bluuebunny/crossref_metadata_2025 huggingface dataset:
The Impact of Antithymocyte Globulin (ATG) Dose in Allogeneic Hematopoietic Stem Cell Transplantation with HLA Mismatched Donors
Authors: Choi Yunsuk , Lee Ho Sup , Lee Je-Hwan , Moon Joon Ho , Shin Ho-Jin , Lee Won Sik
Published: October 2019

Background: Antithymocyte globulin (ATG) is usually included in conditioning regimens in hematopoietic stem cell transplantation (HSCT) from HLA mismatched donors to reduce the incidence of graft versus host disease (GVHD) and rejection. In this study, we intended to assess the clinical impact of ATG dose in HSCT from HLA mismatched donors.

Patients and Methods: We retrospectively analyzed 268 consecutive patients with acute myeloid leukemia (n=159), acute lymphoblastic leukemia (n=49) or myelodysplastic syndrome (n=60) receiving busulfan based conditioning and HSCT with HLA mismatched sibling, unrelated, or haplo-identical family donors in five centers of Korea between 2005 and 2015.

Results: The median age was 44.5 (range, 15-75) years. Fourty-two patients were treated with myeloablative conditioning and 226 patients with reduced intensity conditioning regimen. ATG at a total dose of 0 (n=31), 2.5-7.5 mg/kg (n = 16), 9 mg/kg (n = 164) or 12mg/kg (n=57) was given to patients according to protocol. 140 patients received HSCT in complete remission and 73 patients in relapse or refractory status. The median follow up duration for surviving patients was 33.8 (range, 2.0-100.6) months. The 3-year overall survival (OS) and event free survival (EFS) rate for all patients was 44.7% and 39.2% There was no significant difference of OS according to ATG dose (P=0.153), but ATG dose of 2.5-7.5mg/kg was significantly associated with better EFS than 0, 9 mg/kg, or 12mg/kg of ATG groups (79.1% vs. 45.4%, 38.9%, and 31.7%, respectively, P=0.038) (Table 1). In addition, ATG dose significantly affected on relapse (P=0.046). The relapse incidence in patients receiving 2.5-7.5 mg/kg of ATG was lower compared to those given 0, 9, or 12 mg/kg of ATG (6.7% for 2.5-7.5mg/kg of ATG group vs. 29.5%, 34.0%, and 46.9% for 0, 9, and 12 m/kg of ATG groups, respectively). The incidences of acute GVHD grades II-IV were 32.6%, 25.0%, 29.4% and 24.6% in patients receiving 0. 2.5-7.5, 9, and 12 mg/kg of ATG, respectively. The incidence of extensive chronic GVHD tended to be lower according to the increase of ATG dose (36.8%, 34.5%, 27.4%, and 21.1% for 0. 2.5-7.5, 9, and 12 mg/kg of ATG dose, respectively).In multivariate analysis, ATG dose of 9 mg/kg (P=0.023) and 12 mg/kg (P=0.036) was associated with shorter EFS compared to 2.5-7.5 mg/kg of ATG.

Conclusion: 9mg/kg of ATG dose or more in HLA mismatched busulfan based HSCT with HLA mismatch donors increased the risk for relapse and reduced the EFS.

Disclosures

No relevant conflicts of interest to declare.
Efficacy of CD101, a Novel Echinocandin, in Mouse Models of Aspergillosis and Azole-Resistant Disseminated Candidiasis
Authors: Ong Voon , Bartizal Ken , Miesel Lynn
Published: October 2019

Background: CD101 is a novel echinocandin with long-acting pharmacokinetics and exceptional stability in development for prevention and treatment of serious fungal infections. The in vivo efficacy of CD101 was evaluated using neutropenic mouse models of azole-resistant candidiasis and aspergillosis.

Methods: An azole-resistant strain of Candida albicans (R357; resistant to fluconazole [Flu], voriconazole, and posaconazole but susceptible to amphotericin B [AmB] and echinocandins) isolated from human blood was used for the mouse candidiasis model. A test strain of Aspergillus fumigatus (ATCC 13073) was used for the mouse aspergillosis model. Mice were rendered neutropenic by cyclophosphamide and then infected by injections of C. albicans (105 CFU/mouse) or A. fumigatus (104 CFU/mouse) into the tail vein. Test articles were administered starting 2 hours after infection. In the mouse candidiasis model, groups of 5 mice each received one dose of AmB (3 mg/kg IV), Flu (20 mg/kg orally), or CD101 (3, 10 or 30 mg/kg by intraperitoneal administration [IP]). After 72 hours post-infection, mice were euthanized and C. albicans counts in kidney tissue (CFU/g) were measured. In the mouse aspergillosis model, groups of 10 mice each received one dose ofAmB (2 mg/kg IP) or CD101 (2 mg/kg IV and IP). Survival was monitored daily for 10 days. Differences between vehicle and test article groups were assessed for significance by one-way ANOVA followed byDunnett's test and Fisher's Exact test in the candidiasis and aspergillosis models, respectively.

Results: One dose of CD101 3 mg/kg produced a >99.9% (or > 3-log; P<0.001) reduction in C. albicans CFU compared with vehicle through at least 72hours post-dose following a single IP dose.AmB showed similar, albeit less robust, efficacy (>99% or >2-log reduction in CFU; P<0.05), whereas fluconazole was less efficacious (83.9% or <2-log reduction in CFU). In the aspergillosis model, CD101 administered 2 mg/kg IV or IP showed similar efficacy to that ofAmB 2 mg/kg IP, both with significantly longer survival than vehicle (P<0.05; Figure).

Conclusions: A single dose of CD101 3 mg/kg produced significant reduction in C. albicans burden compared with vehicle (P<0.001) in the neutropenic mouse model of azole-resistant candidiasis, demonstrating efficacy comparable, if not better, to that of AmB at the same dose. One dose of CD101 also demonstrated efficacy in the mouse model of aspergillosis. These data support the continued development of CD101 for treatment of serious infections caused by Candida, including azole-resistant strains, and Aspergillus spp.

