bluuebunny/crossref_metadata_2025 huggingface dataset:
Adding AB Plasma Units to Emergency Release O Negative Red Blood Cells (RBC) Issued to the Emergency Department (ED)
Authors: Andrews Jennifer C. , Weiss Susan , Viele Maurene , Goodnough Lawrence T
Published: October 2019

Abstract 4333

Background:

Recent studies have shown improved mortality and decreased early coagulopathy in trauma patients with administration of RBCs, platelets and fresh frozen plasma in a 1:1:1 ratio (Shaz BH et al. Transfusion 2010;50:493–500). As a result, massive transfusion protocols (MTP) have become common at large academic institutions with level I trauma programs. Our institution recently added 2 units of AB plasma to our standard ED trauma cooler which contains 2 units of O negative RBCs and which is ordered by ED staff upon notification of an incoming trauma patient. The treating physicians can escalate blood component support by ordering an MTP if indicated. The purpose of this study was to assess the use and wastage of plasma and RBCs in the ED trauma cooler.

Methods:

Orders received for the trauma cooler from January 1 2011 to June 30 2011 at a large academic Trauma I center were reviewed retrospectively. Transfusions of RBC and plasma for each order were assessed as well as wastage of plasma or RBCs not transfused. Patients may have used more blood products than initially issued in the ED trauma cooler.

Results:

Ninety one orders were received for an ED trauma cooler during the 6-month period assessed. Fifteen (16%) of 91 orders resulted in transfusion of RBCs. Nine (10%) of 91 orders resulted in transfusion of plasma. Five of 182 (3%) plasma units issued were wasted because temperature parameters were exceeded before return to the transfusion service (TS). No untransfused RBC units were wasted and all were returned to available inventory. See table. The 15 O negative RBC units transfused from the ED trauma cooler during this time period represent 0.8% (15/1891) of our medical center's transfused O negative RBC units, and the 14 AB plasma units transfused or wasted from the ED trauma cooler represent 2% (14/573) of the AB plasma units transfused by the TS.

Conclusion:

Of the 48 academic hospitals participating in the 2009 University Health System Consortium (UHC) Efficient Blood Management Benchmarking Project, the published mean wastage of plasma was 3.36% (range 0.36% – 9.44%). Our 3% wastage rate for AB plasma in the ED trauma cooler was within this range. The addition of 2 units of AB plasma to the ED trauma cooler with 2 units of O negative RBCs was feasible at our institution with acceptable wastage of plasma and no undue strain on our supply and inventory of donor O negative RBC and AB plasma.

Disclosures:

No relevant conflicts of interest to declare.
The Burden of Blood Transfusion: A Utilization Analysis - A Pilot Study
Authors: Reitan John , Kudrik Fred J. , Fox Kayleen , Shreay Sanatan , van Breda Arletta , Corey-Lisle Patricia
Published: October 2019

Abstract 4342

Purpose:

The primary objective of this pilot study is to evaluate the burden of blood transfusion on the institutional outpatient transfusion center. The burden of blood transfusion will be characterized by quantifying time and personnel resources associated with the administration of blood transfusions.

Methods:

This pilot study is a retrospective chart review (starting July 1, 2010, working backwards until 100 evaluable patients are accrued) that evaluated time and resources of blood transfusions at a single local institutional transfusion center located in the US. The record review currently includes 80 RBC transfusions. The mean and standard deviation (SD) were calculated for patient’s age, pre-transfusion Hgb level and other transfusion related activities.

Results:

The mean (SD) patient age was 59 (12) years, and the mean pre-transfusion Hgb was 8.0 (0.8) g/dL. The majority of RBC units administered were 2 units (n =66) with one unit received by n=8 and three units received by n=6. Mean (SD) times to administer the first, second and third units were 102 (12) minutes, 101 (19) minutes and 93 (14) minutes, respectively. The mean (SD) duration of a patient visit to the transfusion center was 313 (59) minutes, with a range of 135 – 448 minutes. When one, two and three units were administered, mean (SD) visit duration was 203 (40) minutes, 321 (46) minutes and 375 (25) minutes, respectively.

