 2019

Abstract
Calreticulin presentation on the cell surface is an important hallmark of immunogenic cell death (ICD), serving as the 'eat me' engulfment signal for professional phagocytes. ERp57 is an interacting and co-translocating protein with calreticulin, but unlike surface calreticulin, surface ERp57 is not known to be immunogenic. Since calreticulin interacts with α-integrins via the conserved GFFKR motif within the integrin cytoplasmic domain, we assessed whether integrin function can modulate cell surface calreticulin and ERp57 levels in ICD. When stimulated to engage integrin substrates, leukemic T-lymphoblasts treated with an ICD-inducer exhibited decreased surface calreticulin and ERp57 compared with cells under non-adherent conditions. The inhibitory effect on surface calreticulin and ERp57 was recapitulated for cells in suspension and treated with various agents that stimulate integrin activation, including Mn2+ and the LIBS antibodies 9EG7 and HUTS21. Similarly, cells expressing a mutant truncated α-integrin bearing only GFFKR as the cytosolic tail, also exhibited low surface calreticulin and ERp57 when treated with ICD-inducers under non-adherent conditions. Furthermore, integrin β1 null cells with overall reduced α-integrin expression exhibited enhanced surface calreticulin and ERp57, consistent with an inhibitory role for integrins in ICD. We generated calreticulin or ERp57 null strains by CRISPR-Cas9, and show that ICD-induced surface calreticulin is ERp57-dependent, while surface ERp57 is not calreticulin-dependent. Using permeabilization techniques that distinguish between cytosolic and ER staining, we found that ICD-inducers promoted the accumulation of cytosolic calreticulin and ERp57 that originated from the ER. In similar fashion, ER to cytosolic trafficking for calreticulin is ERp57-dependent, while for ERp57, it is not calreticulin-dependent. We also showed that integrin-mediated inhibition of surface calreticulin was due to reduced cytosolic to surface translocation coupled with normal ER-cytosolic release, suggesting that activated integrins acts to sequester calreticulin within the cytosol. T-lymphoblasts co-treated with an ICD-inducer and integrin activators exhibited reduced phagocytosis by macrophages when compared with treatment with only ICD-inducer. Our study reveals a previously uncharacterized function of integrins as negative regulators of immunogenic cell death by suppressing the presentation of cell surface calreticulin.

Disclosures
No relevant conflicts of interest to declare.
Cost-Effective Use of White Blood Cell Growth Factors in the Veterans Administration
Authors: Knopf Kevin , Hrureshky William , Love Bryan , Norris Leann , Bennett Charles L. 
Published: October 2019

Abstract
The use of granulocyte colony-stimulating (G-CSF) factors to avoid chemotherapy induced neutropenia is a major cost driver in clinical oncology. Short acting formulations, while less convenient, are 39% the cost of the longer formulation (Pegfilgrastim). We analyzed the use of G-CSF vs. Neulasta in the Veterans Administration (VA) health care system as currently practiced by clinical judgement and analyzed this economically. A survey of 23 sites within the VA on the relative frequency of use of filgrastim, Tbo-filgrastim, Filgrastim-sndz and Pegfilgrastim was conducted. Cost was estimated using 340B pricing.
A maximally cost-effecient strategy of 100% Tbo-filgrastim would result in a cost of $62,336 per 100 patient episodes. Tbo-filgrastim was the preferred treatment in 18 of 23 sites surveyed. Cost results ranged from a minimum of $62,336 (4 of 23 sites) to a maximum of $201,356 (2 of 23 sites) with a mean cost of $99,080. The VA was overall 73% efficient, and therefore highly cost-effective, in minimizing use of Pegfilgrasim.
As in Europe cost-efficiency favors the use of Tbo-filgrastim over Pegfilgrastim; further cost-saving opportunities exist with the following strategies : (1) avoid G-CSF use where there is no convincing evidence of efficacy (i.e. low risk patients, treatment of neutropenia rather than prophylaxis) (2) use daily filgrastim in lieu of pegfilgrastim whenever possible (3) use Tbo-filgrastrim rather than filgrastrimn (4) continue to examine strategies using 2 or 4 day courses of G-CSf as opposed to 8 days which would cut cost an additional 50 to 75%.
Survey data showed that uptake of biosimilar G-CSF in the VA system has been extremely rapid. All sites are using biosimilar GCSF for all new patients; 6 of 23 sites were comfortable shifting current patients on branded G-CSF to the biosimilar. Survey data shows an overwhelming use of filgrastrim compared to pegfilgrastim in the VA system, in contradistinction to the rest of the United States - resulting in an efficiency of 73% (i.e. only 27% of patients are being given pegfilgrastrim). This represents a great savings to the VA system which can be emulated in other sites in the United States. Switching to a Tbo-filgrastim, although heralded by some as cost savings, only engendered a cost savings of 2.2% - small compared to other clinical changes.

