bluuebunny/crossref_metadata_2025_split huggingface dataset:
Hematological Profile in the Saudi Population: Reference Intervals By Gender, Age and Regions
Authors: Alaskar Ahmed , Rehan Hina , Mendoza May Anne , Alsahan Aljowhara , Immanuel Anita , Alhejazi Ayman , Abuelgasim Khadega , Salama Hind , Damlaj Moussab , Gmati Giamal , Alzahrani Mohsen , Rather Mushtaq , Alahmari Bader , Al Mugairi Areej , Ahmed Anwar 
Published: November 2019

Background: Although the prevalence of genetic hematological disorders varies widely between geographical regions, region-specific hematological reference ranges have not been defined in Saudi Arabian adults. Methods: A multicenter retrospective cross-sectional study was conducted with 1127 participants who completed their pre-employment screening and recruitment process between January 2013 and December 2016. Data related to the demographic and hematological indices were extracted. Results: The mean age was 28.0±5.2 years (range 19.4-72.8 years) and gender was distributed equally (47.5% female vs. 52.5% male). The WBC reference range was 3.3-11.4 ×109/L; hemoglobin 111-174 gm/l; platelet 163-412 ×109/L; MCV 80-95.7 fl, and neutrophils 1.2-8.8 ×109/L. A robust regression model was used to evaluate the effect of the participant's characteristics on the hematological indices. Except for WBC, the rest of the hematological indices were significantly influenced by gender, region, and age. The 2.5 percentile hemoglobin values were 135 gm/L in males and 104 gm/L in females, while platelet values were 173 x109/L in females and 159 x109/L in males. Conclusion: The study defined local hematological reference ranges, which were mostly lower than reported in international studies used in our center. Hematological values were mainly influenced by gender and region. A community nationwide screening study is required to create reference ranges specifically for the Saudi population.

Disclosures
No relevant conflicts of interest to declare.
Activation of JAK2/STAT5 Pathway Reduces Expression Level of DNMT3a in MPN Cell Line
Authors: Zhou Jie , Liang Aibin , Li Shaoguang , Zhang Wenjun , FU Jianfei 
Published: November 2019

Introduction: Myeloproliferative neoplasms (MPN) are clonal hematopoietic stem cell (HSC) disorders characterized by overproduction of mature blood cells and increased risk of transformation to acute myeloid leukemia (AML), and JAK2V167F is the most frequent MPN driving mutation detected in >95% of PV and 50-60% ET and PMF. DNMT3A is a de novo DNA methyltransferase that catalyzes the addition of methyl groups into active chromatin in CpG-rich regions leading to gene inactivation. Dnmt3a-/- HSC have enhanced self-renewal and a block in differentiation in vivo. Previous study showed that JAK2V617F and Dnmt3a loss cooperate to induce myelofibrosis through activated enhancer-driven inflammation, while whether JAK2V617F regulates DNMT3a still remains unclear. AZ960 is a potent and selective ATP competitive inhibitor of the JAK2 kinase, and previous studies reported that AZ960 possessed the activity selectively against JAK2. LY2784544 has been identified as a selective inhibitor of JAK2V617F and has undergone clinical trials for the treatment of several myeloproliferative disorders.
Methods: Empty vector (control) and mutant JAK2V617F were transduced into BaF3 cells using a lentivirus system. JAK2V617F-expressing BaF3 cells grow IL-3 independent and were selected by fluorescence-activated cell sorting (FACS) for GFP expression. The protein expression levels of p-STAT5 and DNMT3a were detected by western blotting. JAK2V617F-expressing and control BaF3 cells were incubated with gradient concentration of LY2784544 or AZ960 to inhibit JAK2/STAT5 pathway.
Results: The expression levels of p-STAT5 were obviously up-regulated in the JAK2V617F-expressing BaF3 cells, and DNMT3a was down-regulated. After 1-hour incubation in the serial diluted LY2784544, p-STAT5 were reduced in JAK2V617F-expressing BaF3 cells, with expression of DNMT3a elevated. To further confirm the correlation between JAK2/STAT5 pathway and expression of DNMT3a, another JAK2 inhibitor AZ960 was tested similar to LY2784544. With p-STAT5 expression suppressed, protein level of DNMT3a showed significantly promotion.
Conclusion: We observed that JAK2V167F mutation suppresses protein expression levels of DNMT3a in MPN cell lines. JAK2 inhibition by AZ960 and LY2784544 significantly improved expression levels of DNMT3a. The activation of JAK2/STAT5 pathway reduces expression level of DNMT3a in MPN cell line, and the specific mechanism still needs to be explored.
Figure

