bluuebunny/crossref_metadata_2025 huggingface dataset:
Piezo1-Sensitive Biomechanical Pulsation Stimulates Long-Term Hematopoietic Stem Cell Formation
Authors: Scapin Giorgia , Cillis Jennifer , Patch Taylor , Dharampuriya Priyanka R , Hagedorn Elliott J. , Lundin Vanessa , Tamplin Owen J , Andeson Heidi , Zon Leonard I. , Daley George Q. , Shah Dhvanit I
Published: October 2019

The birth and development of hemat\opoietic stem cells (HSCs) remain a mystery. During fetal development, a subset of endothelial cells transitions to become HSCs in the aorta-gonad-mesonephros (AGM) region. Blood flow-mediated shear stress and activation of nitric oxide synthase (NOS) were demonstrated to stimulate the endothelial-to-HSC transition in the AGM. However, we showed that malbec (mlbbw306), a zebrafish mutant for cadherin 5, produces HSCs despite circulation arrest and the inhibition of NOS, suggesting that other biomechanical forces, mechanosensation pathways, or epigenetic mechanisms might regulate HSC formation and could have utility in developing HSCs. Using zebrafish, murine, and human models, we show that Piezo1-sensitive biomechanical stretching of hemogenic endothelial cells enhances Dnmt3b expression for long-term (LT)-HSC formation.

Our microangiography and time-lapse confocal imaging established that cdh5-MO embryos have a heartbeat and pulsation in blood vessels despite the absence of blood flow. We also employed light sheet and time-lapse confocal microscopy followed by Fourier transform analyses to establish that although pulsation is independent of blood flow in the AGM, it is concurrent with the endothelial-to-hematopoietic transition. To establish the functional link between pulsation and HSC formation, we developed a bioreactor simulating the pulsating blood vessel conditions. We found that the biomechanical stretching of hemogenic endothelial cells or the pharmacological activation of Piezo1 yields three times higher amounts of LT-HSC formation; which reconstitute to normal multi-lineage adult blood even upon serial transplantation. Our gene-silencing, time-lapse imaging, explant culture, and computational analyses further demonstrated that biomechanical stretching activates Piezo1; which enhances epigenetic regulator Dnmt3b expression to stimulate the endothelial-to-HSC transition.

Our results demonstrate how pulsation-mediated biomechanical forces stimulate cell-fate transitions and stem cell formation by activating mechanosensitive channels as well as epigenetic machinery. We present a model that addresses major challenges in HSC transplantation and cellular therapies for treating blood and bone marrow diseases. In addition, we report a scalable bioreactor with potential widespread use and a pharmacological target to develop and expand LT-HSCs.

Disclosures

No relevant conflicts of interest to declare.
ARRB1-Induced NOTCH1 Degradation Is Inhibited By Mir-223 in T-ALL
Authors: Zou Lin , Shu Yi , Wang Yi , Lv Wenqiong , Peng Danyi , Zhang Hang , Jiang Guangjie , Liu Shan , Chen Yanhua , Tang Shi , Fu Guo , Qi Xinkun , Liu Zhidai , Liu Haiyan , Yang Chao , Liu Fangjie , Yu Jie , Zhang Penghui , Qu Bin , Zhao Hui , He Tongchuan
Published: October 2019

Leukemia is the most common malignant tumor in children under 15 years old, which is divided into several subtypes, including acute lymphoblastic leukemia (ALL), acute myeloid leukemia (AML), chronic lymphoblastic leukemia (CLL) and chronic myeloid leukemia (CML), based on the disease phases and effected cells. Each subtype has its specific molecular feature and key regulation factors. In our previous studies, we reported that β-arrestin1 (ARRB1), the pivotal scaffold protein to transduce various cellular signals, could bind with EZH2 to increase Bcr/Abl H4 acetylation level and thus promote CML progression (Brit J Cancer 2014, 111(3): 568-76). ARRB1 could enhance DNMT1 activity and PTEN methylation, decrease PTEN expression and promote self-renew of B-ALL leukemia initiating cells (LICs) (Cancer Lett 2015, 357(1): 170-8.). ARRB1 could increase P300 to bind with SP1 to hTERT promoter, and thus increase hTERT transcription/expression, telomerase activity, telomere length and cell senescence in B-ALL LICs (Cell Death Diff 2017, 8(4): e2756.). However, little is known in the T-ALL, which about 70% have the mutations of NOTCH1 gene. Here, we unveil ARRB1 could curb the progression of T-ALL cells in vitro and in vivo, while the expression of ARRB1 was suppressed by the aberrant increased miR-223. Mechanistically, ARRB1 could recruit DTX1, the E3 ubiquitin ligase, to promote the ubiquitination and degradation of NOTCH1 protein in T-ALL. Furthermore, Overexpression of ARRB1-derived miR-223 sponge BUTR was incompatible with cell proliferation and induces apoptosis in T-ALL cells. Collectively, our results for the first time revealed that ARRB1 acted as a tumor suppressor by promoting NOTCH1 degradation in T-ALL cells where miR-223 effectively antagonized ARRB1 functions. This provides that miR-223 may serve as a valid drug target for developing novel and efficacious T-ALL therapeutics.

Disclosures

No relevant conflicts of interest to declare.
Utilizing the Predictive Value of the 4T Scoring System to Reduce Unnecessary HIT Panel Costs in the Community Hospital Setting
Authors: Potdar Rashmika , Gupta Sorab , Ng Jia Hwei , Robert Schoepe , Berson Andrew , Bansal Reena , Mohiuddin Kamran , Sy Go Mylene
Published: October 2019

Abstract

Study Objective-Heparin Induced Thrombocytopenia (HIT) is a life-threatening immunological response to heparin. The objectives for this study were to determine if the 4T scoring system was being utilized as a tool for predicting HIT, and to look at the costs associated with HIT panels.

Methods-This was a retrospective chart review of patients greater than 18 years of age who had HIT panels performed between January 2013 and June 2014 in a community hospital. Any duplicate HIT panels sent during the same admission period were excluded. Study investigators were trained in two 30-minute intervals in the area of data collection and retrospective calculation of 4T score.

