bluuebunny/crossref_metadata_2025 huggingface dataset:
Usefulness of Dilute Russell’s Viper Venom Clotting Time (DRVVT) for the Routine Monitoring of New Oral Anticoagulants
Authors: Hoppensteadt Debra , Kola Jake , Brayne Mark , Iqbal Omer , Fareed Jawed
Published: October 2019

The new oral anticoagulants such as rivaroxaban (Bayer Healthcare) (R) apixaban (BMS/Pfizer) (A) and dabigatran (Boehringer Ingleheim) (D) have been approved to manage thrombotic and cardiovascular disorders in the US and Europe. Initially it was thought that these agents would not require monitoring at the approved dosages for specific indications. However, there have been reported bleeding complications with some of these agents suggesting that monitoring of these drugs to optimize therapy in some patient populations may be needed. The purpose of this study was to determine which of the available clotting assays could be used to determine the circulating concentration of these new oral anticoagulants in plasma.

Materials: Citrated blood was drawn from ten donors and spun at 3000 rpm to obtain platelet poor plasma (PPP). The PPP was supplemented with A, R and D in a concentration range of 0-1.0 ug/ml. In addition, five plasma samples, for each agent, containing varying concentrations of either A, R or D were also blindly analyzed. The plasma samples were analyzed using PT/INR (Innovin, Seimens, Deerfield,IL), APTT reagent (Platelin, TCoag, Ireland) dilute Russell’s viper venom time (DRVVT) and DRVVT confirm (HemosIL, Instrumentation Laboratories (Bedford, MA).

Results: In the PT and APTT assays a concentration dependent increase in clotting time was observed, however the slope of the line was flatter than with the slope observed in the DRVVT and DRVVT confirm assays. In both the DRVVT and DRVVT confirm assays the concentration-response curves for A,D and R were straight lines with a steep slope. Therefore, it was possible to extrapolate the concentrations of the unknown samples using both of these assays. When using the DRVVT and DRVVT confirm assays , there was a good correlation between the calculated concentration and the actual concentration in the range of 50-1000 ng/ml (r2=0.82). At concentrations < 50 ng/ml, the calculated concentration showed a poor correlation with the actual concentration (r2=0.26).

Conclusions: These results suggest that the PT and APTT assays are not sensitive to the new oral anticoagulants and therefore cannot be used to determine the circulating concentrations of these agents. However, the dRVVT and DRVVT confirm assays are sensitive to A, D and R at concentrations > 50 ng/ml, therefore, A,D and R can be monitored by these assays. Both of these assays are commercially available and can be readily adapted to any automated instrument. These studies warrant clinical validation of these tests in patients populations treated with the newer oral anticoagulant drugs.

Disclosures

No relevant conflicts of interest to declare.
Comprehensive Kinase Profile of Pacritinib, a Non-Myelosuppressive JAK2 Kinase Inhibitor in Phase 3 Development in Primary and Post ET/PV Myelofibrosis
Authors: Singer Jack , Al-Fayoumi Suliman , Ma Haiching , Komrokji Rami S , Mesa Ruben , Verstovsek Srdan
Published: October 2019

BackgroundPacritinib, a JAK2, JAK2 V617F and FLT3 kinase inhibitor is in Phase III development in myelofibrosis. In preclinical models and in Phase 1 studies in both non-Hodgkin lymphoma and myelofibrosis, it appeared unique among this class of agents in its lack of myelosuppression at doses that inhibited the JAK2/STAT3 pathway and resulted in significant clinical efficacy. This potentially important clinical observation was verified in two Phase II studies in myelofibrosis that had no exclusion restriction for low platelets. Pacritinib was effective at reducing spleen size and symptoms regardless of starting platelet counts and there was no apparent decline in either platelet count or hemoglobin levels, or an increase in transfusion requirements.

Aims and Methods: To help elucidate the mechanisms for pacritinib’s lack of myelosuppression, a kinome screening analysis against 429 recombinant kinases at 100 nM pacritinib concentration was performed, followed by titration from 1-100nM against those that were >50% inhibited at 100nM (Anastassiadis et al.; Nature Biotechnology; 2011). In humans, pacritinib achieves steady state free drug levels of approximately 200nM at the standard 400 mg daily dose.

Results:All kinases with IC50 values of <50nM are included in the table.

Table Kinase IC50 (nM) JAK2; JAK2(V617F) 6.0; 9.4 FLT3; FLT3-ITD; FLT3 (D835Y) 6.4; 12.0; 8.3 TYK2 27.0 JAK3 18.3 IRAK1 (IL-1 receptor kinase) 13.6 HIPK4 (PT kinase for P53 ser-9) 14.5 FMS 39.5 c-KIT and most common mutations except (V560G), (V559D/V654A) 43.8 c-Src; (c-Src (T341M)) 47.0 (55.5)

Of note, pacritinib did not affect JAK1 (82% control at 100nM). As the concentrations of pacritinib achieved clinically are likely inhibit signaling the kinases listed in the table, these data suggest that the lack of myelosuppression with pacritinib may be either due to inhibition of JAK2 without affecting JAK1 or to suppression of IRAK1, an IL-1 receptor kinase, downstream from Toll-Like Receptors, and associated with the inflammatory response and suppression of normal hematopoiesis in myelodysplasia models (Rhyasen et al.; Cancer Cell; 24:90, 2013).

Conclusions: The kinase profile for pacritinib is unique among agents in development for myelofibrosis combining lack of suppression of JAK1 with suppression of inflammatory signaling through IRAK1. Moreover, the kinase inhibition profile of pacritinib suggests its potential therapeutic utility in AML, MDS, and particularly in CMML due to CMML’s upregulation of JAK2, c-FMS, and IRAK1.

