bluuebunny/crossref_metadata_2025_split huggingface dataset:
A Report of Fourcases of Severethrombocytopenia with Rh G-CSF Mobilized Allogeneic Peripheral Blood Stem Cell Transplant Donors
Authors: Liu Qingzhen , Yao Jianfeng , Yang Donglin , Pang Aiming , Jiang Erlie , Feng Sizhou , Han Ming-Zhe 
Published: November 2020

Background and Objective: Granulocyte colony-stimulating factor (G-CSF) is used to mobilize progenitor cells into the peripheral bloodin healthy donors. Thrombocytopenia is one of common side effects of G-CSFmobilization, but seriousthrombocytopenia is rare. Herein,we report serious thrombocytopeniain 4 allogeneic peripheral blood stem cell transplantation (Allo-PBSCT) healthy donors induced by recombinant human G-CSF (rhG-CSF) mobilization.
Methods: The donor received 10ug/kg/d G-CSF via subcutaneous injection.The blood parameterswere examined every day, Abdominal ultrasound detected the spleen size before, during, and after rhG-CSF mobilization.
Results: From April 2015 to May 2020, 1127 donors received hematopoietic stem cell mobilization, the percentage of serious thrombocytopenia in healthy donors was 0.35%.The count of white blood cell increased to the 4-7 timesbasic level of (35.4x109/L)after rhG-CSF mobilization 4 days, the lowestcount ofplatelet was 22x109/Lafter single postharvest. After mobilization, length of spleen increased 0.6-1.3 cm (median 10.1 cm), thickness ofspleenincreased 0-1.3 cm (median 3.4 cm). There were no obvious bleeding symptomsafter mobilization in four cases of donor.The quality of stem cell collection was relatively poor in 3 of the 4 donors, the number of mononuclear cells (MNC) was 7.47-18.58×108/kg, and the value of CD34+ was 0.82-2.42×106/kg (CD34+ proportion: 0.11-0.16%). The quality of stem cell collection in the other 1 donor was satisfactory, the number of MNC was 9.29-11.75×108/kg, and the proportion of CD34+ was 0.26~0.38%. Platelet count of 4 donors recovered to the basal level 3-10 days after drug withdrawal.
Conclusion:The severe thrombocytopenia caused by rhG-CSF mobilization healthy donor is very rare.while, the mechanismis unknown. Some studies suggested that it may be related to the insufficient expression of megakaryocytes proliferation-related genes, such as PF4 and PTFN4. Other studies indicated that rhG-CSF may aggravate the immune-mediated thrombocytopenia, which leads to hypersplenism and extensive activation of reticuloendothelial cells, thus increasing the consumption of platelets.In addition, the probability of severe thrombocytopenia was increased in allergic donors after rhG-CSF mobilization.In our report, although there was thrombocytopenia, mobilization was not affected.After the mobilization of rhG-CSF, platelets changes should be closely monitored to keep alert of donor bleeding tendency. While ensuring safe collection of hematopoietic stem cells by donors, patients should not be placed in extreme danger of not being able to receive allogeneic hematopoietic stem cells on time.
Key words:Severe thrombocytopenia;Healthy donor; rhG-CSF; Mobilization; Allogeneic peripheral blood stem cell transplantation.

Disclosures
No relevant conflicts of interest to declare.
Hyperglycemia Cooperates with Tet2 Heterozygosity to Induce Leukemia Driven By Pro-Inflammatory Cytokine Induced Lncrna Morrbid
Authors: Cai Zhigang , Kapur Reuben , Lu Xiaoyu , Zhang Chi , Nelanuthala Sai , Kotzin Jonathan , Williams Adam , Henao-Mejia Jorge , Aguilera Fabiola , Hadley Abigail , Fang Fang , Nephew Kenneth 
Published: November 2020