Figure Percent Survival in a Neutropenic Mouse Model of Aspergillosis Figure. Percent Survival in a Neutropenic Mouse Model of Aspergillosis

Disclosures

Ong: Cidara Therapeutics, Inc.: Employment. Bartizal:Cidara Therapeutics, Inc.: Employment, Other: Stockholder. Miesel:Eurofins Panlabs: Employment; Cidara Therapeutics, Inc: Research Funding.
Invasive Mold Infections in Acute Leukemia Patients Undergoing Allogeneic Hematopoietic Stem Cell Transplantation
Authors: Chien Sheng-Hsuan , Liu Yao-Chung , Fan Nai-Wen , Liu Chia-Jen , Chiou Tzeon-Jye , Gau Jyh-Pyng , Liu Jin-Hwang
Published: October 2019

Introduction:

Patients with acute leukemia exposed higher risk for developing invasive fungal infections and the invasive fungal infection is also an important cause of morbidity and mortality during allogeneic hematopoietic stem cell transplantation (allo-HSCT). In addition to candida species, the invasive mold infection (IMI) is most common in invasive fungal infections and the incidence rate is increasing in recent years. Although recent diagnostic approach and treatments for IMI are advanced, the prognosis remains poor. It is essential to understand the risk factors for developing IMI among acute leukemia patients undergoingallo-HSCT. Here, we conducted a retrospective study to demonstrate demographics, microbiology, and risk factors for the development of IMI among 245 adult acute leukemia patients undergoingallo-HSCT at our institution during a 10-year period.

Method

We reviewed 245 adult acute leukemia patients undergoingallo-HSCT from January 2003 to December 2014. Clinical characteristics including age, sex, underlying disease, type of allogeneic transplant, conditioning regimens, European Group for Blood and Bone marrow Transplantation (EBMT) risk score, and presence of acute graft-versus-host disease (aGVHD) or chronic GVHD (cGVHD) were collected and analyzed. Cox proportional hazard model was adopted to explore the independent risk factors for IMI development. The Kaplan-Meier method was performed to estimate the cumulative incidence and the curve was compared using the log-rank test

Results

Twenty out of 245 patients developed IMI during study period and the median time to onset after transplantation was 391 days (interquartile range, 220-552 days). The cumulative incidence of IMI in this cohort was 1.9 %, 5.1%, and 12.8% at 6 months, 12 months, and 24 months, respectively. Aspergillus species were the most common and presented in 55% of mold infections. The significant risk factors to predict mold infection after transplantation (Table 1) were smoking (hazard ratio [HR] 4.28, 95% confidence interval [CI] 1.40-13.12; P=0.011), EBMT risk score > 2 (HR 4.37, 95% CI 1.35-14.09; P=0.013), and extensivecGVHD(HR 3.10, 95% CI 1.20-8.00; P=0.019). The cumulative incidence of mold infection in smokers was significantly higher than non-smokers (log-rank P < 0.001) and the Kaplan-Meier curve was shown in Figure 1.

Conclusion

We identified three risk factors-smoking, EBMT risk score > 2 and extensivecGVHDto predict IMI among acute leukemia patients undergoingallo-HSCT. Smoking may damage respiratory tract epithelium as well as defense-microorganism mechanism and it would lead to IMI easily duringallo-HSCT. This cohort study suggests that early identification of high-risk patients and to provide better prevention strategies would reduce the incidence and severity of IMI in these patients.

Figure 1 Figure 1.

Disclosures

No relevant conflicts of interest to declare.
Nek2 Stabilization By Usp7 Leads to Activation of NF-Kb in Multiple Myeloma
Authors: Franqui Machin Reinaldo , Zhan Xin , Xu Hongwei , Frech Ivana , Tricot Guido J , Zhan Fenghuang
Published: October 2019

NIMA (Never In Mitosis Gene A)-Related Kinase 2 (Nek2), a centrosomal Serine/Threonine kinase, is a key player in numerous malignancies. Overexpression of Nek2 has been related to many cancers including Multiple Myeloma (MM). In MM, Nek2 is one of the chromosomal instability genes associated with drug resistance and disease relapse. However, very little is known about the mechanisms that lead to these Nek2-driven disparities. Here, we show that the Ubiquitin Specific Peptidase 7 (USP7) stabilizes Nek2 leading to activation of NF-kb pathway.

Using gene expression profile (GEP) data from patients and cell lines we discovered that Nek2 overexpression leads to increases of several targets of the NF-kb pathway. We, thus, hypothesize that Nek2 is activating NF-kb. To address this, we overexpressed Nek2 and tested the classic canonical NF-kb hallmarks proteins by western blotting. Nek2 overexpression led to an increase in phosphorylation of IKK, activator of NF-kb, and to decrease levels of IKb-alpha, a negative regulator of the pathway. Nek2 overexpression also increased nuclear and phosphorylated p65 on residue S536, known as active transcriptional site. To further confirm that Nek2 is activating canonical NF-kb luciferase assay was performed. The luciferase reporter is driven by a p65 promoter and in cells overexpressing Nek2 luciferase levels were increased.

To characterize Nek2 interacting partners a tandem affinity purification/mass spectrometry (TAP/MS) approach was performed. We found that Nek2 binds to Usp7, a deubiquitinase overexpressed in numerous cancers. This led to hypothesize that that Nek2, a known target of the ubiquitin proteasome system, is being stabilized by the Usp7 contributing to its overexpression and the increased activation of the NF-kb pathway. To test our hypothesis, we treated different cancer cell lines with the commercially available Usp7 inhibitor, P5091, or silenced the protein using shRNA. In both case, we found a reduction in Nek2 protein level. Additionally, we overexpressed Usp7 and Nek2 increased confirming that Usp7 stabilizes Nek2. To further show that Usp7 stabilizes Nek2 by de-ubiquitination, we overexpressed Usp7 and analyzed Nek2 ubiquitination after immunoprecipitation. When Usp7 was overexpressed no ubiquitination of Nek2 was detected.

Finally, by using GEP data from MM patients, we found that individuals who overexpressed Nek2 along with an active NF-kb signature have worst event free survival as well as overall survival, indicating Nek2 overexpression leading to increased NF-kb signature has clinical significance.