Disclosures

Singer: CTI BioPharma, Corp: Employment, Equity Ownership. Mesa:Incyte Corporation, CTI, NS Pharma, Inc., Gilead Science Inc., Celgene: Research Funding. Verstovsek:CTI BioPharma, Corp: Research Funding.
The Psychosocial Transplant Evaluation Rating Scale (TERS) Prospectively Predicts Inferior Overall Survival Outcome for High Risk Scoring Patients Undergoing Allogeneic Hematopoietic Stem Cell Transplantation
Authors: Speckhart Dawn , Solomon Scott R. , Zhang Xu , Morris Lawrence E. , Bashey Asad , Holland H. Kent
Published: October 2019

Several studies have begun to examine the relationship between psychosocial variables and outcome in hematopoietic stem cell transplantation (HSCT). Previous studies have also looked at the relationship between the Hematopoietic Cell Transplantation Comorbidity Index (HCT-CI) and disease risk on overall survival in HSCT. Research has shown that disease risk and HCT-CI may have a negative relationship with outcome in HSCT. In this study, psychosocial data was collected prospectively on 437 patients undergoing related and unrelated allogeneic HSCT. All patients were assessed using the Transplant Evaluation Rating Scale (TERS). Based on the patient’s TERS score, each patient was stratified into one of two groups (low/moderate risk (n=371) vs. high risk (n=66)) based on their predicted psychosocial risk for problems during transplant. The 1-year overall survival (OS) for the low/mod risk TERS group versus the high TERS risk group was 78% and 61%, respectively (p=.005). Stratification by CIBMTR disease risk identified 261 patients with low and intermediate risk disease of which 228 had low/mod risk TERS score and 33 had high risk TERS score. The is a significant survival advantage for Low/Mod Risk TERS patients vs. High Risk TERS patients at 100 day, 1 year and overall survival being 97% vs 79% (P=.01), 85% vs 58% (P=.002) and 68% vs 42% (P=.002) with a median follow up of 48 months (graph attached). An adjusted logistic regression analysis showed that psychosocial factors have a significant impact on overall survival despite all patients being required to have 24 hour caregiver support, a pre-transplant psychosocial evaluation and post-transplant psychosocial intervention. The analysis controlled for other variables such as age, gender, performance status, disease risk and transplant type. These results highlight the impact of psychosocial factors on HSCT outcome, as those patients who are expected to do well based on their low/intermediate disease risk may not perform as expected due to psychosocial risk factors. These results suggest the need to perform careful psychosocial assessments on all HSCT patients in order to predict mortality risks for the individual patient and to help the transplant team strategize how to better manage the high risk psychosocial patient.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Loss of IRF8 Inhibits the Growth of Diffuse Large B-Cell Lymphoma
Authors: Xu Yulian , Jiang Lei , Fang Rachel R. , Zhou Jeff Xiwu , Morse Herbert
Published: October 2019

IRF8 is a transcription factor with a critical role in B lymphocyte development and biological functions. Although it has been reported that IRF8 is highly expressed in human diffuse large B-cell lymphoma (DLBCL) and the translocation of IRF8-IgH loci occurs in DLBCL, little information is available regarding the function and mechanisms for the role of IRF8 in DLBCL. In this study, by using several human DLBCL cell lines with shRNA-mediated decrease in IRF8 expression levels, we found that the loss of IRF8 significantly reduced the proliferation of lymphoma cells (Figure 1). Mechanistically, decreasing the levels of IRF8 led to a decrease in p38 and ERK phosphorylation (Figure 2), molecular events critical for B cell proliferation. Furthermore, using a xenograft lymphoma mice model, we found that the loss of IRF8 significantly inhibited the growth of lymphomas in vivo (n=5 for each group) (Figure 3). Analysis of public available data also suggested that the expression levels of IRF8 mRNA in human DLBCL tissues were inversely correlated patientsÕ overall survival time. Taken together, this study showed that IRF8 may play an oncogenic role in human DLBCL by promoting cell proliferation.

Figure 1. Loss of IRF8 decreased the proliferation of DLBCL cells in vitro. Figure 1. Loss of IRF8 decreased the proliferation of DLBCL cells in vitro.

Figure 2. Loss of IRF8 decreased the phosphorylation of p38 and ERK in DLBCL cells. Figure 2. Loss of IRF8 decreased the phosphorylation of p38 and ERK in DLBCL cells.

Figure 3. Loss of IRF8 decreased the growth of DLBCL in vivo. Figure 3. Loss of IRF8 decreased the growth of DLBCL in vivo.

Disclosures

No relevant conflicts of interest to declare.
Constitutive Activation of Shp2 Cooperates with MLL1 Fusion Proteins to Increase Expression of the Homeodomain Transcription Factors Cdx4, HoxA9 and HoxA10
Authors: Bei Ling , Shah Chirag , Wang Hao , Eklund Elizabeth
Published: October 2019

Leukemias with chromosomal translocation or partial tandem duplications involving the MLL (mixed lineage leukemia) gene have exceptionally poor prognosis (referred to as 11q23-leukemias). At the molecular level, 11q23-leukemias are characterized by aberrant expression of a set of homeodomain transcription factors in hematopoietic stem cells (HSC) and differentiating myeloid progenitor cells. This transcription factor set includes HoxB3, B4, A7-11, Cdx2-4 and Meis1. Cdx and Hox proteins are involved in regulating hematopoiesis. Transcription of HOX and CDX genes decreases normal myelopoiesis, but is aberrantly sustained in 11q23-leukemias. Cdx4 activates transcription of the HOXA9 and HOXA10 genes, and HoxA10 activates CDX4 transcription. The events that break this feedback loop, permitting a decrease in Cdx4-expression during normal myelopoiesis, were previously undefined. In the current study, we find that HoxA9 represses CDX4 transcription in differentiating myeloid cells; antagonizing activation by HoxA10. We determine that tyrosine phosphorylation of HoxA10 impairs transcriptional activation of CDX4, but tyrosine phosphorylation of HoxA9 facilitates repression of this gene. Since HoxA9 and HoxA10 are phosphorylated during myelopoiesis, this provides a mechanism for differentiation-stage-specific Cdx4 expression. HoxA9 and HoxA10 are increased in cells expressing Mll-Ell, a leukemia associated MLL1 fusion protein. We find that Mll-Ell induces a HoxA10-dependent increase in Cdx4-expression in myeloid progenitor cells. However, expression of Cdx4 decreases in a HoxA9-dependent manner upon exposure of Mll-Ell-expressing cells to differentiating cytokines. Leukemia associated, constitutively active mutants of Shp2 block cytokine-induced tyrosine phosphorylation of HoxA9 and HoxA10. In comparison to cells expressing Mll-Ell alone, we find that co-expression of Mll-Ell plus constitutively active Shp2 increases CDX4 transcription and Cdx4 expression in myeloid progenitor cells. And, increased Cdx4 expression is sustained upon exposure of these cells to differentiating cytokines. Our results identify a mechanism for increased and sustained CDX4 transcription in leukemias co-overexpressing HoxA9 and HoxA10 in combination with constitutive activation of Shp2. We also demonstrate that inhibition of Shp2-PTP activity decreases Cdx4 expression in Hox-overexpressing human myeloid leukemias.