Disclosures
No relevant conflicts of interest to declare.
Symptom Duration Is Positively Correlated with Factor XIIIa Activity in Acute Pulmonary Embolism
Authors: Brailovsky Yevgeniy , Hoppensteadt Debra , Iqbal Omer , Simpson Kevin , Mcclane Nathan , Fareed Jawed , Darki Amir 
Published: October 2019

Abstract
Background
Acute pulmonary embolism (PE) patients are at variable risk of morbidity, mortality, and response to therapy. Patients often present at various time points from the symptom onset. Several factors may shed light into the state of endogenous thrombotic and fibrinolytic system at the time of presentation. Factor XIIIa plays a critical role in clot stabilization and may impact clot dissolution. Relation of Factor XIIIa activity and symptom duration is not known.
Methods
We prospectively collected blood samples from patients evaluated by Pulmonary Embolism Response Team at a tertiary care center. Blood was centrifuged, and plasma collected for analysis. We used ELISA method utilizing a commercially available kit from Hyphen, BioMed (Neuville-sur-Oise France), specific for measurement of Factor XIIIa, D-dimer, and Pro-TAFI antigens.
Baseline clinical characteristics were collected from electronic medical record. Symptom duration was gathered from patient subjective assessment. Additional workup included radiographic and echocardiographic evaluation.
We performed correlation analysis to test the association between symptom duration and Factor XIIIa activity, D-dimer, and Pro-TAFI antigen. Additionally we performed linear regression analyses to quantify the degree of association of symptom duration and Factor XIIIa activity.
Results
±±9.8, 8 patients were treated with catheter directed thrombolysis, while the rest were treated with anticoagulation alone. Symptom duration was positively correlated with Factor XIIIa activity (r2=0.227). More so, for every one day increase in symptom duration the Factor XIIIa activity was increased by 2.2%. (p=0.014). We demonstrated no correlation between symptom duration and D-Dimer (p=0.58) or symptom duration and Pro-TAFI antigen (p=0.84).
Conclusion
In patients with acute PE, symptom duration positively correlated with Factor XIIIa activity, for every one day increase in symptom duration the Factor XIIIa activity was increased by 2.2%. Future studies are needed to ascertain the impact of Factor XIIIa activity and clot dissolution as well as functional outcomes.
Figure. Figure.

Disclosures
No relevant conflicts of interest to declare.
Second Transplants for Relapsed Acute Leukemia: Real World Experience from Argentina
Authors: Bet Lucia Antonella , Berro Mariano , Paganini Maria Ines , Basquiera Ana , Palmer Silvina , Garcia Juan Jose , Real Juan Jose , Martinez Rolon Juliana , Castro Martin , Kusminsky Gustavo 
Published: October 2019