Disclosures

No relevant conflicts of interest to declare.
Ex-Vivo Fucosylation Improves the Anti-Graft-Versus-Host-Disease Effects of Mesenchymal Stem Cells in the NOD/SCID/IL-2R Null Mouse Model
Authors: Galvez Silva Jorge , Mendt Vilchez Mayela , Nobuhiko Imahashi , Blazar Bruce R. , Zweidler-McKay Patrick , Rezvani Katayoun , Shpall Elizabeth J.
Published: October 2019

BACKGROUND: Graft versus host disease (GVHD) remains the major cause of morbidity and mortality following allogeneic hematopoietic stem cell transplantation (HSCT). Due to their immunomodulatory properties, mesenchymal stem cells (MSCs) have produced durable responses in HSCT patients with steroid resistant GVHD, but their positive effects are not seen in all patients. Sackstein et al have shown that MSCs lack an E-selectin ligand on their surface, which impairs their ability to home to the bone marrow and potentially to sites of inflammation associated with GVHD. Our in vitro results confirmed that fucosylation markedly enhanced the E-selectin ligand expression on the MSC cell surface and did not adversely affected MSCs immunomodulatory properties. Hencewe postulated that ex-vivo fucosylation of MSCs would enhance their homing to sites of inflammantion and thus their clinical efficacy in our NOD/ SCID/IL-2R null (NSG) mouse model of GVHD.

METHODS: On day -1 NSG mice were sub-lethally irradiated (300cGy) and infused with 2x106 fucosylated MSCs. Briefly, healthy bone marrow donor derived MSCs were treated for 30 minutes at room temperature with FT-VI (33ng/ml) and GDP-fucose (1mM) (Targazyme, Carlsbad, CA), and then washed prior to tail vein injection. Control mice received 2x106 unfucosylated MSCs or no MSCs. On day 0 all mice received 1x107 normal human donor peripheral blood mononuclear cells (PBMNCs), representing a MSCs:PBMNCs ratio of 1:5. Mice were then followed for weight loss, clinical GVHD score and survival.

RESULTS: We found that a single dose of FT-VI treated MSCs reduced the acute severity of GVHD compared to unfucosylated MSCs and no MSCs (PBMNCs alone) in this xenogeneic mouse model. This was shown by prevention of weight loss (Fig. 1a), and lower GVHD scores (Fig 1b) in the FT-VI treated MSCs mice. Furthermore, when short-term survival was compared among the subgroups, unfucosylated MSCs did not significantly improve short-term survival over PBMNCs group (P=0.3) (Fig. 2a). However, FT-VI treated MSCs mice showed a significantly longer short-term survival compared to PBMNCs group (P=0.002) (Fig. 2b) and untreated MSCs mice (P=0.02) (Fig. 2c).

CONCLUSION. Our data suggest that fucosylated MSCs exert a better anti-GVHD effect at a similar dose than unfucosylated MSCs. Additional data is being generated to confirm these preliminary results and optimize the dose and schedule of fucosylated MSCs.

Disclosures

No relevant conflicts of interest to declare.
Intra-Tumor Injection of CAR-Engineered NK Cells Induces Tumor Regression and Protection Against Tumor Re-Challenge
Authors: Boissel Laurent , Klingemann Hans , Khan Junaid , Soon-Shiong Patrick
Published: October 2019

In this era of cancer immunotherapy, the long-term goal of treatment is to eliminate tumor cells anywhere in the body through induction of a specific immune memory response. In addition to spontaneous cytotoxicity, ADCC, and cytokine release, NK cells contribute to an adoptive immune response through crosstalk with dendritic cells and T cells. To further characterize the NK cell-induced adoptive response, aNK cells (formerly NK-92) were transduced with a retrovirus construct coding for a second-generation anti-murine CD19-CAR to create targeted activated NK-92 cells (mCD19.taNK). mCD19.taNK cells were injected intra-tumorally (two injections of 5x106 cells, three days apart) into a murine model of syngeneic subcutaneous lymphoma, created by injecting 1x106 A20 cells into BALB/c mice. Tumor size was monitored over time, and mice showing complete tumor regression were re-challenged with another subcutaneous contralateral injection of A20 cells.

mCD19.taNK cells effectively killed murine cancer cells in vitro (>60% killing at E:T ratio of 5:1). In vivo, intra-tumor injection of mCD19.taNK induced significant tumor regression versus saline (tumor volumes of 342 mm3 and 936 mm3, respectively at day 16, p<0.05) and significantly improved survival, with 75% of the mice showing complete tumor regression at day 32 (p<0.05). In contrast, injection of parental aNK cells did not significantly affect tumor size in mice (815 mm3at day 16). Upon re-challenge with A20 lymphoma cells, >80% mice remained free of tumor after 14 days.

In conclusion, the human aNK cell line expressing an anti-murine CD19-CAR (mCD19.taNK) can effectively kill CD19-positive murine cancer cells. Moreover, intra-tumor injections of mCD19.taNK into a fully immunocompetent mouse model can induce tumor clearance and protection from tumor re-challenge, suggesting induction of a memory ("vaccine") effect after the injection of mCD19.taNK.

Disclosures

Boissel: NantKwest, Inc.: Employment. Klingemann:NantKwest, Inc.: Employment, Equity Ownership, Patents & Royalties. Khan:NantKwest, Inc.: Employment. Soon-Shiong:NantKwest, Inc.: Employment, Equity Ownership, Membership on an entity's Board of Directors or advisory committees.
Economic, Transfusion, and Efficacy Outcomes with the Addition of IV Iron Sucrose to Oral Iron Therapy in Pregnancy Associated Iron Deficiency Anemia
Authors: Gaspe Mudiyanselage Nilupa , Elrafei Tarek , Lewis Beth , King Mary , Strakhan Marianna , Alexis Karenza , Gralla Richard
Published: October 2019

Background: Prior studies have indicated that transfusion is unusual (2%) in pregnant women with iron deficiency anemia. Nonetheless, compliance with oral iron replacement can be an issue and physicians may wish to use IV iron therapy in markedly anemic pregnant women.