Disclosures

No relevant conflicts of interest to declare.
Four-Factor Prothrombin Complex Concentrate (4F-PCC) Effectively Reverses Rivaroxaban Induced Bleeding in a Rabbit Model of Acute Bleeding
Authors: Herzog Eva , Mueller-Cohrs Jochen , Kaspereit Franz , Krege Wilfried , Niebl Peter , Doerr Baerbel , Schulte Stefan , Dickneite Gerhard
Published: October 2019

Rivaroxaban is an oral, selective direct factor Xa inhibitor approved for several indications in patients at risk of thrombotic events. One limitation of this agent is the lack of data pertaining to its reversal in situations where urgent response is critical (e.g. acute bleeding events or emergency surgery).

This study therefore evaluated the effectiveness of a four-factor prothrombin complex concentrate (4F-PCC; Beriplex® P/N, Kcentra®, CSL Behring), for the reversal of rivaroxaban-associated bleeding in an in vivo rabbit model of acute bleeding. In addition, the study evaluated the correlation between in vitro coagulation parameters and haemostasis in vivo in order to gain more information on the predictivity of in vitro markers for effective bleeding reversal.

Administration of single intravenous doses of rivaroxaban (150–450 μg/kg) resulted in increased and prolonged bleeding following standardised kidney injury compared to a vehicle treated control group as determined by measurements of total blood loss and time required to achieve full hemostasis. Subsequent treatment with 4F-PCC (25−100 IU/kg) resulted in a dose-dependent reversal of rivaroxaban-associated bleeding signals. Of the in vitro coagulation markers tested, thrombin generation (TGA) and whole blood clotting time (WBCT) correlated well with in vivo measures of PCC-mediated effects. Thrombin generation was highly reagent-dependent, with the assay initiated using tissue factor reagent being most sensitive to the anticoagulant effects induced by Rivaroxaban, while the phospholipid-only reagent being the most predictive of 4F-PCC mediated effective haemostasis in vivo.

In summary, in a rabbit model of acute bleeding, 4F-PCC was able to effectively reduce bleeding to control levels following rivaroxaban 150 μg/kg and 300 μg/kg administrations. The 4F-PCC mediated bleeding reversal correlated best with the in vitro endpoints thrombin generation and whole blood clotting time.

Disclosures

Herzog: CSL Behring GmbH: Employment. Off Label Use: The use of Beriplex P/N for reversal of Rivaroxaban anticoagulation represents off-label use. Mueller-Cohrs:CSL Behring GmbH: Employment. Kaspereit:CSL Behring GmbH: Employment. Krege:CSL Behring GmbH: Employment. Niebl:CSL Behring GmbH: Employment. Doerr:CSL Behring GmbH: Employment. Schulte:CSL Behring GmbH: Employment. Dickneite:CSL Behring GmbH: Employment.
The Use of Recombinant FXIII in a Major Acute Bleeding Episode of a Patient with Congenital FXIII Deficiency – the First Experience
Authors: Boda Zoltan , Arokszallasi Anita , Kerenyi Adrienne , Bereczky Zsuzsanna , Muszbek Laszlo , Pfliegler Gyorgy , Schlammadinger Agota
Published: October 2019

Introduction: congenital FXIII deficiency is a rare bleeding disorder with a high risk of spontaneous intracranial haemorrhage. Life-long preventive supplementation is mandatory if FXIII activity is <1%. A novel recombinant FXIII concentrate (NovoThirteen inj.) was licensed for prophylaxis in congenital FXIII-A subunit deficiency. The efficacy and safety of rFXIII in acute bleeding have not been tested.

Aim: to report the effective use of rFXIII for the treatment of a major bleeding in a patient with severe inherited FXIII-A deficiency.

Patient and methods: a 19-year-old man exhibiting severe bleeding phenotype without clear diagnosis was referred to our haemostasis centre. After presenting negative haemostasis screening tests he was evaluated for FXIII deficiency by the determination of FXIII activity, FXIII-A2B2, FXIII-A and FXIII-B concentrations and by mutation analysis.

Results: severe FXIII-A deficiency was established and prophylaxis with rFXIII was planned. Prior to the start of the prophylaxis he was hospitalized due to a large intramuscular hematoma (19.0x8.3x7.7 cm) in the right thigh and rFXIII, in the prophylactic dose (35 IU/kg) was administered for therapy. Following the administration of rFXIII, the pain disappeared and the swelling of the thigh considerable decreased. Complete resolution of the hematoma occurred without any additional FXIII supplementation. Results of the pharmacokinetic assessment were comparable with those described in the literature.

Conclusion: rFXIII can be successfully used in acute bleeding episode of severe congenital FXIII deficiency.