Results- Of the 154 patients studied, 73 (47.4%) were male, and 81 (52.6%)were female. All patients had a 4T score calculated by study investigators during data analysis, and 1.29% (n=2) had a 4T score calculated before a HIT panel was sent by the team taking care of the patient. 62.3% (n=96) of patients had a low 4T score and 37. 7%( n=58) had an intermediate to high 4T score. Hematology was consulted on 57.7% (n=89) and anticoagulant administration was stopped on 74 % (n=114), while in 26 % ( 40/ 154) heparin was continued despite sending HIT. 25.4%(29/114) were started on alternate anti- coagulants after stopping heparin . Throughout the course of the study, 20 patients died, with only 1 of these patients being HIT positive.

If 100 unnecessary HIT panels were performed in a year, the hospital would be charged more than $20,000 by the diagnostic lab. Additional costs include the halting of Heparin administration and starting an alternate anticoagulant such as Argatroban. 24 hour administration of Argatroban costed $1,000 for continuous infusion. HIT panels have a turnaround time of 4-5 days, resulting in the additional charge of $5,000 just for Argatroban administration. A lengthened stay in the hospital due to HIT panel turnaround time is also a source of increased costs. Combined, the ordering of a HIT panel, alternate anticoagulant administration, and bed charges could amount to $40,000 over the course of four days. This time and money could be put to better use treating the underlying disease of the patient, instead of focusing on the testing for HIT.

Conclusion-Management of HIT in community hospital was sub-optimal. Lack of utilization of the 4T scoring system led to unnecessary ordering of HIT panels. This increased duration of hospital stay, elevated the cost of treatment, and resulted in the holding of prophylactic anticoagulants.

Disclosures

No relevant conflicts of interest to declare.
A New Dosing Model Based on Body Mass Index to Guide Factor VIII Dosing in Patients with Hemophilia A
Authors: Tiede Andreas , Goldmann Georg , Miljić Predrag S , Korsholm Lars , Matytsina Irina , Persson Paula , Lissitchkov Toshko Jelev
Published: October 2019

See attached image file

Disclosures

Tiede: Alnylam, Bayer, Biogen Idec, Biotest, Bristol-Myers-Squibb, Boehringer Ingelheim, CSL Behring, Leo Pharma, Novo Nordisk, Octapharma, Pfizer, Roche, Shire, and SOBI: Consultancy; Alnylam, Bayer, Biogen Idec, Biotest, Bristol-Myers-Squibb, Boehringer Ingelheim, CSL Behring, Leo Pharma, Novo Nordisk, Octapharma, Pfizer, Roche, Shire, and SOBI: Honoraria; Alnylam, Bayer, Biogen Idec, Biotest, Bristol-Myers-Squibb, Boehringer Ingelheim, CSL Behring, Leo Pharma, Novo Nordisk, Octapharma, Pfizer, Roche, Shire, and SOBI: Research Funding. Goldmann:Novo Nordisk and other companies: Honoraria; Novo Nordisk and other companies: Consultancy. Miljić:Novo Nordisk: Other: Investigator fees as a participant of the clinical trial. Korsholm:Novo Nordisk A/S: Employment; Novo Nordisk A/S: Other: Owns stocks. Matytsina:Novo Nordisk A/S: Other: Owns stocks; Novo Nordisk A/S: Employment. Persson:Novo Nordisk A/S: Employment; Novo Nordisk A/S: Other: Owns stocks. Lissitchkov:Novo Nordisk: Other: Investigator fees as a participant of the clinical trial.
Risk Factors and Outcomes of Deep Vein Thrombosis in Pediatric Osteomyelitis
Authors: Ahmad Maria , Knoll Christine M. , Shah Sanjay J. , Mirea Lucia
Published: October 2019

Background:

Estimates of the incidence of DVT in patients with osteomyelitis range widely from 5%-30%, however risk factors and outcomes of DVT in this cohort have not been thoroughly established.

Objective:

This study aims to estimate the incidence of DVT in patients with osteomyelitis, and to assess risk factors and outcomes of DVT.

Design/Method:

After IRB approval, a retrospective chart review was conducted for patients aged 0-18 years seen at Phoenix Children's Hospital between 2012-2016 with ICD 9/10 codes for osteomyelitis. Exclusion criteria included chronic recurrent multifocal osteomyelitis, and chronic DVT. Demographics, clinical factors and outcomes were compared between osteomyelitis patients with and without DVT using the Fisher-exact and Wilcoxon-rank sum tests, as appropriate for the data distribution.

Results:

A total of 179 study subjects with osteomyelitis had a mean (standard deviation) age of 8.4 (5.7) years. DVT was present in 14 (8% of 179) patients, and 4 (28%), 5 (36%) and 5 (36%) patients received anticoagulation for < 6, 6-12 and ≥12 weeks, respectively. Patients with vs without DVT were more likely to be male (86% vs 59%; p-value=0.05), and had significantly higher rates of bacteremia (64% vs 24%; p-value=0.003). Rates of central lines were comparable between DVT and non-DVT patients (71% vs 68%; p-value=1.00); however patients with DVT vs without DVT had significantly longer mean length of stay (18 vs 9 days; p-value <0.0001) and higher rates of ICU admission (71% vs 16%; p-value <0.0001).

Conclusion:

The incidence of DVT among osteomyelitis pediatric patients was estimated at 8%, with risk increased by male sex and bacteremia. Patients with DVT had significantly higher rates of ICU admission and longer length of hospital stay. Many of these patients had standard practice management of their DVT with 6-12 weeks of anticoagulation underscoring the need for future efforts targeting DVT prophylaxis.