Diabetes mellitus (DM) is a risk factor for cancer development. However, the role of DM induced hyperglycemic stress (HG) in the development of blood cancer is poorly understood, largely due to lack of appropriate animal models. Epidemiologic studies show that individuals with DM are more likely to possess higher rate of mutations in genes found in pre-leukemic stem and progenitor cells (pre-LHSC/Ps) including in the epigenetic regulator TET2. TET2-mutant pre-LHSC/Ps require additional hits to evolve into a full-blown leukemia and/or aggressive myeloproliferative neoplasm (MPN). Cell intrinsic mutations have been shown to cooperate with Tet2 to promote leukemic transformation. However, the role of extrinsic factors is poorly understood. Utilizing a novel mouse model bearing haploinsufficiency of Tet2, to mimic the human pre-LHSC/P condition and HG stress, in the form of an Ins2Akita/+mutation, which induces HG and Type-1 DM, we show that the compound mutant mice develop a lethal form of MPN and/or acute myeloid leukemia (AML). RNA-seq revealed that this is in part due to upregulation of pro-inflammatory pathways, thereby generating a feed-forward loop, including the expression of an anti-apoptotic lncRNA Morrbid. Loss of Morrbid in the compound mutants rescues the lethality and mitigates the development of MPN/AML. Our results describe a novel mouse model for age-dependent AML/MPN and suggest that HG stress acts as an environmental driver for myeloid neoplasm, which could be effectively prevented by reducing the expression of inflammation-related lncRNA Morrbid.

Disclosures
No relevant conflicts of interest to declare.
An Ex Vivo Bioengineered Human Liver Microenvironment to Support Hematopoietic Stem Cell Maintenance and Expansion
Authors: Paik Na Yoon , Brown Grace E. , Shao Lijian , Sottoriva Kilian , Hyun James , Khetani Salman R. , Pajcini Kostandin V. 
Published: November 2020

Over 17,000 people require bone marrow transplants annually, based on the US department of Health and Human Services (https://bloodcell.transplant.hrsa.gov). Despite its high therapeutic value in treatment of cancer and autoimmune disorders, transplant of hematopoietic stem cells (HSC) is limited by the lack of sufficient source material due primarily inadequate expansion of functional HSCs ex vivo. Hence, establishing a system to readily expand human umbilical cord blood or bone marrow HSCs in vitro would greatly support clinical efforts, and provide a readily available source of functional stem cells for transplantation.
While the bone marrow is the main site of adult hematopoiesis, the fetal liver is the primary organ of hematopoiesis during embryonic development. The fetal liver is the main site of HSC expansion during hematopoietic development, furthermore the adult liver can also become a temporary extra-medullary site of hematopoiesis when the bone marrow is damaged. We have created a bioengineered micropatterned coculture (MPCC) system that consists of primary human hepatocytes (PHHs) islands surrounded and supported by 3T3-J2 mouse embryonic fibroblasts. Long-term establishment of stable PHH-MPCC allows us to culture and expand HSC in serum-free medium supplemented with pro-hematopoietic cytokines such as stem cell factor (SCF) and thrombopoietin (TPO). HSCs cultured on this PHH-MPCC microenvironment for two weeks expanded over 200-fold and formed tight clusters around the periphery of the PHH islands. These expanded cells also retained the expression of progenitor markers of Lin-, Sca1+, cKit+, as well as the long-term HSC phenotypic markers of CD48- and CD150+. In addition to the phenotypic analysis, the expanded cells were transplanted into lethally irradiated recipient mice to determine HSC functionality. The expanded cells from the PHH-MPCC microenvironment were able to provide multi-lineage reconstitution potential in primary and secondary transplants. With our bioengineered MPCC system, we further plan to scale up functional expansion of human HSC ex vivo and to better understand the mechanistic, cell-based niche factors that lead to maintenance and expansion HSC.

Disclosures
No relevant conflicts of interest to declare.
Oral Posaconazole or Intravenous-Oral Itraconazole in Preventing Invasive Fungal Diseases for Patients with Acute Leukemia
Authors: Liu Li , Pei Xiaolei , Ma Runzhi , He Yi , Zhang Rongli , Wei Jialin , Ma Qiaoling , Zhai Weihua , Jiang Erlie , Pang Aiming , Yang Donglin , Feng Sizhou , Han Ming-Zhe 
Published: November 2020