Disclosures

No relevant conflicts of interest to declare.
Intracellular Expression of Neurotrophic Factors in Mononuclear Cells of Cord Blood and G-CSF-Mobilized Peripheral Blood As a Potential Source for Cellular Therapy
Authors: Lee Young-Ho , Koh Hani , Hwang Kyoujung
Published: October 2019

Purpose: To investigate a possible therapeutic mechanism for cell therapy in the field of neurological disorders using mobilized peripheral blood mononuclear cells (mPBMCs), we compared the expression of inflammatory cytokines and neurotrophic factors in PBMCs and mPBMCs from children with cerebral palsy (CP) to those from healthy adult donors and to cord blood (CB) donated from healthy newborns.

Methods: We evaluated the intracellular expression of neurotrophic factors and inflammatory cytokines in PBMCs and mPBMCs from 14 CP children and 14 healthy adult volunteer donors as well as CB mononuclear cells (CB-MNCs) donated from healthy newborns. For mPBMC, granulocyte colony stimulating factor (G-CSF, 10μg/kg/day) was administered subcutaneously for five consecutive days, and apheresis was performed on the fifth day to collect mPBMC via a central venous catheter. Both PBMC collected prior to G-CSF administration and apheresed mPBMC were cryopreserved. We performed flow cytometric analysis with intracellular staining for various cytokines after thawing PBMCs and mPBMCs as well as CBs.

Results: No significant differences in expression of neurotrophic factors were found between PBMC and mPBMC. However, in cells from CP children, the expression of IL-6 was significantly increased in mPBMC as compared to PBMC, and IL-3 was significantly decreased in mPBMC as compared to PBMC. In healthy adults, the expression of both IL-1β and IL-6 were significantly increased in mPBMC as compared to PBMC. The expression of BDNF in mPBMC from CP children was significantly higher than in CB or mPBMC from healthy adults. The expression of G-CSF in mPBMC from CP children was comparable to that in CB but significantly higher than in mPBMC from healthy adults. Lower expression of IL-1β, IL-3, and IL-6 and higher expression of IL-8 and IL-9 was observed from CB and mPBMCs from CP children rather than in healthy adults. Lower expression of IL-1β, and higher expression of IL-8 was observed from mPBMCs from CP children rather than in healthy adults.

Conclusion: The altered expression of neurotrophic factors and anti-inflammatory cytokines in mPBMC in CP children and CB from healthy children could provide a new potential source for cellular therapy for individuals with neurologic diseases.

Disclosures

Lee: Korea Healthcare Technology R&D Project (A101712), Ministry for Health & Welfare, Republic of Korea: Research Funding.
Study on Leukemia Cell Differentiation and Drug Resistance By SATB1 Gene
Authors: Wang Chunyan , Tan Huo , Lei Liu , Xu Lihua
Published: October 2019

Background and objective:

In this article, four types of lymphoma cell lines U937, Raji, Hut-102, Akata, and three types of leukemia cell lines K562, HL-60, Jurkat were used the expression of SATB1 in leukemia cells, Through cell differentiation induced by all-TRANS Retinoic acid and DMSO, finds the SATB1 gene expression is reduced, suggest that the SATB1 gene may be involved in cell differentiation. By gene silencing, reduce the SATB1 gene expression, sensitivity to chemotherapy drugs is increased, suggest that the SATB1 gene may be associated with drug resistance.

Methods:

1. Western Blot Assay for detection of SATB1 gene expression in cell lines.

2. DMSO and ATRA to HL-60, Jurkat cells for different times, SATB1 gene expression in the cells are detected by Western Blot.

3. Building SATB1-shRNA, transfected Jurkat cells and to HL-60.

4. Testing inhibition rate of chemotherapy drugs on HL-60, HL-60-SATB1-ctr, HL-60-SATB1-sh, Jurkat, Jurkat-SATB1-ctr, Jurkat-SATB1-sh cell, suggesting its relationship with the drug resistance.

Results:

1. SATB1 gene was expression in all the plant cells.

2. The expression of SATB1 in U937, Raji, Hut-102, Akata four types of lymphoma cell line is low.

3. It was highly expression in HL-60, Jurkat cells, but low in K562.

4. DMSO and ATRA treat HL-60, Jurkat cells for 48 and 96 hours, HL-60 cells become larger, rounded, Jurkat cells into smaller, is more obvious for 96-hour.

5. For HL-60, Jurkat cells, after DMSO and ATRA treated, the expression of SATB1 was decreased, more obvious for 96-hour.

6. HL-60-SATB1-sh1, Jurkat-SATB1-sh1 cell compared with the control group, the SATB1 protein content decreased significantly, successfully building SATB1-shRNA HL-60 and Jurkat cell line.

7. HL-60-SATB1-sh1, and Jurkat-SATB1-sh1 of daunorubicin (DNR) sensitivity has improved significantly.

Conclusion:

1. The SATB1 gene expression in leukemia and lymphoma cells, and higher expression in leukemia cells.

2. After DMSO and ATRA treatment, the SATB1 expression significantly reduced, suggest that SATB1 may be involved in cell differentiation.

3. Successfully built SATB1-shRNA, and successfully transferred to HL-60 and Jurkat cell lines.

4. HL-60-SATB1-sh1, and Jurkat-SATB1-sh1 of daunorubicin (DNR) sensitivity increased significantly, prompting SATB1 may be related to drug-resistance.

Disclosures

No relevant conflicts of interest to declare.
Decitabine Act As Demethylation Modulators in Acute Myeloid Leukemia for Reversal of Drug Resistance
Authors: Wang Zhixiang , Jiang Xuejie , Huang Kaikai , Yin Changxin , Zhong Qingxiu , Zhu Qiuhua , Ding Bingjie , Meng Fan Yi
Published: October 2019

Background and objectives The DNA methyltransferase inhibitors decitabine represent archetypal drugs for epigenetic cancer therapy. One alternative approach for the treatment of (Acute Myelocytic Leukemia) AML is the use of hypomethylating agents, including the 5-aza-2-deoxycytidine (decitabine; DAC) and other epigenetic regulators. However, the exact mechanism of DAC for drug resistance AML remains poorly understood. We conducted this study to explore how DAC regulated the sensitivity to adriamycin and aclacinomycin of AML cell line HL60/ADR and assessed the efficacy and safety of decitabine-based induction treatment patients with refractory acute myeloid leukemia.