Disclosures
No relevant conflicts of interest to declare.
Deformability Based Sorting of Stored Red Blood Cells Reveals Donor-Dependent Aging Curves
Authors: Islamzada Emel , Matthews Kerryn , Guo Quan , Santoso Aline , Scott Mark D. , Ma Hongshen 
Published: November 2019

A fundamental challenge in the transfusion of red blood cells (RBCs) is that not all donated RBC units will confer the same benefit to the recipient. While some RBC units will remain in circulation for long periods after transfusion maintaining homeostasis, some will be cleared rapidly, leading to the need of an increased number of transfusions. This variability stems, in part, from the inherent ability of donor RBCs to withstand the physical and chemical insults of cold storage, which ultimately dictate their survival in circulation. The loss of RBC deformability during cold storage is well-established and has been identified as a potential biomarker for the quality of donated RBCs. Previous methods for characterizing RBC deformability have been limited in their sensitivity and consistency. As a result, these methods have only been able to characterize pathological and chemically degraded RBCs, but have not been able to distinguish differences between healthy donors.
Recently, we developed a microfluidic device to sort RBCs based on deformability using a matrix of asymmetrical tapered constrictions that form microfluidic ratchets. Due to the geometric asymmetry of the taper, the force required to deform cells through the constriction along the direction of taper is less than against the direction of taper. Coupling this deformation asymmetry with an oscillatory flow creates a ratcheting effect that selectively transports cells based on their ability to squeeze through each microscopic constriction. Importantly, this oscillatory flow also minimizes the contact between cells and the filter microstructure to prevent clogging and fouling to ensure that a consistent filtration force is applied to each cell.
We measured the deformability profiles of eight healthy donors using the microfluidic ratchet device. We found the deformability profile to be consistent for each donor over multiple donations. We also found significant variability across different donors. We then cold stored donated RBCs in SAGM media in plastic test tubes for 14 days to accelerate cold storage related damage. We find that the aging curve of RBC deformability varies significantly across donors, but is also consistent for each donor over multiple donations. Specifically, certain donors seem capable of providing RBCs that maintain their deformability during two weeks of accelerated aging.
This study illustrates a potential route to identify high-quality donors, or super-donors, that can provide RBCs that maintain their integrity during cold storage. Being able to identify these donors will enable blood bankers to reserve high-quality RBC units for chronic transfusion recipients, which will reduce the total number of transfusions for these patients and increase the overall blood supply.

Disclosures
Ma: Patent (US 9880084): Other: Inventor.
The Role of Protein Tyrosine Phosphatase PTP4A3 in T-Cell Acute Lymphoblastic Leukemia Progression
Authors: Wei Min , Blackburn Jessica 
Published: November 2019

The tyrosine protein tyrosine phosphatase PTP4A3 has been extensively reported to play a causative role in numerous cancers, including several types of acute leukemia. We found PTP4A3 to be highly expressed in T-cell Acute Lymphoblastic Leukemia samples, and show that PTP4A3 accelerates T-ALL onset and increases the invasive ability of T-ALL cells in a zebrafish model, and is required for T-ALL engraftment and progression in mouse xenograft. Our in vitro studies showed that PTP43A3 enhances T-ALL migration, in part via modulation of SRC signaling. However, whether SRC is a direct substrate of PTP4A3, and whether the phosphatase activity of PTP4A3 actually plays a role in T-ALL or other types of leukemia progression is unknown and remains a major question in the field.
We used a BioID-based proximity labeling approach combined with PTP4A3 substrate trapping mutant pull down assay to capture the PTP4A3 substrates candidates. BioID, a biotin ligase, was fused to PTP4A3 to generate a Biotin-PTP4A3 (BP) fusion protein. The overexpression of BP in T-ALL cell lines led to biotin modification of 288 PTP4A3 proximal proteins, including the potential direct PTP4A3 substrates. PANTHER pathway analysis showed that PTP4A3 interacting proteins are largely clustered in the T-cell activation, PDGF signaling, and angiogenesis. We are in process of validating potential substrates using immunoprecipitation and phosphoenrichement assays.
Finally, we are using a novel zebrafish Myc+PTP4A3 induced T-ALL model to assess the function of PTP4A3 in leukemia progression. We have created several PTP4A3 protein mutants, including a phosphatase-dead mutant, a mutant unable to bind magnesium transporter, and a prenylation deficient mutant, and are in process of assessing the effects of these mutants in T-ALL onset and progression in our in vivo model. In total, these studies will allow us to better understand function of PTP4A3 in T-ALL progression, and may provide a strong rationale for the development of PTP4A3 inhibitors for use in leukemia.