Conclusions:

In this preliminary analysis when transfusion is administered, the majority of patients received 2 units of RBCs and the mean time to administer these 2 units was about 3.3 hrs. The duration of patient visits on average was 5.2 hrs which could be a considerable burden on cancer patients. The cost and labor quantification of mean time of administering RBC transfusion would elicit burden to the transfusion center.

Disclosures:

Reitan: Amgen: Research Funding. Shreay:Amgen: Employment. Corey-Lisle:Amgen: Employment.
Platelet Usage Trends-Could It Be Less?
Authors: Kocak Ulker , Belen Fatma Burcu , Okur Arzu , Kulali Ferit , Egritas Odul , Yenicesu Idil
Published: October 2019

Abstract 4343

Thrombocytopenia as a risk factor for major bleeding is a life threatening complication of critical disease situations. Prophylactic or therapeutic platelet transfusions are widely used for hemorrhagic problems.

The aim of this study was to find out the indications, the threshold for platelet transfusion for the pediatric patients in a tertiary care hospital throughout one year to determine the optimal platelet transfusion practice.

The study was conducted upon the records of the hospital's blood bank and the files of the patients were retrospectively reviewed.

A total of 378 platelet units were used in 104 patients with an age range of 0–18 years. The majority (48.1 %) of the transfusions were in mainly leukemia patients in the hematology department. Transfusions per patient were higher in hematology (mean 4.8 ± 7.4 transfusions) and neonatology (mean 4.8 ± 7.6 transfusions) (p > 0.05) departments.

256 (67.7 %) of the transfusions were single donor apheresis whereas 122 (32.3 %) were random donor platelet products. About 1/3 of the apheresis products were given when the platelet count was below 10 × 109/L and 1/3 were given when the platelet count was between 10–20 × 109/L. 118 (31.2 %) of the transfusions were used for prophylaxis in hematology and oncology departments when the platelet levels were below 10×109/L (mean platelet count of 9.8 ± 7.7 × 109/L and 8.8 ± 5.2 × 109/L). These levels are lower than other departments (p < 0.001). 59 % of the apheresis units were preferred for therapeutic reasons. The highest platelet levels were observed when platelet concentrates were used for treatment of bleeding or prior to any procedure (Table 1).

Platelet usage shows a discrepancy between the departments of pediatrics. Hematology and oncology patients were the major consumers of the platelet concentrates. Although it is known that when given in appropriate dosage no difference was observed between the platelet concentrates in terms of increasing the platelet counts, there was a tendency for using apheresis products which are the most expensive (188.61 dollars) among all. This practice could be abandoned by continuous education. Besides the guidelines should be reviewed periodically to remind the physicians the dilemma of transfusion; life saving on one hand and the risks on the other.

Disclosures:

No relevant conflicts of interest to declare.
Efficacy of Baxter’s Longer Acting rFVIII (BAX 855) in Mouse Models of Hemophilia A
Authors: Schiviz Alexandra , Barbara Eder , Lawo John-Philip , Ehrlich Hartmut J. , Scheiflinger Friedrich , Turecek Peter L , Schwarz Hans Peter , Muchitsch Eva-Maria , Hoellriegl Werner
Published: October 2019

Abstract 4350

Factor VIII (FVIII) is a critical component of the intrinsic coagulation pathway. FVIII concentrates are used in patients with hemophilia A to provide a hemostatic FVIII level sufficient to treat and prevent bleeding episodes. Multiple prophylactic administrations per week are necessary to maintain a FVIII level of at least 1% of normal to effectively prevent or reduce spontaneous bleeding episodes. A longer acting FVIII concentrate would reduce the frequency of infusions. Baxter and Nektar have developed a longer-acting PEGylated variant (BAX 855) of Baxter’s recombinant FVIII (ADVATE process). The aim of the presented studies was to evaluate the efficacy of BAX 855 in two primary pharmacodynamics in hemophilia A mice: a tail-tip bleeding model (TTBM) and a carotid occlusion model (COM).

BAX 855 was tested at a dose of 200 IU/kg rFVIII. ADVATE™ served as the active reference item, and formulation buffer as the negative control item. Mice received intravenous treatment with either BAX 855 or ADVATE™ 12–40h prior to the experiment. Buffer was administered 5–15min prior to the experiment.