Disclosures

No relevant conflicts of interest to declare.
Epigenetic Therapy Targets Th1/Th17 Polarization to Reversing Immune Evasion and Treating Leukemia Relapse Post Allogeneic Stem Cell Transplantation in Non-APL AML Patients
Authors: Yang Xi , Zheng Hong , Dai Jingying , Li Chenglong , Zhang Rong , WANG Xiaodong , Wang Chunsen , Huang Xiaobing
Published: October 2019

To reverse non-APL AML patients with post-transplantation relapse before hematological relapse(early-stage relapse), we investigated the safety and efficacy of a new epigenetic therapy(DNA methyltransferase inhibitors, Decitabine and histone deacetylase inhibitors, Chidamide, plus Thymalfasin simultaneously). Twenty-four patients were enrolled in this observational study. The most common adverse event was CTCAE grade 2 thrombocytopenia (20/24). No acute GVHD was observed in patients received this treatment regimen. Strikingly all 24 patients responded to this epigenetic therapy with minute residual disease(MRD) decreasing (ORR 100%). At the more than 3-year follow-up, overall survival rate is 87.5%(21/24), with relapse-free survival rate of 75%(18/24). With this epigenetic therapy for 3 months, Th1 cells and CD3+CD4-CD8+T cells increased gradually, and Th17 cells decreased. The status of Th1 and Th17 cells remained at the time of 3 months after discontinuation. Interestingly, the Th1/Th17 ratio was low in patients with relapse disease and then increased gradually during this treatment regimen, correlated with the down trend of MRD. Therefore, our new epigenetic therapy is safe and tolerable for non-APL AML suffering post-transplant early relapse. The polarization for Th1/Th17 and the significant elevated ratio of Th1/Th17 seemed to better reflect the epigenetic therapy-mediated immune effect, which maybe a good biomarker for the early relapse monitoring.

Disclosures

No relevant conflicts of interest to declare.
Increased Risk of Second Primary Hematologic and Solid Malignancies in Patients with Mycosis Fungoides
Authors: Goyal Amrita , O'Leary Daniel , Lazaryan Aleksandr
Published: October 2019

Importance: Mycosis fungoides (MF) is a rare, generally indolent non-Hodgkin T-cell lymphoma of the skin. It has previously been associated with increased risk of second hematologic malignancies and melanoma, but the association with second solid malignancies has not been well characterized.

Objective: This retrospective analysis seeks to assess the risk of developing a second primary hematologic or solid malignancy in patients with MF.

Design: We performed (1) an analysis of patients diagnosed with MF from 2000-2014 in the 18 population-based United States cancer registries of the Surveillance, Epidemiology, and End Results Program (SEER-18), and (2) a retrospective cohort study of patients with MF treated at the University of Minnesota (UMN) from 2005-2017 with a control group of patients with seborrheic dermatitis.

Setting: This study is a combination of population-based data and a single-center cohort.

Participants: SEER-18 consists of a population-based sample of patients across the United States with a diagnosis of MF (n=6196). The UMN cohort included a cohort of 172 patients with MF treated at the UMN from 2005-2017.

Results: In the SEER-18 cohort, relative risks were estimated using the standardized incidence ratios (SIR). Of the 6196 patients in the MF cohort there were 514 (8.3%) second cancer events (SIR 7.3, 95% confidence interval [CI] 6.7-7.9). Patients with MF were at substantially increased risk for both. non-Hodgkin (SIR 46.51, CI 39.16-55.0) and Hodgkin lymphoma (SIR 69.8, CI 34.8-124.9). MF patients were also at higher risk of melanoma (SIR 7.2, CI 4.5-10.9), lung (SIR 6.2, CI 4.8-7.8), female breast (SIR 8.0, CI 5.8-10.6), prostate (SIR 4.1, CI 3.1-5.2), bladder (SIR 3.6, CI 2.0-5.8), colon (SIR 5.2, CI 3.7-7.2) and renal (SIR 3.9, CI 1.8-7.3) cancers.

In the UMN cohort, patients with MF were significantly more likely to develop a second malignancy than patients with seborrheic dermatitis (relative risk [RR] 8.1, p<0.0005, CI 5.1-13.2). Patients with tumor stage MF were more likely to develop a second malignancy than those with patch/plaque stage disease (RR 3.2 p<0.01, CI 2.6-10.4). Similarly, patients with stage IIB or higher were significantly more likely than those with early stage disease (RR 3.0, p<0.003, CI 3.1-13.5) to develop a second malignancy. Analysis of treatments and imaging undergone by each patient revealed no significant difference in systemic, topical, radio- or phototherapy, or number of CT scans between patients who did and did not develop second malignancies.

Conclusions and Relevance: Patients with MF are at increased risk of developing second malignancies, particularly those with advanced stage disease or tumors. These findings warrant the development of targeted screening strategies for patients with MF.

Disclosures

No relevant conflicts of interest to declare.
The Impact of Selenium Deficiency on a Sickle Cell Disease Mouse Model
Authors: Jagadeeswaran Ramasamy , Lenny Hong , Zhang Helen , Afranie-Sakyi Jennifer , Molokie Robert E. , Lavelle Donald , Diamond Alan , Rivers Angela
Published: October 2019