Objectives: to evaluate the effectiveness of adding intravenous iron sucrose concentrate (ISC) to pregnant patients already taking oral iron in terms of effect on hemoglobin, effect on ferritin levels, rates of transfusion, and cost.

Methods: We analyzed all referrals from Obstetrics to Hematology clinic and Obstetrics consultation (Internal medicine) clinic from January 2014 to June 2016. Of the 176 pregnant patients, 98 were referred for anemia, including 81 patients with Hgb < 12 g/dl and ferritin < 20 ug/L. All had previously been given oral ferrous sulfate prescriptions. Patients with hemoglobinopathy were excluded. All 81 patients were advised to continue on the oral iron, and 40 were given IV iron sucrose (ISC group).

Results: The average cumulative dose of iron sucrose was 700 mg, a mean of 5.575 doses (initiated in the third trimester in 38 of 40 patients). The lowest antepartum Hgb was 8.18 g/dl in the ISC group and 9.58 in the oral only group; there was an average Hgb increase of 2.17 vs 1.76 g/dl respectively (p=.107 NS and the 0.41 g/dl difference was considered to be of no clinical consequence). 89% in the ISC group vs 30% in the oral achieved a ferritin >20 (p=0.000015). No adverse events in the IV iron group were reported. There was 1 transfusion in the oral iron group attributable to iron deficiency (2.4%) vs none in the IV iron group (p = 0.107 NS). Two patients were transfused in the antenatal period before IV iron was started and 1 transfused because of post-partum hemorrhage. The total cost of the IV iron therapy would add an average of $1,500 per patient. Thus, and additional cost of $60,000 in IV iron would be required to prevent 1 transfusion [40:1].

Conclusions: ISC corrects ferritin in more patients than oral iron replacement, but did not significantly increase Hgb levels or have a meaningful impact on the transfusion rate. The additional cost and lack of clinically improved outcomes with IV iron argue against its use and in favor of strategies to ensure compliance with oral iron.

Disclosures

No relevant conflicts of interest to declare.
Endogenous Adamts-13 Regulates Angiogenesis in Cultured Human Endothelial Cells
Authors: Tang Huiyuan , Lee Manfai , Ho Kim Eun , Bishop Daniel Allen , Rodgers George M.
Published: October 2019

ADAMTS-13, a disintegrin and metalloproteinase with a thrombospondin type 1 motif, member 13, is a zinc-containing metalloprotease enzyme that cleaves von Willebrand factor (vWf). Previous publications by our laboratory have shown that ADAMTS-13 may also be involved in angiogenesis; for example; addition of exogenous ADAMTS-13 enhances angiogenesis in cell migration and proliferation assays. For this study, we report the successful transient knockdown of endogenous ADAMTS-13 in human umbilical vein endothelial cell (HUVEC) via siRNA. 100pmols of ADAMTS-13 siRNA inhibited HUVEC ADAMTS-13 expression levels by 60% after 7hr incubation, whereas control siRNA did not reduce endogenous ADAMTS-13. The effects of reduced endogenous ADAMTS-13 on HUVEC function were studied in proliferation and scratch wound assays. Transfection of HUVEC with 100pmols of ADAMTS-13 siRNA resulted in a 47% decrease in proliferation after 19hr incubation. The effects of ADAMTS-13 knockdown on migration of HUVEC across a scratch wound were also studied. 9hr after transfection with control siRNA, cells initiated migration across the scratch wound. This migration did not occur in ADAMTS-13 knockdown cells. The activity of the AKT pathway, one of the angiogenesis downstream signaling pathways was down regulated by ADAMTS-13 siRNA while ERK, a component of the MAP kinase pathway was not affected upon knockdown of ADAMTS-13. These data indicate that one role of endogenous ADAMTS-13 in HUVEC is regulation of angiogenesis, mediated through the AKT signaling pathway. Overall our data suggest an additional model of endogenousADAMTS-13 functionality, beyond that of cleaving von Willebrand factor.

Disclosures

No relevant conflicts of interest to declare.
Prospective, Non-Randomized, Open-Label, Single-Arm, Multi-Center Clinical Trial to Evaluate the Efficacy and Safety of Intravenous Immunoglobulin 10% Formulation in Patients with Immune Thrombocytopenia (ITP)
Authors: Bang Soo-Mee , Mun Yeung-Chul , Shin Ho-Jin , Jung Chul Won , Bae Sung Hwa , Sohn Sang Kyun , Kim Kyoung Ha , Kim Hawk , Yhim Ho-Young , Hong Junshik , Kim Hye Joo , Lee Jae-hoon , Oh Doyeun
Published: October 2019

This study investigated the safety and efficacy of IV-Globulin SN, a 10% intravenous immunoglobulin (IVIg), in patients with severe ITP (platelet count ¡Â20 x 109/L). Among 81 eligible patients, 31 patients were newly diagnosed, 7 patients had persistent ITP, and 43 patients had chronic ITP. Five patients had received splenectomy. Patients received IV-Globulin SN 1 g/kg/day on two consecutive days; infusion rate was initially 1 mg/kg/minute and then doubled every 30 minutes to a maximum of 8 mg/kg/minute.

Fifty-eight patients (72%) attained the primary efficacy endpoint of clinical response (platelet count ¡Ã 50 x 109/L within 7 days). Their median time to response was 2 days and the median duration of response was 10 days (range from 1 to 28 days). Complete response (platelet count ¡Ã 100 x 109/L over 7 days) was obtained in 14 patients (17%). Response rates were not significantly different when compared the patients with newly diagnosed, persistent or chronic ITP; previous treatment with immunosuppressant or not; splenectomized or not. IV-Globulin SN 10% was well tolerated and the frequency of mucocutaneous bleeding was decreased during the study period (figure 1). There was no unexpected adverse event. The mean half-life and Cmax of study drug were 28.9 days and 34.6 g/L in 25 tested patients. There were no unexpected adverse events. In conclusion, IV-Globulin SN was efficacious in adult ITP patients regardless of their disease status as well as safe given that the resolution of bleeding and minimal infusion-related adverse events. (NCT02063789)