Background and Objectives:
Invasive fungal diseases (IFDs) are major and lethal infectious complications for patients with neutropenia after chemotherapy for acute leukemia (AL). We aimed to compare the use of oral posaconazole vs. itraconazole in an intravenous-oral regimen for primary antifungal prophylaxis in terms of efficacy, adverse events and costs among neutropenic patients with AL after chemotherapy.
Methods:
This single-center, retrospective study enrolled patients with AL. Prophylaxis with oral posaconazole or intravenous-oral itraconazole (intravenous formulation × 2 days + oral suspension) was administered for patients recovering from neutropenia after chemotherapy. Propensity-score matching was used to assemble a cohort of patients with similar baseline characteristics.
Results:
Among 342 eligible episodes from 186 patients, 110 episodes received oral posaconazole and 110 episodes received itraconazole in sequential regimen had similar propensity scores and were included in the analyses. The incidence of breakthrough IFDs was 0.0% (0/110) and 0.0% (0/110) in the posaconazole group and itraconazole group, while the incidence of proven, probable and possible IFDs was 1.8% (2/110) and 8.2% (9/110), respectively (P=0.030). In clinical failure analysis, the failure rate of posaconazole group was lower as compared to the itraconazole group (2.7% vs. 10.9%, P=0.016). The proportion of patients who needed systemic antifungal treatment were lower in posaconazole group than that in itraconazole group (2.7% vs. 10.0%, P=0.027). Five cases (4.5%) experienced adverse events possibly associated with posaconazole and thirteen cases (11.8%) with itraconazole (P=0.049). There was no significant difference of survival between patients on posaconazole or itraconazole. The acquisition costs of posaconazole were higher than those of itraconazole (P=0.000).
Conclusions:
Both oral posaconazole and intravenous-oral itraconazole are effective in preventing IFDs for acute leukemia patients recovering from neutropenia after chemotherapy, while posaconazole is slightly better. Oral posaconazole is safer and more tolerable but costs higher than intravenous-oral itraconazole.

Disclosures
No relevant conflicts of interest to declare.
Removal of Abnormal M Protein in Patients with Multiple Myeloma By Plasma-Free Exchange
Authors: Zhou Fuling , Liu Dandan , Wu Tianzhi 
Published: November 2020

ObjectiveBased on the albumin substitution fluid, we performed plasma-free exchange in the removal of abnormal M protein for patients with multiple myeloma using Fresenius COM.TEC machine
MethodsFrom February 2016 to August 2018, 70 patients with multiple myeloma, who have indication of plasma exchange because of increased M protein, were treated with plasma-free exchange. Record the relevant basic information, the clinical symptoms and related disease indicators, and use SPSS 20.0 to analyze the changes of erythrocyte sedimentation rate, blood routine, albumin, renal function, serum calcium and immunoglobulin, and report adverse reactions.
ResultsAfter treatment, the patient's erythrocyte sedimentation rate, serum calcium, serum creatinine, β2-microglobulin, uric acid, IgG, IgA were significantly reduced; hemoglobin, white blood cell level were not statistically significant. The creatinine remission rate was 97%, and the platelet decline was only 17.8%, and no adverse reactions related to thrombocytopenia such as hemorrhage and ecchymoses were observed. Patients with IgG type responded better than IgA-free treatment to plasma-free exchange. There were no serious adverse reactions after treatment, 1 case of blood pressure drop, 2 cases of low calcium reaction, all in the normal range, and recovered after the end of treatment.
ConclusionPlasma-free exchange is an effective technology reducing M protein, which can effectively improve the erythrocyte sedimentation index, renal function index and blood viscous state. Patients with IgG type or higher ISS stage have more obvious effects. Plasma-free exchange significantly get rid of the application of fresh frozen plasma.

Disclosures
No relevant conflicts of interest to declare.
CRISPR/Cas9 Chemogenetic Profiling Identifies Candidate Biomarker Genes That Modulate Sensitivity to Selinexor
Authors: Kwanten Bert , Landesman Yosef , Daelemans Dirk 
Published: November 2020