Sickle cell disease (SCD) is caused by a mutation in the β-globin gene resulting in a disease that affects more than 100,000 Americans and millions worldwide. Though pain is the hallmark of SCD, patients also suffer damage to most organ systems. Sickle cell hemoglobin (HbS) polymerization occurs when deoxygenated, rendering red blood cells rigid and fragile. Production of excessive reactive oxygen species (ROS) and intracellular hypoxia in RBCs further accelerates the pathology associated with SCD. Recently, vaso-occlusive crisis (VOC) and organ damage were established to be strongly associated with oxidative stress in RBCs. This occurs when there is an increase in oxidants that exceeds the cellular anti-oxidant defenses. Excessive ROS can trigger a cascade of oxidative reactions that damage membrane lipids, and essential enzymatic antioxidants such as GPx-1, ultimately leading to hemolysis and multi-organ dysfunction. ROS generation in RBCs of SCD patients is due to factors such as HbS auto-oxidation and potentially aberrant mitochondrial function. We recently determined that red blood cells obtained from SCD mice and SCD patients retain their mitochondria compared to control subjects. Mitochondria retained SCD RBCs are also associated with elevated levels of ROS and hemolysis. Oxidative stress in the RBCs of SCD patients may be elevated by lower levels of antioxidant proteins such as the selenium-dependent enzyme GPX1. GPX1 was first described as an enzyme capable of protecting hemoglobin from ROS and has been reported to be lower in the RBCs in SCD. Selenium levels are lower among African Americans in the Chicago area and elsewhere. In this regard, it is notable that in the United States, African Americans represent the majority of those with SCD. To investigate the relationship between selenium levels and SCD, we have utilized a mouse model of SCD to examine the impact of a reduced intake of selenium on parameters associated with SCD pathology. SCD mice on a selenium-deficient diet (<0.01 mg/kg diet) were compared to mice fed with a selenium-adequate diet (0.1mg/kg). SCD mice in the selenium-deficient group exhibited an increase in mitochondria retaining RBCs (Se deficient: 26%±6.9%, n=3 vs. Se adequate: 5 % ± 3.5%, n=3, p<0.01), reduced Hb levels (Se deficient 5.7± 0.17 g/dl, n=3 vs. Se adequate 7.0± 0.83 g/dl, n=4 p<0.05), and an increased RBC oxygen consumption rate (OCR). These results support the hypothesis that low selenium status likely results in reduced levels of anti-oxidant selenoproteins and enhanced SCD severity.

Disclosures

Lavelle: Global Blood therapeutics: Research Funding.
Boosting Gamma Delta T Cells-Mediated ADCC By PD-1 Blockade in Follicular Lymphoma
Authors: Bezombes Christine , Rossi Cédric , Gravelle Pauline , Bordenave Julie , Decaup Emilie , Poupot Mary , Tosolini Marie , Franchini Don-Marc , Laurent Camille , Morin Renaud , Lagarde Jean-Michel , Ysebaert Loic , Savina Ariel , Klein Christian , Perez-Galan Patricia , Fournié Jean-Jacques
Published: October 2019

Follicular lymphoma (FL) is a common non Hodgkin's lymphoma subtype in which immune escape mechanisms are implicated in resistance to chemo-immunotherapy. Although molecular studies point to qualitative and quantitative deregulation of immune checkpoints, in depth cellular analysis of FL immune escape is lacking. Here, by functional assays and in silico analyses we show that a subset of FL patients displays a 'high' immune escape phenotype. These FL cases are characterized by abundant infiltration of PD1+ CD16+ TCRVgamma9Vdelta2 gamma delta T lymphocytes and high expression of gamma delta cytolytic markers. In a 3D co-culture assay (MALC), gamma delta T cells could mediate both direct and indirect (ADCC in the presence of anti-CD20 mAbs) cytolytic activity against FL cell aggregates. Importantly, PD-1, which is expressed by most FL-infiltrating gamma delta T lymphocytes with ADCC capacity, impaired these functions. In conclusion, we identify a PD1-regulated gamma delta T cell cytolytic immune component in FL. Our data provide a treatment rational by PD-1 blockade aimed at boosting gamma delta anti-tumor function in FL.

Disclosures

Morin: IMACTIV-3D: Employment. Lagarde:IMACTIV-3D: Employment. Savina:Institut Roche: Employment. Klein:F. Hoffmann-La Roche Ltd.: Employment, Other: stock, Patents & Royalties.
Lymphadenopathy in a TB-Endemic Region: Employing Focused Techniques in a Rapid Access Lymph Node Biopsy Clinic
Authors: Antel Katherine Rae , Verburgh Estelle , Louw Vernon J. , Oosthuizen Jenna , Malherbe Francois , Gary Maartens , Nicol Mark
Published: October 2019

Introduction

Lymphoma and extrapulmonary tuberculosis (EPTB) are common in HIV and may cause peripheral lymphadenopathy. EPTB can be difficult to diagnose and consequently, patients with lymphadenopathy in TB-endemic regions are frequently placed on empiric TB treatment. This may result in a missed or delayed diagnosis of lymphoma.

In the local setting, in Cape Town (South Africa), primary level doctors have difficulty accessing lymph node excision biopsies which rely on an over-burdened surgical service. Core-needle biopsy is not currently routinely performed for peripheral lymphadenopathy in the investigation of lymphadenopathy.

Methods

A prospective study of patients with peripheral lymphadenopathy >2cm was performed. A rapid access lymph node biopsy clinic examined the performance of three methods of investigation on a lymph node: the fine-needle aspirate for cytology (FNAC), the Xpert MTB/Rif Ultra (on both aspirate and tissue specimens), and the ultrasound-guided core-needle biopsy. In all patients a final histological diagnosis was sought, including referral for excision biopsy if core-needle histology was inconclusive. If >0.5ml of pus was aspirated this was tested with the Xpert MTB/Rif and a biopsy was not performed.

Results

Sixty-two patients including 48% HIV+ have undergone investigation. The final diagnosis is known for 56 patients: lymphoma in 34%, TB in 30%, cancer in 12%. Ultra had a 64% sensitivity on the aspirate (95% CI 31-89%), and 73% on tissue (95% CI 39-94). In both cases specificity was 100%. FNAC was diagnostic in 0% of lymphoma cases (reported atypical in 24%); it was diagnostic for TB (+AFBs) in 27% with TB; and diagnostic in 36% of disseminated cancer cases. The core biopsy was able to make a diagnosis in 83% of cases (including sub-typing in 89% of lymphoma cases).

Conclusion

The Ultra shows promising sensitivity and should routinely be performed for both HIV+ and -patients at the first visit in the investigation of unexplained lymphadenopathy in a TB-endemic region. If the Ultra is negative, a core-needle biopsy is the most appropriate next investigation. In a TB-endemic region, FNAC should be reserved for patients >50 years who are HIV negative. There is no role for empiric TB treatment for patients with peripheral lymphadenopathy in the era of new TB diagnostic tests, and the application of these investigations with early biopsy for patients with negative TB tests will result in earlier detection of lymphoma.