Bleeding rate according the anatomic sites during the study period in 81 patients. *Visit 2 means day 1; visit 3, day 2; visit 4, day 4¡¾1; visit 5, day 6¡¾1; visit 6, day 8¡¾1, visit 7, day 11¡¾1; visit 8, day 15¡¾1; visit 9, day 22¡¾3; visit 10, day 29¡¾3) Bleeding rate according the anatomic sites during the study period in 81 patients. / *Visit 2 means day 1; visit 3, day 2; visit 4, day 4¡¾1; visit 5, day 6¡¾1; visit 6, day 8¡¾1, visit 7, day 11¡¾1; visit 8, day 15¡¾1; visit 9, day 22¡¾3; visit 10, day 29¡¾3)

Disclosures

Kim: Green Cross Corp.: Employment. Lee:Green Cross Corp.: Employment.
Assessment of Intracranial Hemorrhage Demographics in Children with Hemophilia: A Multicentre Study in China
Authors: Haque Qareen , Feng Xiaoqin , Li Chunfu
Published: October 2019

Objective: Intracranial hemorrhage (ICH) is a severe complication in hemophilia patients with high mortality and disabling sequelae. This study is to provide key insights on the demographics of hemophilic patients with ICH in China.

Method: The database and medical notes ofHemophilia patients who were diagnosed between 1995 and 2014 are reviewed with a goal to find the effective factors which relate to ICH such as age of onset, type, severity, cause, and ICH position, presence of inhibitor and neurological sequelae. A total of 145 hemophilic boys who are diagnosed with ICH and come from multiple hemophilia centers in China, are included.

Result: The median age of ICH is 18 months. 123/145 (85%) are Hemophilia A and 22/145 (15%) are Hemophilia B. Intracranial hemorrhages most commonly occur in patients with severe disease 79/145 (54%), while 58/145 (40%) and 8/145 (5%) in moderate and mild disease respectively. There is obvious trauma history in 53/145 (37%) children. Neurological disabilities are seen only in 34/145 (23%). No deaths from ICH have been reported in our study. Data about the position of ICH is available only in 30% of patients; including involvement of multiple areas is seen in 50%. ICH in epidural space is 14%, brain parenchyma is 14%, subdural is 11% and subarachnoid is 5%.Out of 30 patients inhibitor is found only in 9 cases while in other 115 patients inhibitor data is unavailable. After the onset of ICH only 53/145 (37%) children are put on regular prophylaxis to prevent further episode of ICH.

Conclusion: Risk of ICH is high in patients with severe hemophilia. In most cases of ICH, there is no trauma history and relatively less neurological consequences. About half of children with ICH multiple areas of brain are involved. We need more data to conclude the other risk factors in the incidence of ICH.

Disclosures

No relevant conflicts of interest to declare.
Evaluation of Direct Oral Anticoagulants in the Treatment of Venous Thromboembolism for Inherited Thrombophilia Disorders
Authors: McBride Ali , Diri Reem , Krishnadasan Ravitharan , Chalasani Pavani , Abraham Ivo , Campen Chris
Published: October 2019

Background

Venous thromboembolism can be classified according to the presence of either environmental or genetic risk factors. Risk factors for thrombosis can include activated protein C resistance, and heritable including deficiencies of antithrombin, protein C or protein S. Factor V Leiden deficiency and prothrombin gene mutations are some of the more common thrombophilias, with a slight increased risk for venous thromboembolism (VTE). Current guidelines suggest the use of low-molecular weight heparins for secondary prophylaxis in patients with VTE. However, there is a lack of data on the use of Direct Oral Anticoagulant (DOACs) in patients with inherited thrombophilia. We evaluated our use of rivaroxaban in patients with thrombophilia disorders treated for secondary DVT prophylaxis.

Method

We performed a retrospective evaluation of patients in our institution with inherited thrombophilia with an active VTE diagnosis who received DOACs for secondary prophylaxis from November 2013 until April 2016. Data collected included patient demographics, inherited thrombophilia mutation, previous history of VTE, prior treatments, and efficacy and safety of anticoagulation with DOACs.

Results

We had 13 patients with inherited thrombophilia mutation and 4 patients diagnosed with concomitant cancer (non-Hodgkin lymphoma, melanoma, and 2 with breast cancer) (Table 1). Out of 13 patients 3 failed warfarin, and one failed fondaparinux prior to switching to a DOAC. Mutation with heterozygous Factor V Leiden deficiency was reported in 7 patients, while mutations with Protein C and/or S deficiency were found in 4 patients. One patient had both Factor V Leiden and Protein C deficiency mutations. The prothrombin gene mutation was identified in one patient. The median of length of therapy was 2 years with 8/13 still on rivaroxaban in April 2016. The shortest treatment duration was 41 days for a patient who failed rivaroxaban with a second clot and was switched to apixaban without subsequent treatment failure. Two patients experienced 4 non-major episodes of gastrointestinal bleeding, nose bleeding and dark stool. One patient developed rash with noted bruising during their rivaroxaban therapy.

Conclusion:

This is the first report on outcomes for secondary DVT prophylaxis with DOACs in patients with underlying thrombophilia mutations. Safety and efficacy of DOACs for secondary VTE prophylaxis yielded favorable results; however, future prospective studies in the setting of thrombophilia are warranted.

Table 1 Summary of baseline characteristics and outcomes. Table 1. Summary of baseline characteristics and outcomes.

Disclosures

McBride: Sanofi: Research Funding.
The Effectiveness and Safety of Direct Oral Anticoagulants (DOACs) Vs. Traditional Anticoagulants in Patients with Cirrhosis
Authors: Hum Justin , Jou Janice , Deloughery Thomas G. , Shatzel Joseph
Published: October 2019

Introduction: The coagulopathy associated with cirrhosis is complex and places patients at risk for both bleeding and thrombosis. Direct oral anticoagulants (DOACs) have been shown to have superior efficacy and safety compared to vitamin K antagonists; however their efficacy and safety in cirrhotic patients is not clear. The aim of this study is to retrospectively compare the effectiveness and bleeding complications of DOACs as compared to traditional anticoagulants in cirrhotic patients.