Abstract
INTRODUCTION
Acute leukemia relapsing after allogeneic stem cell transplantation (AlloSCT) is usually associated with dismal prognosis. Treatment options in this population remain limited. We evaluated the outcomes of second AlloSCT in patients with acute leukemia relapsed after previous AlloSCT in centers from Grupo Argentino de Trasplante de Médula Ósea (GATMO)
METHODS AND RESULTS
We carried out a retrospective analysis in 21 adult patients who received a second transplant as a salvage treatment for relapsed acute leukemia. Main outcomes were overall survival (OS) and leukemia free survival (LFS). Variables evaluated were re-induction chemotherapy regimen, conditioning regimen, donor type, early immunosuppression tapering, and the use of maintenance treatment.
Mean age was 28 years old (range 16-53). Thirteen were under 30 years, 11 were women, 16 were acute myeloid leukemia. Median time from the first transplant to the relapse was 16,3 months (range 2 to 91 moths). Fourteen patients were MRD negative at the time of second AlloSCT (4 of them in aplasia) and 7 were either positive MRD or refractory disease. Almost all patients received a different re-induction chemotherapy regimen as well as a different conditioning compared to the first transplant. Sixteen out of 21 patients were transplanted with a different donor. Patients without graft versus host disease (GVHD) at day 100 started immunosuppression tapering. Considering the 16 patients alive/relapse free at day 100, half of them received maintenance therapy (6 azacitidine, one sorafenib and one dasatinib).
With a median follow up of 24 months, 1-year OS and LFS was 62% and 38%.
Age was a strong predictor of survival. There were no patients ≥30 years alive or relapse free beyond 6 months of the procedure, while LFS at 2 years was 58% (8/13) in patients younger than 30 years (p=0.0007). One year LFS was 60% for patients receiving any type of maintenance, compared to 33% of patients who did not receive maintenance (p=0.2). There were no differences in changing the donor or the pre-transplant status.
CONCLUSION
Second AlloSCT for relapsed acute leukemia is a potential curative option. Younger patients showed better outcomes. Patients relapsing after 6 months could have better prognosis. Maintenance treatment after second transplant should be evaluated, however prospective studies are needed in this scenario to draw better conclusions.
Figure. Figure.

Disclosures
No relevant conflicts of interest to declare.
Single Cell RNA Sequencing of Serial Tumor and Blood Biopsies from Lymphoma Patients on an in Situ Vaccination Clinical Trial
Authors: Shree Tanaya , Sathe Anuja , Czerwinski Debra K. , Long Steven R. , Ji Hanlee , Levy Ronald 
Published: October 2019

Abstract
The critical determinants of effective antitumor immune responses, whether native or induced by therapy, remain poorly understood due to the complexity and plasticity of the immune system. To better profile and track these responses, we have employed the novel approach of performing single cell RNA sequencing for paired gene expression and immune repertoire analysis on tumor fine needle aspirates and peripheral blood of lymphoma patients undergoing immunotherapy on a clinical trial (NCT02927964).
In this in situ vaccination study, patients with low-grade lymphoma received local low-dose radiation and intratumoral SD-101 (a TLR9 agonist) to one site of disease, with systemic ibrutinib (a BTK inhibitor) added in the second week of treatment. Tumor fine needle aspirates and peripheral blood samples were obtained prior to treatment, at one week (prior to ibrutinib initiation) and at six weeks after treatment start. Single cell preparations were processed using 10X Genomics' single cell RNA transcription and library preparation protocol, followed by sequencing on the Illumina platform. Cells were sequenced to an average depth of 50,000 reads/cell for gene expression libraries and 5000 reads/cell for TCR sequencing. Identification of variable genes, principal component and/or canonical correlation analysis, graph-based clustering and differential expression analysis of single-cell gene expression data was performed using the Seurat algorithm. Single-cell TCR repertoires were analyzed using TCR-specific analysis software. This data is being integrated with data from multiparameter flow cytometry and functional immune assays for these same patients, as well as with their clinical outcomes.
Sequencing libraries have been prepared from 37 samples from 4 patients thus far. We have successfully generated single cell gene expression and TCR libraries from 3,000-10,000 cells from tumor fine needle aspirates and peripheral blood, with excellent sequencing quality metrics obtained. From detailed analyses of one patient's samples thus far, we have identified distinct immune populations in blood and tumor (Figure 1), including light-chain restricted B-cells, with good concordance with flow cytometry. Preliminary results show changes occurring in immune cell frequencies and phenotypes at the treated tumor site, at distant tumor sites and in the peripheral blood when samples from before and after treatment are compared. Sample collection, sequencing, and analysis are ongoing.
Deep profiling of serial biopsies during immunotherapy using single cell RNA sequencing promises to illuminate underlying cellular dynamics, and paired with clinical outcome data, determinants of response. Ultimately, this may provide a roadmap for successful translation of single-cell genomics into the clinic for treatment monitoring and response prediction.

Disclosures
No relevant conflicts of interest to declare.