Disclosures

No relevant conflicts of interest to declare.
Genotypic Groups As Risk Factor for Cardiac MR Abnormalities and Complications in Thalassemia Major
Authors: Pepe Alessia , Meloni Antonella , Salvadori Stefano , Macchi Silvia , Vitucci Angelantonio , Murgia Mauro , Maggio Aurelio , Fiorenza Maria Flavia , Massei Francesco , Commendatore Francesca Valeria , Giuliano Pietro , Positano Vincenzo , Pistoia Laura
Published: October 2019

Introduction. Beta thalassemia major (β-TM) displays a great deal of genotypic heterogeneity, not fully investigated in terms of cause-effect. This prospective and multicentre study aimed to detect if different genotypic groups could predict the development of cardiovascular magnetic resonance (CMR) abnormalities and cardiac complications (CC).

Methods. We considered 708 β-TM patients (373 females, 30.05±9.47 years), consecutively enrolled in Myocardial Iron Overload in Thalassemia (MIOT) network. Data were collected from birth to the first CMR imaging scan. Myocardial iron overload was assessed by the multislice multiecho T2* technique. Biventricular function parameters were quantified by cine images. Late gadolinium enhancement (LGE) images were acquired to detect myocardial fibrosis.

Results. On the basis of the type of gene mutation, three groups of patients were identified: homozygotes β+ (N=158), compound heterozygotes β+ / β° (N=298) and homozygotes β° (N=252). Table 1 shows the effect of genotype group on the development of different cardiac outcomes. Compared to the milder genotype group homozygotes β+, the other two groups showed a significantly higher risk of myocardial iron overload (MIO) and left ventricular dysfunction. We recorded 90 (13.0 %) cardiac events: 46 heart failures (HF), 38 arrhythmias (33 supraventricular, 3 ventricular and 2 hypoinetic) and 6 pulmonary hypertensions (PH). No prospective association was detected between genotype group and HF and PH. The homozygous β° group showed a significantly higher risk of arrhythmias than the homozygous β+ group and at the limit of significance than the compound heterozygotes. Globally, homozygotes β° showed a significantly higher risk of CC than homozygotes β+.

Conclusion. Different genotypic groups predict the development of MIO, left ventricular dysfunction, arrhythmias and CC in β-TM patients. These data support the knowledge of the different genotypic groups in the clinical management of β-TM patients.

Table Table.

Disclosures

Pepe: Chiesi Farmaceutici S.p.A., ApoPharma Inc., and Bayer: Other: No profit support.
Metabolic Reprogramming Overcomes T Cell Inhibition By AML Cells
Authors: Uhl Franziska Maria , Chen Sophia , O'Sullivan David , Buck Michael , Büscher Jörg , Pfeifer Dietmar , Finke Jürgen , Duyster Justus , Bertz Hartmut , Shoumariyeh Khalid , Börries Melanie , Richter Gesa , Madl Tobias , Pearce Erika L. , Zeiser Robert
Published: October 2019

Allogeneic hematopoietic cell transplantation (allo-HCT) is the treatment strategy of choice for the majority of patients with acute myeloid leukemia (AML). However, about 40% of patients relapse, which is associated with a poor prognosis. Donor lymphocyte infusions (DLIs) induce remission only in a minority of relapsed AML patients.

In order to understand the impact of AML cells on T cell bioenergetics, we investigated the metabolic phenotype and immunological profile of CD8+ T cells isolated from AML patients at primary diagnosis, during remission and relapse after allo-HCT. At diagnosis, T cells exhibited robust metabolic activity and strong immune activation. In contrast, during relapse, T cells demonstrated reduced glycolytic and respiratory activity compared to their status at remission. The metabolic phenotype was accompanied by reduced cytotoxic ability. Metabolomics revealed lactate accumulation and substantial arginine depletion in the serum of these patients.

The results observed in the human setting could be reproduced in a murine graft versus leukemia (GVL) model where T cells isolated from AML bearing mice exhibited lower metabolic activity compared to control animals.

In vitro co-culture experiments enabled tightly controlled conditions for mechanistic studies. The metabolic impairment induced by AML cells was mediated by soluble factors, mirroring the major changes in serum metabolites of patients, and interfered with the crucial metabolic reprogramming in the priming phase of T cells. Well-known mechanisms such as nutrient deprivation could be excluded. Extensive transcriptomic and metabolomic analyses have been undertaken in order to identify the molecules responsible for the diminished metabolic activation.

Influencing mitochondrial architecture thereby boosting respiratory activity prior to infusion led to enhanced anti-leukemic activity of T cells in vivo. The pre-treatment favored an immunological memory phenotype essential for a potent GVL effect. Using metabolic stimuli as pre-treatment of DLIs could promote T cell resistance against tumor evasion and increase anti-tumor immunity. This could greatly improve the success of DLIs and other adoptive T cell transfer strategies in the treatment of AML relapse.

Disclosures

Finke: Neovii: Consultancy, Honoraria, Other: travel grants, Research Funding; Medac: Consultancy, Honoraria, Other: travel grants, Research Funding; Novartis: Consultancy, Honoraria, Other: travel grants, Research Funding; Riemser: Consultancy, Honoraria, Research Funding.
The Expression of EIF2S3 in Acute Myeloid Leukemia and Its Prognostic Significance
Authors: Lu Jielun , Xu Lihua , Luo Xiaodan , Chen Shuyi , Zou Yawei , Tan Huo
Published: October 2019

Background: Eukaryotic translation initiation factor 2 subunit gamma (EIF2S3) is a component of eIF2 ,which functions in the early steps of protein synthesis by forming a ternary complex with GTP and initiator tRNA and binding to a 40S ribosomal subunit. The preliminary research suggested that in multiple malignant carcinomas, the expression level of EIF2S3 between the normal tissue and malignant one is Significantly different.It indicated that EIF2S3 might play an important role in the occurrence and development of malignant tumors .The purpose of this study is to investigate the expression status of EIF2S3 and its prognostic significance in AML patients.