Methods: This study was a retrospective review of patients treated at a single academic center between 2012-2015 who were prescribed a DOAC (apixaban or rivaroxaban), or a traditional anticoagulant (warfarin or low molecular weight heparin), with an ICD-9 code for the diagnosis of cirrhosis. The primary outcomes of interest are recurrent thrombosis or stroke (efficacy failure), or bleeding events (safety failure). Major bleeds were characterized as fatal bleeding, symptomatic bleeding in critical organ area, or bleeding causing a fall in hemoglobin level >2 or leading to transfusion of 2+ units of packed red blood cells.

Results: During the study period, 27 cirrhotic patients were prescribed a DOAC and 18 were prescribed a traditional anticoagulant (either LMWH or warfarin). Both groups had similar total bleeding events (8 DOAC vs. 10 traditional anticoagulation, p = 0.12). There were significantly less major bleeding episodes in the DOAC group, (1 (4%) vs. 5 (28%), p = 0.03) and less intracranial bleeding (3 (17% ) vs. 0 (0%) p=0.06). Recurrent thrombosis or stroke occurred in 1 (4%) patient in the DOAC group and 1 (6%) patient in the traditional group (p = 1.0).

Conclusions: Anticoagulation with DOACs in cirrhotic patients may be as safe as traditional anticoagulants with respect to bleeding events. Patients with cirrhosis at our center prescribed DOACs had less major bleeding events, while maintaining efficacy at preventing stroke or recurrent thrombosis.

Table Table.

Disclosures

No relevant conflicts of interest to declare.
R4 Rgs Subfamily Proteins Negatively Regulates SDF-1/CXCR4 Signaling in CD34+ Hematopoietic Stem and Progenitor Cells
Authors: Leung Kam Tong , Wong Yorky Tsin Sik , Li Karen , Chan Kathy Yuen Yee , Zhang Xiao-Bing , Leung Ting Fan , Ng Pak Cheung , Li Chi Kong
Published: October 2019

RGS family proteins are known to negatively regulate G-protein-coupled receptor signaling through their GTPase-accelerating activity. In several types of hematopoietic cells (e.g., B lymphocytes and megakaryocytes), responses to stromal cell-derived factor-1 (SDF-1) are subjected to regulation by R4 subfamily RGS proteins. However, their expression patterns and functional roles in hematopoietic stem and progenitor cells (HSC) are poorly characterized. Here, we showed that human CD34+ HSC derived from cord blood (CB, n = 10) expressed 7 out of 10 R4 RGS proteins at mRNA level (RGS1-3, 5, 13, 16 and 18), whereas expressions of RGS4, 8 and 21 were undetectable. Exposure of CB CD34+ cells to SDF-1 significantly increased RGS1, 2, 13 and 16 expressions and decreased RGS3 and 18 expressions (P ≤ 0.0402, n = 5). Expressions of RGS1, 13 and 16 were significantly higher in bone marrow (BM, n = 10) CD34+ cells when compared to mobilized peripheral blood (MPB, n = 5) CD34+ cells (P ≤ 0.0160), while RGS3 and 18 expressions were lower in BM CD34+ cells (P ≤ 0.0471), suggesting a SDF-1- and niche-dependent regulation of RGS expressions. To investigate the potential involvement of RGS proteins in SDF-1-mediated homing-related functions, we introduced RGS overexpression constructs into CB CD34+ cells by lentiviral transduction. With >80% transduction efficiency, we showed that overexpression of RGS1, 13 and 16 but not RGS2 significantly inhibited migration of CD34+ cells to a SDF-1 gradient (P ≤ 0.0391, n = 4-5). Similarly, RGS1, 13 and 16 overexpression suppressed SDF-1-induced Akt phosphorylation (n = 2), but none of them affected SDF-1-mediated actin polymerization (n = 3). In the NOD/SCID mouse xenotransplantation model, preliminary results showed that bone marrow homing was impaired in RGS1- (16.3% reduction), RGS13- (12.7% reduction) or RGS16-overexpressing CD34+ cells (33.7% reduction). Taken together, we provided the first evidence that expressions of R4 RGS proteins are regulated by the SDF-1/CXCR4 axis in human CD34+ HSC. We also presented evidence that specific R4 RGS proteins (RGS1, 13 and 16) negatively regulate in vitro SDF-1-mediated responses and in vivo homing of CD34+ cells, suggesting that RGS proteins may serve as a feedback mechanism to regulate SDF-1/CXCR4 signaling. Strategies to inhibit RGS signaling could thus be a potential method for enhancing efficiency of HSC homing and long-term engraftment, which is particularly important in the setting of CB transplantation.

Disclosures

No relevant conflicts of interest to declare.
Clinical Observation for the Treatment of Umibilical Cord Mesenchymal Stem Cells(UCMSC) on Severe Aplastic Anemia(SAA)
Authors: Liu Qingchi , Zheng Zhuanzhen
Published: October 2019

Objective To investigate the safety and the efficacy of Umibilical cord mesenchymal stem cells(UCMSC) on severe aplastic anemia(SAA).

Methods 30 cases who diagnosed SAA in our department from October 2013to January 2016 and treated with UCMSC on the basis of conventional treatment,were enrolled in this study prospectively.The cultured UCMSC using Serum-free culture system in vitro were given to patients with AA, interval of 15 days, 3 times for a course.the conventional treatments include CsA,male hormone and Chinese herb.Observed the changes of Th / Ts before and after treatmen ,Clinical efficacy and UCMSC infusion adverse reactions. Compared the efficacy of UCMSC therapy with conventional therapy of 30 cases of SAA at the same time.

Results:Among 30 cases using umbilical cord mesenchymal stem cell infusion with UCMSC each infusion volume 3.2 × 106 / kg ~ 4.2 × 106 / kg, the average of 2.9 × 106 / kg, the average influsinon three times , 8 cases acquired basic cure, 8 cases acquired remission , 6 cases acquired significant progress in 6 cases, 8 cases were ineffective,with an effective rate of 73.3%.Before UCMSC treatment Th / Ts 0.67 ± 0.41, after1.33 ± 0.45, there was significant difference (P <0.05). There were 4 patients experienced transient fever during the influsion ,disappeared after symptomatic treatment, no other serious adverse reactions.Among 30 cases of SAA recived conventional therapy at the same period , 3cases acquired basic cure, 5cases acquired remission ,8cases acquired significant progress in 6 cases, 14 cases were ineffective,with an effective rate of 53.3%.The difference was significant (P <0.05).