Methods In this study, we analyzing cell proliferation of HL60, HL60/ADR and KG1-¦Á cells treated by DAC against control group detected with MTS assay. Then the changes of genes methylation were determined using Methylation specific PCR (MSP), stained with anti-CD11b, before being analyzed for cell markers with flow cytometric analysis. The cell protein levels were checked with Western blotting as described elsewhere. Data were collected from refractory acute myelogenous leukemia (AML) patients treated with homoharringtonine, cytarabine, and aclarubicin after decitabine (D-HAA). Control group consisted of fludarabine, cytarabine with or without Granulocyte Colony-Stimulating Factor between 2012 and 2014.

Results Different dose of DAC had proliferative inhibition effect on HL-60 cells, with time-dose dependent relationship. But the proliferative inhibition of DAC in KG1-¦Á and HL-60/ADR cells was not so effective. After been treated with 1¦ÌM DAC for 72 hours, HL-60/ADR cells were cultured with 0.5µg/ml adriamycin 1 hour. Adriamycin positive cells in DAC group were higher than control group, and the proliferative inhibition effect of adriamycin (P<0.001) and aclacinomycin (P<0.001) on HL60/ADR cells was increased (Figure 1). 1¦ÌM DAC significantly induced expression of CD11b on HL-60/ADR and KG1-¦Á cells, accompanied by cellular changes of differentiation (decreased nuclear¨Ccytoplasmic ratio, nuclear condensation, segmentation or lobulation, cytoplasmic granulation or vacuolization). Followed the increased the sensitivity of adriamycin, DNMT1 and SPRPs gene methylation was down-regulated. And ¦Â-catenin protein in nuclear was depressed after treatment of DAC. We enrolled 75 patients, of whom 71 were included in the intention-to-treat analysis. 65.2% patients in the D-HAA group achieved complete remission versus 29.2% in the FA/FLAG group (P=0.031). 2-year disease-free survival and overall survival were better in D-HAA group than FA/FLAG group (P=0.020, P=0.025; Figure 2).

Conclusion These ﬁndings provide evidence for clinic protocols using DAC to reversed drug resistance in refractory AML cell.

Disclosures

No relevant conflicts of interest to declare.
Fev Expression Is Functionally Required for Fetal HSCs and LSCs Indicating Prenatal Origin of Leukemia
Authors: Hong Dengli , Liu Tianhui , Zhang Fang , Liu Feng , Chen Guo-Qiang
Published: October 2019

Cancers, particularly in the childhood and adolescence, can originate prenatally or postnatally. How to distinguish however and what clinical implication of the distinct origin of cancers remains unknown. Here we have identified and characterized transcription factor fev an indicator of prenatal origin of leukemia in the human context of normal and leukemic hematopoiesis. Fev is expressed in fetal and neonatal and not in adult hematopoietic cells. Functional assays showed fev is essential for self-renewal in fetal HSCs. More interestingly, fev is expressed in leukemic cells of some cell lines and patients, which is associated with more frequent relapse. Fev-knockdown in leukemic cells markedly compromised their capacity of leukemic reconstitution in xenograft recipients. Leukemogenic targeting experiments by forced expression of MLL-AF9 or N-Myc showed fev-regulated self-renewal is retained from fetal HSCs rather than reactivated in adult HSCs. Therefore, fev-expression can distinguish LSCs from normal HSCs and indicate its prenatal origin.

Disclosures

No relevant conflicts of interest to declare.
Remission of a Heavily Pre-Treated Relapsed/Refractory ALK-Positive Anaplastic Large Cell Lymphoma after Treatment with Metronomic Therapy
Authors: Shunzong Yuan , Brad Haverkos , Su Hang , Yu Jianhua
Published: October 2019

The prognosis of ALK-positive, anaplastic large cell lymphoma is usually favorable, but patients that are resistant to primary chemotherapy or who relapse early have always been associated with an aggressive clinical course and unfavorable prognosis. For a recurrent relapsing case of ALK-positive, anaplastic large cell lymphoma heavily pretreated by nearly 50 cycles of chemotherapeutic agents (Fig 1.), we developed a treatment schema of metronomically scheduled low-dose methotrexate, which successfully controlled the relapse more than 12 months (Fig. 2). The metronomic therapy that we identified from this case was an effective and economic option that caused less damage to other organs than other treatments; thus, it should be given consideration for the treatment of similar cases, especially in the setting of where the recurrence seems to be only skin disease or when other effective regimens are unavailable. Our study represents the first report of a well-documented case of ALK+ALCL which was refractory to various lines of therapy but which was subsequently successfully treated with a combination of prednisone and etoposide followed by low-dose MTX maintenance metronomic therapy.

Figure 1 Figure 1.

Figure 2 Figure 2.

Disclosures

No relevant conflicts of interest to declare.
Platelets from Patients with Myocardial Infarction Can Activate T Cells in Vitro
Authors: Solomou Elena E. , Gizas Vasilios , Babali Theodora , Perperis Angelos , Verigou Evgenia , Ntavlouros Periklis , Gogos Charalambos , Alexopoulos Dimitrios
Published: October 2019

Specific aim: Our aim was to examine whether T cells can be activated by the circulating activated platelets from patients with myocardial infarction (with ST elevation-STEMI)

Methods: After written informed consent was obtained, peripheral blood mononuclear cells (PBMCs) were isolated from heparinized venous blood obtained from 20 patients with STEMI (18 men and 2 women) at the time of hospital admission at diagnosis, before receiving any treatment, as well as 5 days and 30 days later. We also analyzed 10 healthy subjects (8 men and 2 women), and 5 patients with unstable angina who served as the disease control group. PBMCs were analyzed by flow cytometry with the following markers and their isotypic controls: CD4, CD25, CD69, and FOXP3. We also isolated platelet rich plasma or plasma alone from the patients and the healthy subjects, and used in mixed cultures with PBMCs.