Disclosures
No relevant conflicts of interest to declare.
A Refractory B-ALL Patient Relapsed with Leukemia Cells Expressing CD19 and Anti-CD19 CAR Gene 14 Days after CR from Anti-CD19 CAR T-Cell Therapy
Authors: Meijing Liu , Cui Rui , Mu Juan , Yuan Ji-jun , Mou Nan , Yang Zhen-xing , Yan-yu Jiang , Juanxia Meng , Deng QI 
Published: November 2019

Purpose: To investigate the immunophenotypic characteristics of CD19 CAR transfected into leukemia cells with CD19 expression during CAR-T cell preparation.
Patient and Methods: Here we reported a TP53 positive refractory B-ALL patient relapsed 14 days after Complete response (CR) from the CD19 CAR-T therapy. But her leukemia cells expressed CD19 and expressed aberrantly the anti-CD19 CAR gene at the same time. We transfected CD19 CAR gene into Nalm-6 cells to detect the expression of CD19 CAR Nalm-6 cells and the sensitivity to anti-CD19 CAR T-cell and anti-CD22 CAR T-cell. The binding of CD19 antigen on the surface of CD19 CAR Nalm-6 cells and Nalm-6 cells to CD19 CAR in CD19 CAR Nalm-6 cells was observed by confocal microscopy. The proportion of Nalm-6 cells in the transfection system was increased gradually to observe the change of CD19 expression on CD19 CAR Nalm-6 cells.
Results: The patient received a CD22 CAR-T therapy followed by a haploid hematopoietic stem cell transplantation and achieved CR with vanishment of TP53 mutation. To date, she had leukemia-free survival (LFS) and complete donor chimerism for 8 months. We found that there had 1.41% leukemic cells in the peripheral blood mononuclear cells (PBMC) during anti-CD19 CAR T-cell preparation, which may contribute to the relapse of the first CD19 CAR-T therapy. The CD19 expression was only 1.19% in CD19 CAR Nalm-6 cell culture system with a transfection rate of 46.5±3.78%, while the proportion of CD19 expression increased gradually when we increased the proportion of Nalm-6 cells in the transfection system. CD19 CAR of CD19 CAR Nalm-6 cells occupied the CD19 expression site of the cells themselves, meanwhile occupied the CD19 expression site of Nalm-6 cells. We found that the CD19 CAR Nalm-6 cells were sensitive to anti-CD22 CAR T-cell, althought they were resistant to anti-CD19 CAR T-cell.
Conclusions: There had a presence that CD19 CAR gene was transferred into leukemia cells during T cell manufacturing. The immunophenotype of leukemia cells of these refractory B-ALL patients who relapsed after CR from anti-CD19 CAR T-cell therapy were CD19+ or CD19-. It was determined by the proportion of CD19 CAR leukemia cells and leukemia cells. Anti-CD22 CAR T-cell therapy could be as a salvage measure to such patients.
Figure

Disclosures
No relevant conflicts of interest to declare.
Loss of IRF4 Results in Multiple Myeloma Cell Apoptosis through the Transcriptional up-Regulation of the BH3-Only Proteins Bmf and BIM
Authors: Fedele Pasquale L. , Liao Yang , Gong Jianan , van Delft Mark , Low Michael S.Y. , Tai Lin , Herold Marco J , Tellier Julie , Grigoriadis George , Huang David C.S. , Shi Wei , Nutt Stephen , Willis Simon 
Published: November 2019