Methods:Total RNA was extracted from bone marrow of 42 patients. The expression of EIF2S3 was examined by Realtime quantitative PCR. The relationship of EIF2S3 expression and other clinicopathological charactors and itsprognosis significance were statistically analyzed.

Results:The relative expression of EIF2S3 was much higher in patients who had achieved CR (median value 0.088, range: 0.0179-0.238;) compared with newly diagnosed AML patients(median value 0.037, range: 0.003-0.203),p value was 0.020.Furthermore, the expression of EIF2S3 was markedly associated with Complete Remission (P = 0.012) and outcome(P=0.029). Patients with lower EIF2S3 expression had a relatively poor overall survival(OS) (P = 0.016) and short disease-free survival(DFS) (P = 0.001). Moreover, EIF2S3 expression was an independent prognostic factor for both OS (P = 0.001) and DFS (P <0.001).

Conclusions:Patients who had achieved CR usually had a higher level of EIF2S3 expression compared with newly diagnosed AML patients. The over-expression of EIF2S3 is associated with good prognosis in AML patients, and was an independent prognostic factor for both OS and DFS.

Disclosures

No relevant conflicts of interest to declare.
Yearly Percent Incidence of Suspected Novel Oral Anticoagulant Interference on Thrombophilia Testing: A Nine Year Experience at a Reference Coagulation Laboratory
Authors: Wong Edward C.C. , Worfolk Laura A. , Dlott Jeffrey S.
Published: October 2019

Introduction: With the recent FDA approval of novel oral anticoagulants (NOACs) for anticoagulation, the diagnosis of thrombophilia risk factors, such as protein S deficiency, protein C deficiency and presence of lupus anticoagulant, has become problematic, especially when clot-based assays are utilized in the presence of Xa and direct thrombin inhibitors. Falsely elevated protein S and C activities and false positive lupus anticoagulant testing may occur with NOAC use. Little is known about the yearly incidence of such interference during the time period when these NOACs were introduced in the United States (dabigatran, 10/2010; rivaroxaban, 7/2011; apixaban, 12/2012; edoxaban, 1/2015). To answer this question, we undertook a retrospective study that examined interference before and after NOACs were introduced to the US market.

Materials and Methods: Retrospective review of venous thrombophilia testing from January 2008 to December 2016 at a major coagulation reference referral laboratory. To identify potential NOAC interference, samples were tested with both a dilute Russell Viper Venom test (dRVVT), which is a lupus anticoagulant test, and a protein C activity test. NOAC interference was strongly suspected when dRVVT testing demonstrated an inhibitor pattern and elevated protein C activity (>200%). The annual percentage of thrombophilia tests with strongly suspected NOAC interference was determined and compared to that of tests with elevated protein C activity and without a dRVVT inhibitor pattern (less strongly suspected interference).

Results: The percentage of suspected NOAC interference increased annually from 2008 to 2016, which corresponds to the introduction of NOACs to the US market. From 2008 to 2016, test results of suspected NOAC interference increased from 0.21% to 3.45% (16.4-fold increase), peaking in 2015 (24.1-fold increase); test results of elevated protein C activity without a dRVVT inhibitor pattern increased from 0.48% to 7.22% (15.0-fold increase), peaking in 2014 (31.5-fold increase). Suspected interference decreased between 2015 and 2016, which may be related to education efforts by our laboratory in 2014 and 2015, when we notified practitioners of possible interference.

Conclusions: Suspected NOAC interference, which can potentially cause false-positive test results for lupus anticoagulant and protein C activity, increased when NOACs were introduced into the US market. Further education of licensed independent practitioners may help prevent these interferences and provide more accurate assessment of thrombophilia risk factors.

Figure. Figure.

Disclosures

Wong: Quest Diagnostics: Employment, Other: stock . Worfolk:Quest Diagnostics: Employment. Dlott:Quest Diagnostics: Employment, Other: stock.
Improved Pharmacokinetic Profile for BAY 81-8973 Due to Increased Branching and Sialylation of N-Linked Glycans of Recombinant Factor VIII
Authors: Teare John M. , Kates David S. , Shah Anita , Garger Stephen
Published: October 2019

The circulatory half-life of recombinant factor VIII (rFVIII) products is affected by glycosylation of the FVIII protein, including N-linked glycan branching and terminal sialic acid occupancy, primarily through receptor-mediated hepatic clearance (eg, asialoglycoprotein receptor [ASGPR] and lipoprotein receptor-related protein [LRP]). BAY 81-8973 (Kovaltry®, Bayer, Berkeley, CA) is an unmodified full-length rFVIII for treatment of hemophilia A. The BAY 81-8973 manufacturing process results in a product of enhanced purity with a consistently high degree of branching and sialylation of N-linked glycans. This study evaluated whether a relationship exists between N-linked glycosylation patterns and pharmacokinetic (PK) characteristics of BAY 81-8973 and 2 other rFVIII products (sucrose-formulated rFVIII [rFVIII-FS; Kogenate® FS, Bayer] and antihemophilic factor (recombinant) plasma/albumin-free method [rAHF-PFM; Advate®, Shire, Westlake Village, CA]).

N-linked glycans or terminal carbohydrates were enzymatically removed from immobilized BAY 81-8973, rFVIII-FS, and rAHF-PFM proteins and analyzed using high-performance liquid chromatography to determine the percentage of individual N-linked glycan structures and degree of sialylation of each structure. PK data were available from 2 separate phase 1 crossover studies in which the PK profile of BAY 81-8973 was compared with that of rFVIII-FS (n=26) and rAHF-PFM (n=18) in patients with severe hemophilia A who received a single 50-IU/kg dose of each product.

BAY 81-8973 and rFVIII-FS had increased N-linked glycan branching with higher levels of sialylation compared with rAHF-PFM. Levels of trisialylated glycans were 29.0% for BAY 81-8973 versus 11.5% for rFVIII-FS and 4.8% to 5.5% for rAHF-PFM; tetrasialylated glycans were 12.0% versus 2.8% and 0.6%, respectively. Degree of sialylation was 96% for BAY 81-8973, 94% for rFVIII-FS, and 78% to 81% for rAHF-PFM. Based on chromogenic assay results from the single-dose phase 1 PK studies, BAY 81-8973 half-life was 15% longer than that for rFVIII-FS and 16% longer than rAHF-PFM.