Conclusions:In vitro serum-free culture system can be used to culture and expand clinical scale UCMSC.UCMSC infusion can improve the SAA patient's Th / Ts proportion of T lymphocyte subpopulation , can improve the efficacy of new SAA, also more secure, no serious adverse reactions.

Disclosures

No relevant conflicts of interest to declare.
Either Scl or Lyl1 Is Required to Maintain Primitive Erythropoiesis
Authors: Chiu Sung Kai , Tremblay Cedric , Saw Jesslyn , Curtis David J.
Published: October 2019

SCL (TAL1) and LYL1 are the predominant bHLH transcription factors expressed in erythropoiesis. Using a conditional allele of Scl, we have previously demonstrated redundancy of Scl in adult erythropoiesis. Similarly, adult erythropoiesis is maintained in Lyl1-deficient mice. To determine if these factors can compensate for each other, we deleted Scl with Cre recombinase under the control of the Epo receptor, which is active in late erythroid progenitors from embryonic day 8.5. Embryos lacking Scl in erythroid progenitors (EpoR-Cre SclD/D) were born at the expected Mendelian frequency with only a mild anemia in adult mice, indicating Scl was not required for primitive or definitive erythropoiesis. In contrast EpoR-Cre SclD/D mice lacking Lyl1 died at e11.5 due to erythropoietic collapse. Gene expression profiling of yolk sacs prior to the loss of erythrocytes (E9.5) revealed reduced Gata1, Fog1, Klf1 and the primitive b-globins. In contrast, expression of Gata2 and Runx1 were increased despite absence of Scl and Lyl1. Development of embryonic stem cells from these mice will determine if Gata1 is key downstream target of the Scl/Lyl1 complex for primitive erythropoiesis.

Disclosures

No relevant conflicts of interest to declare.
Demonstration of Pharmacokinetic and Pharmacodynamic Equivalence in Healthy Volunteers for B12019, a New Proposed Pegfilgrastim Biosimilar
Authors: Roth Karsten , Gastl Barbara , Lehnick Dirk , Jacob Karin , Jankowsky Ruediger
Published: October 2019

B12019 is being developed as a biosimilar to Neulasta® (INN pegfilgrastim), a long-acting, pegylated form of recombinant human granulocyte-colony stimulating factor (r-metHuG-CSF, INN filgrastim) for the prevention of chemotherapy-induced neutropenia. A comprehensive analytical, functional and preclinical comparability program has already demonstrated a high degree of similarity of B12019 as compared to Neulasta®. In order to confirm the similarity on the clinical level, a pharmacokinetics/pharmacodynamics (PK/PD) study was conducted with B12019 in comparison to EU-authorised Neulasta®.

The study was designed as a single-dose, randomized, double-blind, two-way crossover study. The study statistics were based on a two-stage design derived from to Potvin et al, 2007 to address potential high variability for the PK endpoints. 172 healthy male volunteers were enrolled in stage 1 of the study, whereas stage 2 would allow the recruitment of additional 102 subjects. The subjects received B12019 as well as Neulasta®. The primary PK endpoints were the Area Under the Curve for concentration (AUC0-last) and maximum concentration (Cmax). The primary PD endpoint was the Area Under the Effect Curve (AUEC) for Absolute Neutrophil Count (ANC). PK endpoints were assessed with a 94.32% confidence interval (CI) accounting for the two-stage study design, whereas the PD endpoint was assessed with a 95% CI. Furthermore, safety and immunogenicity were investigated.

161 subjects were eligible to contribute to the model-based PK and PD comparison based upon the first stage of the study. For the PK endpoints, the 94.32% CIs for the geometric mean ratios were 86.60-104.73% for AUC0-last and 84.36-102.18% for Cmax. Both PK endpoints fulfilled the predefined acceptance criteria of being within the range of 80-125%. For the PD endpoint, the 95% CI for the geometric mean ratio of the ANC AUEC was 98.67-101.75%, also falling within the predefined acceptance criteria of 80-125%. Since the primary PK endpoints were met in stage 1 of the study, stage 2 was not required. The safety profile of B12019 did not show any clinically meaningful difference as compared to Neulasta®. Neither anti-G-CSF nor neutralising antibodies were detected for both, B12019 and Neulasta®.

The study demonstrated PK and PD comparability as well as comparable safety and immunogenicity profiles of B12019 as compared to EU-authorised Neulasta®. No clinically meaningful differences were detected between B12019 and Neulasta®. The high analytical and functional similarity of B12019 was confirmed on clinical level.

Disclosures

Roth: Cinfa Biotech: Employment. Gastl:Cinfa Biotech: Consultancy. Lehnick:Nuvisan GmbH: Employment. Jacob:Cinfa Biotech: Employment. Jankowsky:Cinfa Biotech: Employment.
TP53-Induced Glycolysis and Apoptosis Regulator Protects from Spontaneous Apoptosis and Predicts Poor Prognosis in Chronic Lymphocytic Leukemia
Authors: Hong Ming , Xia Yi , Zhu Yu , Zhao Huihui , Xie Yue , Fan Lei , Wang Li , Miao Kourong , Wu Wei , Zhu Huayuan , Chen Yaoyu , Xu Wei , Qian Sixuan , Li Jianyong
Published: October 2019