Introduction
Selinexor (KPT-330) is a first-in-class selective inhibitor of XPO1-mediated nuclear export that has recently been approved by the US FDA for the treatment of relapsed/refractory multiple myeloma and relapsed diffuse large B-cell lymphoma. Additionally, the drug is being evaluated in ongoing clinical trials (phase I to III) on late-stage haematological and solid tumours. To improve outcomes and personalize therapeutics, specific biomarkers and underlying mechanisms of response to treatment need to be uncovered.
Material and method
We performed genome-wide CRISPR/Cas9-mediated loss-of-function genetic screening in two multiple myeloma (KMS-28-BM and SK-MM-1) and one chronic myeloid leukemia (HAP1) cell lines. After transduction with a knockout library, cells were divided and subsequently cultured into three parallel groups (control, IC20 and IC50) for at least 20 cell doublings. This setup allows monitoring of subtle relative changes between groups. After DNA extraction, sequencing of guide RNAs and statistical analysis, we identified genes that upon disruption confer either increased susceptibility or resistance to selinexor.
Results and discussion
For the multiple myeloma cell lines, the TGF-β/SMAD4 signalling pathway stood out as an important mediator of resistance to selinexor. Besides SMAD4 as a major hit in both cell lines, we identified five other genes from this pathway leading to resistance in KMS-28-BM cells, suggesting that downregulation of this pathway is a potential biomarker of response to SINE treatment in multiple myeloma. In addition, screens revealed significant variability across cell lines with hit genes involved in mRNA processing, proteasomal degradation and cell cycle regulation. Interestingly, we could simultaneously uncover resistance and sensitivity genes involved in the same pathway, suggesting that these have antagonistic effects. Notably, we identified knockout of the poorly characterized gene ASB8 as a strong, common sensitizer between the three cell lines.
Conclusion
Until now, most chemogenetic CRISPR/Cas9 knockout screens have focused on determining resistance genes, and few have employed the technique to screen for genes that enhance cell kill. We performed such screens with selinexor and found a number of general and cell type-specific hits. Genes identified to confer increased resistance are candidate biomarkers while genes inducing increased susceptibility represent new targets for drug combination therapies.

Disclosures
Kwanten: Karyopharm Therapeutics Inc.: Patents & Royalties: employees of KULeuven. KULeuven has a license agreement (royalties) with Karyopharm Therapeutics on XPO1 inhibitors (selinexor). Landesman:Karyopharm Therapeutics Inc: Current Employment, Current equity holder in publicly-traded company. Daelemans:Karyopharm Therapeutics Inc.: Patents & Royalties: employees of KULeuven. KULeuven has a license agreement (royalties) with Karyopharm Therapeutics on XPO1 inhibitors (selinexor).
Utilizing Medical Students As Patient Mentors: An Intervention to Improve Transition Readiness in Young Adults with Sickle Cell Disease
Authors: Viola Adrienne , Drachtman Richard A. , Kaveney Amanda , Sridharan Ashwin , Savage Beth , Porter Jerlym S. , Devine Katie 
Published: November 2020

Background:Advances in medical care have resulted in nearly 95% of all children with sickle cell disease (SCD) living to adulthood. Standardized transition programming does not currently exist, resulting in high rates of mortality and morbidity among young adults (YA) during the transition period. Mentoring and social support have been used to impact health behavior change. Medical student mentors can serve as mentors offering specialized support for YA. This study examined the feasibility and preliminary efficacy of a medical student mentor intervention to improve transition outcomes for YAs with SCD.
Methods:24 YA with sickle cell disease and 9 medical student mentors enrolled in the intervention. Feasibility and acceptability of the intervention was assessed through enrollment rates, reasons for refusal, retention rates, engagement with the intervention, satisfaction, and reasons for drop-out. The preliminary efficacy of the intervention among patient participants was assessed using dependent t-tests to evaluate changes in transition readiness, health related quality of life, self-efficacy, SCD knowledge medication adherence, and health literacy. Among medical student mentor participants, changes in attitudes towards chronic illness and SCD knowledge were explored.
Results:Patient participants demonstrated adequate enrollment (63.2%), retention (75.0%), and adherence to the intervention (88.3%) and rated the intervention components highly. Patient participants demonstrated significant improvements in transition readiness (p= .001), self-efficacy (p= .002), medication adherence (p= .02), and health literacy (p= .05). Medical students also demonstrated significant improvements in SCD knowledge (p= .01).
Discussion:A medical student mentor intervention to facilitate transition from pediatric to adult care for young adults with SCD is both feasible and acceptable to patients and medical students. Preliminary evidence suggests such an intervention may provide dual benefit for both patients and students. A randomized controlled trial is needed to evaluate efficacy.