The transcription factor IRF4 is essential for the survival of both normal plasma cells and the plasma cell malignancy multiple myeloma (MM). However the basis for this dependency remains uncertain, with the prevailing view that IRF4 loss in MM induces "death by a thousand cuts."
Here we have explored the genetic basis for IRF4 addiction through CRISPR-Cas9 mediated deletion of IRF4 in three lenalidomide sensitive MM cells lines (MM1S, OPM2 and H929) and one resistant cell line (RPMI-8226). Loss of IRF4 resulted in marked loss of viability, irrespective of lenalidomide sensitivity, which was driven by two distinct mechanisms: 1) induction of apoptosis, which was rescued through prior deletion of apoptotic mediators BAX and BAK, and 2) proliferative arrest, which persisted despite rescue from apoptosis.
RNA-sequencing following IRF4 inactivation identified a core group of 86 genes whose dysregulation was shared across all 4 MM cell lines. We identified 31 common down-regulated genes (IRF4 activated), including several with previously described roles in survival (such as KLF2, TNFRSF17/BCMA and MYB). Furthermore, MYC, a previously identified target of IRF4 known to play an important role in MM pathogenesis and survival, was down-regulated in 3 of the 4 cell lines.
Most surprisingly, IRF4 inactivation resulted in 55 common up-regulated genes that included two BH3-only pro-apoptotic proteins, BMF and BCL2L11 (BIM). Up-regulation of BMF appeared more marked than BIM with an average fold change of 6.5x (range 2.7 - 10.3) vs 1.9x (range: 1.8 - 2.2) respectively. Remarkably, genetic ablation of BMF in the OPM2 cell line and BMF/BIM in the MM1S largely protected from the cell death observed following IRF4 inactivation, suggesting that IRF4 maintains MM survival through the transcriptional repression of BMF and BIM.
Our results confirm the vital role of IRF4 for MM proliferation and survival, and shed further light into its critical downstream transcriptional targets. Most interestingly, we have demonstrated that the 'killer blow' appears to be the concerted effect of the BH3-only pro-apoptotic proteins BMF and BIM.

Disclosures
Huang: Genentech: Patents & Royalties: DCSH is an employee of the Walter and Eliza Hall Institute which receives milestone and royalty payments related to venetoclax.
Implementing a Structured Health Care Transition (HCT) Process for Sickle Cell Disease (SCD) Using Quality Improvement (QI): Early Results of a SCD HCT Learning Collaorative
Authors: Osunkwo Ifeyinwa (IFY) , Cheryl Courtlandt , Patterson Charity , Laura Noonan , Cornette Jennifer , McManus Peggy , Patience White , Lawrence Raymona Holloway 
Published: November 2019

Background: A regional SCD HCT learning collaborative (SCD-LC) was formed from engaging 14 pediatric and adult paired sites (including required community participation) using a standardized HCT quality improvement process. The SCD-LC involved a kick-off meeting and monthly calls to establish a common understanding of clinical recommendations, QI methods, and implementation of the Six Core Elements of HCT (6CE).
Methods: Implementation of 6CE was measured as part of a SCD HCT Study conducted in 14 paired SCD clinical sites with pediatric, adult and community partners who care for 1625 individuals aged 16-25. They represent a variety of clinical programs:12 urban and 2 rural, 12 academic and 2 nonacademic, 6 small and 8 large. Got Transition's Process Measurement Tool (HCTPMT) was customized for SCD and used to assess 6CE implementation. HCTPMT were completed by 14 pediatric and 14 adult sites at baseline (2018) and after 6 months. Pre-post results were compared overall and by type of practice.
Results: All 14 participating sites made substantial progress towards implementing a structured HCT process consistent with the within the first 6 months of the project. Overall, baseline scores averaged 19.4 (SD 14.1) and after 6 months of QI support, scores increased to 33.7 (SD 19.4) (average change 14.3, p<0.0001). For the 14 pediatric sites, scores went from 24 to 43 (p 0.0016), and for the 14 adult sites, from 15 to 24 (p 0.0017). Pediatric sites had higher scores at 6 months compared to adult sites (p=0.0068). Although their change over time was higher, it did not reach statistical significance at 0.05 (p=0.0759). Large sites went from 21 to 34 (delta 14, p=0.0048), small went from 18 to 32 (delta 15, p=0.0018).
Conclusion: All 14 pediatric and adult sites reported significant progress within 6 months of implementing a structured HCT process aligned with the 6CE.

Disclosures
Osunkwo: Novartis: Consultancy, Speakers Bureau; Pfizer: Consultancy; Terumo: Speakers Bureau; Micella Biopharma: Other: DSMB Member .
Pre-Emptive Tyrosine Kinase-Inhibitor Intervention Based on BCR/ABL1 Monitoring in Philadelphia Chromosome-Positive Acute Lymphoblastic Leukemia after Allo-HSCT: Two Independent Observational Cohort Study
Authors: Hui Liu , Xuan Li , Lin Ren , Deng Lan , Huang Fen , Fan Zhiping , Nie Danian , Li Xudong , Liang Xinquan , Xu Dan , Zhang Yu , Xu Na , Ye Jieyu , Jin Hua , Lin Dongjun , MA Liping , Sun Jing , Huang Xiaojun , Liu Qifa 
Published: November 2019