Results: We first examined T cell activation by measuring CD69 expression on CD4 T cells following incubation with platelets obtained from patients with STEMI. T cells treated with platelets from patients with STEMI showed increased expression of CD69 (as an activation marker) compared with T cells treated with platelets from healthy subjects (p<0.05, Figure 1). There was no T cell activation following incubation with plasma alone from patients or healthy controls.

We then examined the percentages of CD4+CD25+hi (regulatory T cells). There was no statistical difference in Tregs between patients at presentation and controls (healthy subjects and disease control group). Five days later, patients with STEMI displayed increased levels of Tregs compared with the 2 control groups; one month later, Treg numbers returned to the initial presentation levels (p<0.05, Figure 2)

Conclusion: To our knowledge we describe for the first time that platelets from patients with STEMI can activate T cells in vitro. In patients with STEMI, an increase in Tregs possibly in an effort to suppress immune system activation secondary to platelet activation, appears shortly after the infarct and normalizes a month later.

Figure 1. T cell activation after treatment with platelets from patients with STEMI compared with the platelets from healthy controls or plasma alone Figure 1. T cell activation after treatment with platelets from patients with STEMI compared with the platelets from healthy controls or plasma alone Figure 2. Increased T regs in patients with STEMI , 5 days after admission (STEMI EXIT) ( UA;unstable angina, STEMI 1mfup; STEMI after one month) Figure 2. Increased T regs in patients with STEMI , 5 days after admission (STEMI EXIT) ( UA;unstable angina, STEMI 1mfup; STEMI after one month)

Disclosures

No relevant conflicts of interest to declare.
Lower Dosages of Intravenous Immunoglobulin (IVIg) in Treating Immune Thrombocytopenia with Long-Term Follow-up of Three Years
Authors: Zhou Zeping , Qiao Zhuoqing , Li Huiyuan , Zhang Xian , Xue Feng , Yang Renchi
Published: October 2019

This study compared the effects of different dosages of intravenous immunoglobulin (IVIg) against immune thrombocytopenia. A total of 167 patients, 91 adults and 76 children, with ITP, followed-up for 3 years in the case-control study, were each divided into three subgroups according to the dosages of IVIg administered: group A (0.2g/kg/day), group B (0.3g/kg/day), group C (0.4g/kg/day). The therapeutic response in 91 adult patients did not differ significantly among the three groups of IVIg dosages (P=0.459). The response rate of IVIg treatment in the three adult groups was 97.1% for group A, and 97.2% for group B, 100% for group C. The mean time for raising platelets to 30 ×109/L in group A was 2.5 days, group B 3.2 days, group C 2.9 days (P=0.324). The median IVIg consumption in group A was 0.83 g/kg, group B 1.22 g/kg, and group C 1.64 g/kg (P<0.01). Similar results were shown in the children groups. The follow-up results showed no significant difference of clinical outcome between groups A, B and C. In conclusion, low-dose IVIg treatment is shown to be as effective as high-dose regimen without increasing the risk of developing the patients into chronic ITP conditions, suggesting that ITP patients could be treated more cost-effectively by lower than conventional dosage of IVIg regimen.

Disclosures

No relevant conflicts of interest to declare.
ONC201 Exhibits Mutation-Independent Efficacy with Superior Potency in Non-Hodgkin Lymphoma and Multiple Myeloma
Authors: Allen Joshua , Tarapore Rohinton , Garnett Mathew J , Benes Cyril
Published: October 2019

ONC201 is a first-in-class small molecule inducer of the integrated stress response that is currently in phase II clinical trials in select advanced cancers with promising early clinical results. The efficacy of this novel agent has been demonstrated in numerous preclinical advance cancer models in multiple indications with an exceptional safety profile that has translated well to the clinic. To determine the preclinical sensitivity profile of ONC201 in cancer, we performed an in vitro efficacy screen across >1,000 human cancer cell lines that represent a diverse array of tumor types and genetic aberrations. Sensitivity profiling was assessed by cell viability assays using dose responses curves at concentrations up to 20uM and at 72 hours post-treatment. Ranking the sensitivity dataset by tumor type, non-Hodgkin's lymphomas and multiple myeloma were the most sensitive tumor type to ONC201.

The mutation-agnostic efficacy that is most pronounced in lymphomas and multiple myeloma is in accordance with the recent findings that ONC201 induces the integrated stress response through a novel target to trigger is downstream late apoptotic effects. B-cell malignancies are particularly susceptible to induction of apoptosis via the integrated stress response, as they have relatively high basal activation of this pathway due to ER stress conferred by immunoglobulin production. Confirmatory studies revealed that multiple myeloma cell lines indeed possess pronounced sensitivity with nanomolar GI50s, unlike most other tumor types, that is particularly encouraging given the systemic concentrations observed in the first-in-man study. Together, these studies suggest specific advanced cancer indications, such as non-Hodgkin's lymphoma and multiple myeloma, as promising lead indications for this novel agent that are being evaluated in phase II clinical trials.

Disclosures

Allen: Oncoceutics, Inc: Employment, Equity Ownership. Tarapore:Oncoceutics, Inc: Employment, Equity Ownership.
Phase I Study of Single Agent Ibrutinib in Recurrent/Refractory Primary and Secondary CNS Lymphoma
Authors: Christian Grommes , Thomas Kaley , Abdel-Wahab Omar , Omuro Antonio M. , Ingo Mellinghoff , DeAngelis Lisa
Published: October 2019

BACKGROUND: Primary CNS Lymphoma (PCNSL) is an aggressive primary brain tumor. Standard treatment of PCNSL may include radiation, methotrexate-based therapy, and anti-CD20 antibody therapy (rituximab) and is associated with substantial morbidity and treatment recurrence. Outcome and treatment options for patients with recurrent/refractory disease are poor. There is only limited use of targeted agents in this patient population. Ibrutinib has shown promising clinical response in some B-cell malignancies. This phase I trial investigates the maximal tolerated dose of ibrutinib in patients with recurrent/refractory PCNSL and secondary CNS lymphoma (SCNSL).