Increases in the percentage of sialylated tri-antennary and tetra-antennary N-glycans correlated well with longer half-life of rFVIII in humans (adjusted R2=0.978 and 0.892 for tri-antennary and tetra-antennary N-glycans, respectively). Higher percentages of sialylation (ie, sialic acid capping) correlated with a longer half-life (adjusted R2=0.697), but the relationship was not as strong as that between glycan branching and half-life. Improved PK for BAY 81-8973 relative to rFVIII-FS and rAHF-PFM as seen in single-dose crossover PK studies might be related to this greater level of branching and sialylation, which may prolong the time BAY 81-8973 remains in the circulation.

Disclosures

Teare: Bayer: Employment. Kates:Bayer: Employment. Shah:Bayer: Employment. Garger:Bayer: Employment.
RDW Is a Prognostic Marker for Patients with Aggressive Peripheral T-Cell Lymphoma
Authors: Beltrán Brady E. , Castro Denisse A. , Huerta- Collado Yesenia M. , Sotomayor Eduardo , Castillo Jorge J.
Published: October 2019

Introduction: Red blood cell distribution width (RDW) has emerged as a potential prognostic factor in solid tumors and lymphomas. Previous studies have shown an association between increased levels of RDW and inflammatory diseases, being a surrogate marker of inflammation and a predictor of poor outcome. Data on the impact of RDW in outcomes of patients with aggressive peripheral T-cell lymphoma (PTCL) are scarce.

Methods: We performed a retrospective study on consecutive patients with a de novo diagnosis of aggressive PTCL diagnosed and treated at our institution between 2010 to 2016. All patients included were treated with chemotherapy with a curative intent. The RDW was collected from the hemogram of PTCL patients at diagnosis. RDW-CV >14% and RDW-SD >49 fL were our cut-offs for the categorical analysis. We fitted univariate and multivariate Cox proportional hazard regression models for overall survival (OS). P<0.05 were considered statistically significant.

Results: A total of 101 patients with de novo aggressive PTCL were included. This study included 53 patients (52%) with PTCL, not otherwise specified, 16 patients (16%) with extranodal natural killer/T-cell lymphoma,15 (15%) with adult T-cell lymphoma/leukemia,14 patients (14%) with anaplastic large cell lymphoma, 2 patients (2%) with angioimmunoblastic T-cell lymphoma, and 1 patient (1%) with enteropathy-associated T-cell lymphoma. Median age was 57 with a male predominance (69%). Clinically, 45 patients (45%) were 60 years or older, 41 patients (34%) had ECOG ≥2, 63 patients (65%) had increased LDH levels, 68 patients (67%) had > 1 extranodal site, 65 patients (64%) had stage III/IV disease, and 55 patients (54%) had B symptoms. RDW-CV was >14% in 64 patients (64%). RDW-SD was >49 fL in 35 patients (35%). 38% of patients received CHOEP, 29% received CHOP, and 33% received other regimens. The overall response rate was 68%; 50% had a complete response and 18% had a partial response. At 5 years, median overall survival (OS) was 46%. In the univariate Cox regression analysis, ECOG ≥2 (p<0.001), high RDW-CV >14% (p<0.001), high RDW-SD >49 fL (p=0.007) and high neutrophil-lymphocyte ratio (NLR >4) (p=0.002) were associated with a worse survival. In the multivariate Cox regression analyses, ECOG ≥2 (HR 3.1 CI 1.6-6.0; p=0.001) and high RDW-CV >14% (HR 2.7, 95% CI 1.2-6.0; p=0.01) were independent predictors of poor survival. RDW-CV was an independent factor of survival when was compared with IPI and NCCN-IPI score.

Conclusions: RDW is an independent marker for adverse prognosis in aggressive patients with aggressive PTCL treated with curative intent.

Figure. Figure.

Disclosures

Castillo: Genentech: Consultancy; Pharmacyclics: Consultancy, Research Funding; Janssen: Consultancy, Research Funding; Beigene: Consultancy, Research Funding; Abbvie: Consultancy, Research Funding; Millennium: Research Funding.
Morning Vs Bedtime Aspirin Intake in Patients with Essential Thrombocythemia
Authors: Cacciola Emma , Gentini Cacciola Elio , Vecchio Veronica , Cacciola Rossella Rosari
Published: October 2019

The essential thrombocythemia (ET) is a myeloid neoplasm characterized by platelet hyperreactivity and thrombotic risk. The treatment with aspirin (ASA) is recommended in ET patients at risk of first-time or recurrent thrombotic events. An unexplored topic is the optimal timing of once daily ASA intake. On the basis of the presumptions that 1) platelet aggregation is higher in the morning and that 2) the platelet inhibitory effect of ASA is not sustained during the usual 24-hour (h) dosing interval and that 3) a higher gastric mucosal resistance in the evening, we evaluated platelet count, β-thromboglobulin (β-TG) and platelet factor 4 (PF4), as markers of platelet activation, the clotting time (CT), clot formation time (CFT) and maximum clot formation/firmness (MCF), as indicators of aspirinated platelet contribution to clot formation/firmness. We studied 60 patients (20 men, 40 women; mean age 51 years, range 32-70) with ET according to WHO criteria. The mean duration of disease was 11 years. All patients were on ASA 100 mg once daily. Of these, 30 took ASA on awakening and 30 took ASA at bedtime. Of the 60 patients, 45 were on anagrelide hydrochloride (daily dose 1.5 mg) (10 men, 35 women), 15 were on hydroxyurea (daily dose 2 mg) (10 men 5 women). None had inherited or acquired thrombotic risk factors. Sixty subjects served as controls. Platelets were measured by automated analyzer. β-TG and PF4 were determined by ELISA. CT, CFT and MCF were measured by ROTEM delta. The mean platelet count was 455±200x109/L. The awakening ASA patients had normal β-TG and PF4 (12±5 IU/ml and 4±1 IU/ml), normal CT (CT, unit: s. n.v. 100-240 s) ( 110±20 s), normal CFT (CFT, unit: s, n.v. 30-110 s) (45±5 s) and normal MCF (MCF, unit: mm, n.v. 50-72 mm) (61±2 mm), whereas the bedtime ASA patients had high β-TG and PF4 (200±15 IU/ml vs 20±11 IU/ml and 170±50 IU/ml vs 6±2 IU/ml, respectively) (p<.0001 and p<.0001, respectively), shortened CT (CT, unit: s. n.v. 100-240 s) ( 80±10 s), CFT (CFT, unit: s, n.v. 30-110 s) (15±5 s) and MCF (MCF, unit: mm, n.v. 50-72 mm) (40±2 mm). These findings suggest that in ET patients the optimal timing of once daily ASA intake is in the morning.