Disclosures
No relevant conflicts of interest to declare.
CD33 Targeting Primary CAR-NK Cells Generated By CRISPR Mediated Gene Insertion Show Enhanced Anti-AML Activity
Authors: Naeimi Kararoudi Meisam , Likhite Shibi , Elmas Ezgi , Schwartz Maura , Sorathia Kinnari , Yamamoto Kenta , Chakravarti Nitin , Moriarity Branden S , Meyer Kathrin , Lee Dean Anthony 
Published: November 2020

Human peripheral blood natural killer (NK) cells have intense antitumor activity and have been used successfully in several clinical trials. Modifying NK cells with a chimeric antigen receptor (CAR) can improve their targeting and increase specificity. Recently, we described an efficient method for gene targeting in NK cells using Cas9/ribonucleoprotein complexes (PMID: 29985369 and 32603414). Here we combined this approach with single-stranded (ss) or self-complementary (sc) Adeno-associated virus (AAV)-mediated gene delivery for gene insertion into a safe-harbor locus using a wide variety of homology arms for homology repair (HR) and non-homologous directed CRISPR-assisted insertion tagging (CRISPaint) approaches. We demonstrated that expansion of NK cells on feeder cells (CSTX002) expressing membrane-bound IL21 increases expression of HDR-related genes and provides optimum biological condition for targeted gene insertion. For proof-of-concept, we successfully generated stable mCherry-expressing primary NK cells (up to 89% mCherry+) and determined that sc vectors with 300bp homology arms were optimal. Then, we generated CD33-targeting CAR NK cells with differing transmembrane and signaling domains (CD4/4-1BB+CD3ζ and NKG2D/2B4+CD3ζ), which continued to show robust expansion on CSTX002 and stably maintained their CAR expression. This resulted in CAR-NK-cells of high number and purity (mean 68% CAR+) that demonstrated enhanced antileukemic activity against acute myeloid leukemia (AML) cell lines. This efficient method for site-directed insertion of genetic materials into primary NK cells has broad potential for fundamental discovery and therapeutic applications.
Keywords: CRISPR, NK, Cas9/RNP, AAV6, CRISPaint, HR, CD33CAR-NK
Figure

Disclosures
Naeimi Kararoudi: Kiadis Pharma Netherlans B.V: Patents & Royalties. Lee:Kiadis Pharma Netherlands B.V: Consultancy, Current equity holder in publicly-traded company, Membership on an entity's Board of Directors or advisory committees, Patents & Royalties.
Integrin αIIbβ3 Regulates Platelet-Procoagulant Activity in the Lung
Authors: Brzoska Tomasz , Kaminski Tomasz W. , Vats Ravi , Tutuncuoglu Egemen , Gladwin Mark T , Sundd Prithu 
Published: November 2020

Pulmonary thrombosis is a major complication associated with high morbidity. Despite advances in diagnosis and treatment, the pathophysiology of pulmonary thrombosis remains incompletely understood. New clinical evidence suggests that in situ platelet activation resulting in enhanced procoagulant activity may promote pulmonary thrombosis. Improved understanding of the etiological mechanism would enable the development of new therapies for pulmonary thrombosis.
Collagen and thromboplastin (TF) were administered intravascularly (IV) to C57BL/6 (WT) mice and the pulmonary microcirculation was visualized using quantitative fluorescence intravital fluorescence lung microscopy (qFILM). Fluorochrome-conjugated anti-mouse CD49b Ab and dextran was administered IV for in vivo staining of circulating platelets and visualization of blood vessels, respectively. Pulmonary thrombosis was defined as occlusion of blood vessels with platelet aggregates leading to pulmonary ischemia. Additionally, quantitative microfluidic fluorescence microscopy (qMFM) was used to study the effect of platelet αIIbβ3 inhibition on platelet procoagulant activity in human blood under vascular mimetic flow conditions.
Collagen and TF triggered dose-dependent pulmonary thrombosis in mice in vivo, which involved development of platelet-rich thrombi in the pulmonary arteriolar bottlenecks (junction of pulmonary arteriole and capillaries), resulting in a transient ischemia in the arteriole and the down-stream capillary tree. The pulmonary arteriole thrombosis triggered by IV collagen or TF was protracted, lethal and completely abrogated following IV administration of αIIbβ3 receptor inhibitor (eptifibatide). Inhibition of platelet αIIbβ3 also significantly reduced platelet procoagulant activity, fibrin formation and thrombus formation in human blood flowing through microfluidic channels ex vivo.
Our current findings suggest that αIIbβ3-dependent platelet procoagulant activity promotes pulmonary thrombosis. Both our models have potential application in investigating the molecular determinants of pulmonary thrombosis in diverse pulmonary disorders as well as evaluating efficacy of new antithrombotic drugs.

Disclosures
No relevant conflicts of interest to declare.