METHODS: Eligible patients had a recurrent or refractory PCNSL or SCNSL, age≥18, KPS≥50, normal end-organ function, and unrestricted number and type of prior therapies. In patients with SCNSL disease, systemic disease needed to be absent or not requiring any active treatment.

RESULTS: Three patients have been enrolled at dose level 1 (560 mg daily) and one patient at dose level 2 (840mg daily) of whom three were women with a median age of 70 years (range 21-80). Three had recurrent PCNSL and 1 recurrent SCNSL. Two patients presented with parenchymal and two with leptomeningeal relapse. Treatment was generally well tolerated. There was one drug-related grade 4 toxicity (neutropenia) that resolved after the drug was held for 4 days. No drug related grade 3 toxicities have been observed to date. Most common grade 2 toxicities were decreased neutrophil count and hyperglycemia. All patients continue on study at a median follow up of 87 days. Three out of four patient were evaluated for response so far. There were two responses: one complete (in the CSF) and one partial, both in recurrent/refractory PCNSL as well as one stable disease in the patient with recurrent SCNSL. The patient with partial response had failed multiple prior treatment regimens including methotrexate-based chemotherapy, radiation, and rituximab/temozolomide. Serum and CSF pharmacokinetic analysis is initiated. Dose level 2 (840mg) is accruing.

CONCLUSION: Patients with CNS lymphoma tolerate Ibrutinib at 560 and 840mg well. Dose escalation will continue. Targeted agents might be an alternative therapeutic approach to be investigated for refractory/recurrent CNS lymphoma patients.

Disclosures

No relevant conflicts of interest to declare.
Evaluation of Modified Japanese Ministry Health and Labor Welfare Diagnostic Criteria for DIC to Detect Early Phase of DIC
Authors: Wada Hideo , Takeshi Matsumoto , Ohishi Kohshi , Katayama Naoyuki
Published: October 2019

Evaluation of the modified Japanese Ministry Health, Labour and Welfare (JMHLW) diagnostic criteria for DIC to detect the early phase of DIC

Introduction: Disseminated intravascular coagulation (DIC) is a syndrome characterized by the systemic activation of blood coagulation, which generates intravascular thrombin and fibrin, resulting in the thrombosis of small-to medium-sized vessels and ultimately organ dysfunction and severe bleeding. Three different diagnostic criteria incorporating similar global coagulation tests have been established by the International Society of Thrombosis and Haemostasis (ISTH), Japanese Ministry Health, Labour and Welfare (JMHLW) and Japanese Association of Acute Medicine (JAAM). The JMHLW score is well correlated with the severity of DIC and can be used to predict the outcome of the disease, however, it has a relatively low sensitivity for the early phase of DIC (pre-DIC).

Material and Methods: The Modified JMHLW criteria are shown in the Table 1. From 1996 to 2014, 282 patients with hematopoietic disorder type, 220 patients with infection type and 249 patients with other type were retrospectively diagnosed with DIC according to the JMHW criteria. There were 184 patients with pre-DIC and 256 patients without DIC.

Table 1. Modified Japanese Ministry Health, Labor and Welfare criteria for DIC Hematopoietic disorder type Othertype Infectiontype Platelet count(X107/ml) none 8 < < 12: 1 point, 5 < < 8: 2 points,(> 30% decrease: plus 1 point)< 5: 3 points FDP (ƒÊg/ml) 10 < < 20: 1 point, 20 < < 40; 2 points, 40< : 3 points Fibrinogen (g/L) 1 < < 1.5 : 1 point, < 1: 2 points none PT ratio 1.25 < < 1.67: 1 point, 1.67 < : 2 points AT (%) < 70 : 1 point TAT or SF 2X (upper limit of normal range): 1 point DIC 4 points = or < 6 points = or < 5 points = or < AT: antithrombin, TAT: thrombin AT complex, SF: soluble fibrin

Results: The agreement rate between the JMHLW and modified JMHLW criteria was 95.9%. The diagnosis rate of pre-DIC was 0% using the JMHLW criteira, 8.3% using the ISTH diagnostic criteria, 63.0% using the JAA criteria and 38.6% using the modified JMHLW criteira for DIC. The mortality of DIC was 42.5% using the JMHLW criteria, 45.5% using the ISTH criteria, 41.3% using the JAA criteria and 42.6% using the modified JMHLW criteria.

Discussion and conclusion: The agreement rate between the JMHLW and modified JMHLW criteria is high. As the modified JMHLW criteria adds the score for antithrombin (AT) activity, thrombin-AT complex (TAT) or soluble fibrin (SF), these criteria increase the sensitivity for pre-DIC.

Disclosures

No relevant conflicts of interest to declare.
Incidence of Pulmonary Embolism with Concomitant Deep Vein Thrombosis at an Urban Community Teaching Hospital
Authors: Goldberg Uri , Nasiri Wazhma , Alibakhiet Mohamed , Jean Henry , Mukherji Rajat , Kalavar Madhumati
Published: October 2019

Introduction

Though venous thromboembolism (VTE) is thought to be under-diagnosed, roughly 900,000 people are estimated to be affected by VTE each year in the U.S. According to CDC figures, VTE accounts for roughly 60,000 to 100,000 annual deaths. Recent research has demonstrated that the risk of mortality in patients with pulmonary embolism (PE) who were found to have concomitant deep vein thrombosis (DVT) was higher in comparison to patients with PE who did not have concomitant DVT (Beccatini et al. Chest 2015). We have sought to evaluate the risk of concomitant DVT among patients diagnosed with PE in our hospital population.