The circulating chronic lymphocytic leukemia (CLL) cells appear not to be overly utilizing aerobic glycolysis. However, CLL cells' recurrent contact with the stromal microenvironment leads to an increase of aerobic glycolysis and the cells' overall glycolytic capacity, which promotes cell survival and proliferation. TP53-induced glycolysis and apoptosis regulator (TIGAR) has been directly implicated in cellular metabolism in the control of glycolysis. TIGAR inhibits glycolysis and protects cells from intracellular reactive oxygen species (ROS)-associated apoptosis. Here, we investigated correlation between TIGAR expression and apoptosis in CLL primary cells, along with its relationship with the clinical characteristics and prognosis in 102 newly diagnosed CLL patients. Our data showed TIGARoverexpression correlated with protection from spontaneous apoptosis in CLL cells, and is strongly associated with advanced Binet stage, unmutated immunoglobulin heavy-chain variable region (IGHV) status, CD38 positivity, β2-microglobulin and p53 aberrations. Higher expression of TIGAR was associated with briefer treatment-free survival (median: 3 months vs. 51 months, p = 0.0108) and worse overall survival (median: 74 months vs. not reached, p = 0.0242), as well as their diverse responses to fludarabine based chemotherapy. TIGAR expression of the patients who were resistant to chemotherapy was significantly higher than those who were sensitive to chemotherapy (mean: 0.3859±0.1710 vs. 0.0974±0.0291, p = 0.0290). Taken together, our findings depict how bioenergetics characteristics could be therapeutically exploited in CLL.

Disclosures

No relevant conflicts of interest to declare.
Selective Advantage of the Recurrent Isodicentric Chromosome 20 in Myeloid Disease
Authors: Schleede Justin B , Papenhausen Peter , Schwartz Stuart , Gadi Inder K.
Published: October 2019

Interstitial deletion of the long arm of chromosome 20 is a common genomic imbalance associated with myeloid hematologic disorders. Originally identified by conventional cytogenetics and fluorescent in situ hybridization, it has been hypothesized that deletion of 20q results in haploinsuffciency of tumor suppressor gene(s) causing a proliferative advantage. The advent of microarray analysis has revolutionized the detection of copy number gains and losses associated with a variety of oncogenic mechanisms. Initial studies using microarray analysis have shown significant heterogeneity in the breakpoints associated with interstitial deletions of 20q, ruling out the possibility of a recurrent genomic fusion, but establishing a critical region which appears to support a tumor suppressor gene loss model. We have documented six individuals, referred for microarray analysis of myelodysplasia, with alterations of chromosome 20 involving a similar complex rearrangement. The rearrangement is an isodicentric chromosome 20 consisting of 1) a deletion of the majority of the short arm pter to p11.1, 2) retention of the region flanking the centromere from p11.1 to q11.21, and 3) an adjacent interstitial deletion of the long arm that can vary in size similar to the common 20q deletion. The breakpoints of the region flanking the centromere at 20p11.1 are within a block of segmental repeats and virtually identical in all 6 cases while the break points in 20q11.21 are within ~1.1 MB (30,785,977-31,887,129). In addition, one of the six cases shows additional copies of the region flanking the centromere from p11.1 to q11.21 consistent with supernumerary marker chromosomes found by conventional cytogenetics. These results suggest that there is a selective advantage associated with a gene(s) within the chromosome 20p11.1->q11.21 segment. The PDRG1 gene is localized to 20q11.21and has been shown to be upregulated in number of malignant cell types including colon, rectum, ovary, lung, stomach, breast and uterus. As our results show, PDRG1 is duplicated or amplified in all 6 cases with the isodicentric chromosome 20. Therefore, these results suggest PDRG1 upregulation may contribute to the pathogenicity of myelodysplasia by conferring a selective advantage in individuals with the isodicentric chromosome 20.

Disclosures

No relevant conflicts of interest to declare.
Apatinib Inhibits Proliferation and Induces Apoptosis of Acute Myeloid Leukemia Stem/Progenitor like Cell Line(kg1α cells) By Inhibition PI3K/AKT Signal Pathway
Authors: Xu Bing , Li Zhi-feng , Deng Man-man , Zha Jie , Zhou Yong , Fang Zhi-hong
Published: October 2019

OBJIECTIVE To investigate the effect of apatinib, a small-molecule vascular endothelial growth factor receptor-2 tyrosine kinase inhibitor, on the proliferation and apoptosis of acute myeloid leukemia stem/progenitor like cell (kg1α) and its molecule mechanism. METHODS The kg1α cells were treated with a serial of concentrations of apatinib for 48 h and 72 h, the inhibitory ratio was measured by CCK8 assay, the apoptosis percent was measured by flow cytometry. Western bolt was used to analyzed AKT/p-AKT、p-Raf and p-PTEN expression after treatment with 0、10 and 20μM apatinib. RESULTS ①After treatment with a serial of apatinib (2.5、5、10、20、40μM) on AML stem-like cell line(kg1α)for 48 and 72 hours, the cell proliferation were significantly inhibited in a dose- and time-dependent mode. For treating with 48h, the cell viability were respectively (92.32±0.82) %、 (80.59±4.95) %、 (61.75±0.47) %、 (51.51±4.10) %、 (26.42±4.20) %, and all the differences had statistical significance compared with control group, and with the 50% inhibitory concentration(IC50) value of (16.98±0.08) μM; However, the viability were respectively (88.42±2.91) %、 (70.83±4.45) %、 (47.87±1.59) %、 (31.41±3.57) %、 (13.26±1.96) % for 72h, and with the 50% inhibitory concentration(IC50) value of (10.05±0.08) μM .②The results of Annexin V/PI showed that various concentration of apatinib induced significantly apoptosis on kg1α cells. After the treatment of 10、20、30 and 40μM apatinib for 48 hours, the apoptosis percentage was significantly higher than control group (6.12±1.26) %, respectively (4.57±1.16) %、 (12.6±2.34) %、 (16.37±4.38) % and (19.77±2.55) % ,and the differences(except for 10μM) had clearly significance compared with control group(F=18.85,P<0.001);for 72h, the apoptosis percentage of 10、20、30 and 40μM apatinib were seperately 10.83±2.96 (%、 (15.38±2.78) %、 (24.95±2.70) % and (35.17±1.36) %, compared with untreated group (5.03±1.11) % had obviously significance(F=79.425,P<0.001). Western blot results indicated decrease the expression of AKT/p-AKT and p-Raf, however, upregulated the level of p-PTEN. CONCLUSION Apatinib can inhibit the proliferation and induce the apoptosis of kg1αcells. Its mechanism may be related with the PI3K/AKT signal pathway.

Disclosures

No relevant conflicts of interest to declare.