Relapse is a major cause of treatment-failure in persons with Philadelphia-chromosome-positive acute lymphoblastic leukemia (Ph+ALL) receiving an allogeneic hematopoietic cell transplant. There is controversy whether post-transplant tyrosine kinase-inhibitors (TKIs) reduces relapse risk. We evaluated a pre-emptive strategy in 103 subjects in a prospective study with prophylactic group as historical control. Subjects were tested for measurable residual disease (MRD) by assaying levels of BCRABL1 transcript. MRD was considered positive if the levels of BCRABL1 transcript were >=10-5. Subjects with positive MRD post-transplant received TKI-therapy. The chosen of TKIs was based on BCRABL1 mutation, and donor lymphocyte infusion (DLI) was recommended in those who were not responsive to TKIs. The primary endpoint was relapse (including hematological relapse and central nervous system relapse). Twenty-three of 103 subjects were MRD-positive post-transplant. Seven of whom had BCRABL1 mutations including T315I (N=5). Pre-emptive therapy resulted in 21 subjects being MRD-test negative. The overall 3-year cumulative incidence of relapse (CIR) post-transplantation was 11% (95% confidence interval [CI], 4, 17%) and 34% (22,1, 46.5%) in the pre-emptive and prophylactic maintain group (P=0.030). The 3-year cumulative survival and LFS post-transplants in the pre-emptive and prophylactic group were 87% (80, 94%) and 63% (51, 75%), and 83% (76, 91%) and 58% (46, 70%) (P=0.000 and P=0.000, respectively). The multivariate analysis showed that positive MRD pre-transplantation was the risk factor for relapse, LFS and survival [HRs=8.977(3.344, 24.096; P=0.000), HRs=4.763 (2.153, 10.538; P=0.000) and HRs=7.535 (2.977,19.071; P=0.000], while pre-emptive therapy was the protective factor for relapse, LFS and survival [HRs=0.176 (0.077, 0.339; P=0.000), HRs=0.222 (0.107, 0.462; P=0.000) and HRs=0.188 (0.082,0.43; P=0.000)]. Our data suggest a pre-emptive strategy based on MRD-test monitoring and mutation analyses might reduce CIR and improve survival after allotransplants for Ph+ALL (NCT 01883219).

Disclosures
No relevant conflicts of interest to declare.
On Demand Plerixafor As a Rescue Strategy in Healthy Donors to Achieve Adequate Stem Cells - a Pilot Study from South India
Authors: Kumar Kishore , Kumar Kishore , Subash Chezhian 
Published: November 2019

Aims & Objectives: The aim of the study is to check the safety and feasibility of using Plerixafor in healthy donors who have failed to mobilise > 2 x 10 6/kg of CD34 cells on day 1 of aphresis. The indication at present remains off label.
Patients/Materials & Methods: Seventeen healthy donors, all more than 18 years of age were enrolled in this study after a proper written informed consent who failed to mobilise adequately on day 1 collection. The dose of Plerixafor used was 0.24mg/kg body weight of donor, 10-11 hours before the aphresis procedure. The GCSF was given from day -5 and the last dose was given 4 hours before the second day aphresis. CD34 was done in pre-aphresis peripheral blood and in product bag and compared with day 1 data.
Results: The median donor bw was 54 kg (IQR, 42 kg to 69 kg) and the median recipient bw was 58 kg (IQR, 44 kg to 76 kg).The median CD34+ count in peripheral blood was 11.2/µl on day 4 after G-CSF alone and 24.7.0/µl on day 5 after G-CSF plus Plerixafor. The use of Plerixafor increased the number of circulating CD34 cells in peripheral blood by 2.2 fold.There were no major side effects except for the manageable bone pains which can be attributed to GCSF also. There were no differences in the engraftment statistics and rates of GVHD in comparison to historical cases. Though this is not a randomised control study to compare with second day high dose GCSF cases, the amount of increase in stem cells were statistically significant comparing with historical controls(p< 0.05). After a median followup of six months, no adverse effects were noted in donors.
Discussion & Conclusion: Plerixafor is well tolerated in healthy donors and can be used safely in situations of poor mobilisers or when there is a significant difference in weight of donor to receipient. Its a small pilot study. We need a proper randomised control study and a longer follow up to look infor the side effects on healthy donor.

Disclosures
No relevant conflicts of interest to declare.