Methods

This is an ongoing retrospective cohort study examining the incidence of PE with concomitant DVT among patients admitted to an urban community teaching hospital between January 2011 and March 2015. Radiological findings for patients who underwent computed tomography angiography (CTA) and venous duplex ultrasound were reviewed. Patients found to have PE were sub-divided into two groups: those with concomitant DVT and those without concomitant DVT. Mortality risk, correlation with inferior vena cava (IVC) filter placement, and association with other comorbidities continue to be evaluated.

Results

Results of 1,777 CTAs were reviewed of which 160 demonstrated a positive finding of PE: 103 women (64.4%) and 57 men (35.6%). Patient age ranged from 19 to 97 years old. The mean and median ages for all PE-positive patients were 61.9 and 63,respectively. Several patients demonstrated multiple emboli; to wit, 273 distinct PEs were noted among the 160 individual patients. Of the 160 patients with PE, 81 were found to have concomitant DVT (50.6%).

Discussion

The combination of high mortality and rapid-though often overlooked-onset make VTE a uniquely vexing condition. Though the nature of the relationship between DVT and the increased mortality risk among patients with PE is unclear, the correlation has been convincingly demonstrated by prior research and may signal the need for more aggressive management for patients with concomitant thromboemboli. Our findings, consistent with the literature, suggest that the prevalence of concomitant PE and DVT may be substantial in the hospitalized population.

Disclosures

No relevant conflicts of interest to declare.
Radiosensitivity of Fancd2-/- mouse Bone Marrow Stromal Cells Is Altered By Abrogation of TGF-β Signaling
Authors: Chen Katherine , Franicola Darcy , Shields Donna , Epperly Michael W. , Zhang Xichen , Greenberger Joel S.
Published: October 2019

Both marrow-transplanted and non-transplanted Fanconi Anemia (FA) patients are often radiosensitive. Due to an increased risk of developing secondary malignancies, these patients require dose and volume modification during radiotherapy. To determine whether abrogation of TGF-β signaling alters the radiation sensitivity of Fancd2-/- mice, cell lines derived from double knockout (DKO) (SMAD3-/- Fancd2-/-) mice were compared with those from Fancd2-/-, SMAD3-/-, and wild-type mice for ionizing irradiation sensitivity.

Bone marrow stromal cell lines were derived from long-term bone marrow cultures of DKO, Fancd2-/-, SMAD3-/-, and wild-type SMAD3+/+ (129/Sv) X Fancd2+/+ (B6) F1 mice. Radiation sensitivity was determined using clonogenic irradiation survival curves. There was no significant difference in radiosensitivity comparing DKO cells (Do = 1.95 ± 0.06 Gy, ň = 4.3 ± 0.7) to the wild type SMAD3+/+ (129/Sv) X Fancd2+/+ (B6) F1 cell line (Do = 2.00 ± 0.11 Gy, and ň = 5.1 ± 0.7, p = 0.7003 and 0.4820, respectively). The Fancd2-/- cell line was more radiosensitive with a Do of 1.37 ± 0.09 Gy compared to 1.95 ± 0.07 and 2.00 ± 0.11 for DKO and wild type cells (p = 0.0063 and 0.0360, respectively. In contrast, the SMAD3-/- cell line was more radioresistant with an increased shoulder on the irradiation survival curve (ň = 12.1 ± 2.9) compared to the DKO or wild type SMAD3+/+ (129/Sv) X Fancd2+/+ (B6) F1 cell lines (ň = 4.335 ± 0.7 or 5.1 ± 0.7, p = 0.00277 or 0.0426, respectively). This confirms and extends results with SMAD3-/- mouse derived cell lines on another background strain (C57BL/6J) (Epperly, et al., Radiation Research, 165:671-677, 2006). TGF-β signaling was abrogated in both DKO and SMAD3-/- mouse cell lines (measured by TGF-β inhibition of fresh marrow CFU-GEMM in vitro), confirming the phenotype of altered TGF-β signaling.

Therefore, radiosensitivity associated with the Fancd2-/- genotype was abrogated by interruption of the TGF-β signaling pathway in the same cells.

Supported by research grant NIAID/NIH, U19A168021.

Disclosures

No relevant conflicts of interest to declare.
Association of the RUNX1/AML1 Gene Mutation and Familial MDS/AML: A Case Report and Review of Literature
Authors: Chen Wancheng , Li Hongbo
Published: October 2019

Myelodysplastic syndrome (MDS) and Acute Myeloid Leukemia (AML) are mostly sporadic hematopoetic malignancies, which generally thought inheritable, though specific gene abnormalities may play a role in the progression of myelodysplasia. Recently, many reports have revealed that mutations in the RUNX1/AML1 gene are involved in defective hematopoiesis. Moreover, patients with such mutations have a significant poorer prognosis than cases without RUNX1 mutations. Since familial occurrence of MDS/AML is rare, there are few reports about the relationship between RUNX1 mutations and familial MDS/AML. In the study of a familial MDS/AML, we found a61-year-old male with MDS who developed into AML later. He had a history of leukocytopenia and thrombocytopenia for decades before developing into pancytopenia. We identified a RUNX1mutationin this patient through sequencing. Furthermore, the same mutation was present in his son (II-1) who suffered from MDS as well as his granddaughter. Intriguingly, II-1 was 10years old when diagnosed with MDS, nearly 50 years earlier than his father. Myelodysplastic features in bone marrow sections of these two patients were not obvious in the early stage. They mainly showed active erythrocyte proliferation, together with an increased mean corpuscular volume, which can indicate inflection points in MDS progression. To date, patients with familial MDS had no antecedent hematological abnormalities in the early stage. In this study, we found a new inheritable mutation of the RUNX1 gene in the familial MDS which is frequently deregulated in hematological malignancies. As RUNX1 was present in the early stage of familial MDS, this genetic mutation may serve as an early diagnostic marker and contribute to the treatment of MDS/AML.

Disclosures

No relevant conflicts of interest to declare.