Disclosures

No relevant conflicts of interest to declare.
CD25 (IL2RA) Is a Direct Transcriptional Target of PRDM1 in Activated Human Natural Killer Cells
Authors: Akman Burcu , Kursun Deniz , Hatipoglu Tevfik , Hu Xiaozhou , Chan Wing C , Kucuk Can
Published: October 2019

Background: PRDM1 is a transcription factor that regulates differentiation and/or homeostasis of B and T lymphocytes. It may also play a role in natural killer (NK) cell homeostasis, and it is frequently inactivated in NK cell-derived lymphomas. Many target genes of PRDM1 have been reported; however, there is little insight into genes directly targeted by PRDM1 in NK cells. IL2 receptor signaling is crucial for many features of NK cell activation including sustained proliferation and survival. Here we evaluated whether CD25 (IL2Rα), a critical gene involved in NK cell activation, is transcriptionally repressed by PRDM1 in activated human NK cells.

Methods: ChIP-Seq was applied on normal primary NK cells activated using a special co-culture system that involved genetically engineered NK cell target cells (i.e. K562-Cl9-mb21) to determine genomic locations occupied with PRDM1. PRDM1 binding site screen in CD25 was computationally performed using two different ChIP-Seq peak callers. Two malignant NK cell lines (NK92 and KHYG1) were transduced with a retroviral construct that co-expresses PRDM1α and GFP. CD25 transcription expression was evaluated with DNA microarray, RNA-Seq and RT-qPCR on GFP-sorted NK cells 48h post-transduction. CD25 expression was stably knocked-down with two different shRNAs in NK92 and KHYG1 cell lines. CD25 was ectopically expressed in primary human NK cells activated through coculturing with K562-Cl9-mb21. GFP competition assays were performed on CD25 shRNA-transduced NK cell lines or primary NK cells with ectopic CD25 expression in limiting IL2 concentrations, respectively.

Results: PRDM1 occupancy on the 1st intron of CD25 was detected with CisGenome and MACS softwares in activated primary NK cells. Direct binding of PRDM1 on CD25 was further validated by ChIP-qPCR in normal activated NK cells. Two PRDM1α-transduced PRDM1-nonexpressing NK cell lines showed significant transcriptional repression of CD25 by DNA microarray and RNA-Seq, the repression of which was cross-validated with RT-qPCR. Stable knock-down of CD25 inhibited growth of two NK cell lines cultured in limiting IL2 concentrations. By contrast, ectopic expression of CD25 resulted in positive selection of ex vivo cultured primary NK cells.

Conclusions: Altogether these results establish CD25 as a direct transcriptional target of PRDM1 in human NK cells. These observations provide additional support for the role of PRDM1 in termination of NK cell activation and growth, with implications on neoplastic transformation or NK cell function when it is deregulated.

Disclosures

No relevant conflicts of interest to declare.
Gene Mutation Profile and Risk Stratification in AML1-ETO-Positive Acute Myeloid Leukemia Based on Next-Generation Sequencing
Authors: Yu Guopan , Yin Changxin , Wu Fuqun , Jiang Ling , Zheng Zhongxin , Xu Dan , Zhou Jiaheng , Jiang Xuejie , Liu Qifa , Meng Fanyi
Published: October 2019

Gene mutations play a critical role in leukemogenesis of AML1-ETO-positive acute myeloid leukemia (AE-AML). Nevertheless, gene mutation profile in this subtype leukemia remains unclear, and their clinical effect might be underestimated. In this study, we detested gene mutations at diagnosis and relapse with next-generation sequencing in 64 newly diagnosed AE-AML patients, and verified the results with Sanger sequencing at the same time. Our results showed that 68.8% patients presented recurrent mutations at diagnosis and 6/11 cases underwent genetic alterations at relapse. C-KIT mutation was the most common event at diagnosis, with an incidence of 42.2%, followed by ASXL1 (15.6%), MET (12.5%), MLH1 (9.4%) , TET2 (7.8%), and FBXW7, TP53 and DNMT3A (7.8%), etc. Also, C-KIT mutation was the most common molecular event associated with relapse (7/11, 63.6%). No significant difference in the clinical characteristic between the gene mutation and wild type (WT) groups was observed, except of higher incidence of additional cytogenetic abnormalities (ACAs) (P=0.025) in TP53 mutation patients. C-KIT (exon 8, 17) mutation but not exon 10 adversely affected on survival. Also ASXL1 and TP53 mutation were poor for the RFS (P<0.05), and ASXL1, MET, FBXW7 and TP53 mutation negatively impacted on the OS (P<0.05). Multivariate analysis showed C-KIT (exon 8, 17) and ASXL1 mutations were the independent adverse factors for survival. Further, co-mutation of these two genes showed even worse effect on survival. Taking together, additional gene mutations are critical in the development and progression of AE-AML. C-KIT and ASXL1 mutations are the two most common molecular events in this subtype leukemia. Single mutation of C-KIT (exon 8, 17) or ASXL1 poorly affects on survival, even worse in the co-mutation.

Figure. Figure.

Disclosures

No relevant conflicts of interest to declare.