Animals were anesthetized using ketamine and xylazine. In the TTBM (n=16) the tip of the tail was cut off and total blood loss [mg] was assessed over 60 minutes. In the COM study (n=10) the left carotid artery was exposed and the endothelium was denuded by topical application of FeCl3. Time to occlusion [min] was assessed.

Buffer-treated animals had a median blood loss of 951 mg in the TTBM and did not show vessel occlusion within 30 min in the COM. In the TTBM, a prophylactic effect of ADVATE™ could be observed for up to 18 h, leading to a reduction in blood loss to 121 mg after this treatment interval. Prophylactic efficacy after treatment with BAX 855 could be observed for up to 40 h with a median blood loss of 73 mg after 30 h and 436 mg after 40 h. In the COM, treatment with ADVATE™ 12 h before the experiment shortened the time to occlusion to 3.8 min. With BAX 855, a similar efficacy could be observed for up to 24 h after administration (5.2 min).

In summary, the results of our studies show that BAX 855 is efficacious in these two mouse models of hemophilia A and that the efficacy of BAX 855 is prolonged compared with an unmodified rFVIII. Furthermore, the intravenous injection of BAX 855 was well tolerated in all animals across all treatment groups without any signs of acute toxicity.

Disclosures:

Schiviz: Baxter Innovations GmbH: Employment. Barbara:Baxter Innovations GmbH: Employment. Lawo:Baxter Innovations GmbH: Employment. Ehrlich:Baxter Innovations GmbH: Employment. Scheiflinger:Baxter Innovations GmbH: Employment. Turecek:Baxter Innovations GmbH: Employment. Schwarz:Baxter Innovations GmbH: Employment. Muchitsch:Baxter Innovations GmbH: Employment. Hoellriegl:Baxter Innovations GmbH: Employment.
Plasminogen Activator Inhibitor-1 and Vitronectin Levels in Children with Acute Lymphoblastic Leukemia, and Their Relationship with Risk Groups
Authors: Ayduran Semra , Devecioglu Omer , Unuvar Aysegul , Ekmekci Hakan , Karakas Zeynep , Ozturk Gulyuz , Anak Sema , Agaoglu Leyla , Kavakli Kaan
Published: October 2019

Abstract 4365

Fibrinolytic system plays a role in tumor growth, invasion and metastasis in cancer. High levels of plasminogen activator inhibitor-1 (PAI-1) which is a part of fibrinolytic system is a sign of poor prognosis in a number of cancer types. Acute lymphoblastic leukemia (ALL) is the most frequent malignant disease of childhood. Risk groups of ALL are used to determine treatment strategies in childhood ALL. In this study, we aimed to ascertain the prognostic and predictive roles of PAI-1 and vitronectin in children with ALL, and their relationship with risk groups of ALL. In the literature, there are few studies with PAI-1, but no study with vitronectin and ALL in children. In this study, we analyzed the plasma PAI-1 and vitronectin levels of 37 recently diagnosed ALL patients (age range: 9 months–17 years), and 25 age-matched healthy children as a control group by using the ELISA technique. The mean PAI-1 levels was 27.36±16.53 ng/ml, vitronectin levels 97.54±29.82% in the ALL group; and, 23.60±10.44 ng/ml, 85.50±20.85% in the control group, respectively. It was found that the plasma vitronectin levels of ALL group was high as nearly statistically significant (p=0.06). Although, the PAI-1 levels of ALL group was higher than the control group, there was no statistically significance (p=0.27). The ALL group was divided into three risk groups (standard, moderate, and high) according to the BFM protocol. When the standard and moderate-high risk groups are compared with control group, the vitronectin levels of standard risk group was higher than the control group (p=0.048), but no difference was found for PAI-1 levels (p=1.00). There was no significant difference for vitronectin levels (p=0.76), and PAI-1 levels (p=0.25) between moderate-high risk groups and the control group, although PAI-1 levels were higher in moderate-high risk groups. In conclusion, we could not find any relationship between the parameters of fibrinolytic system and ALL risk groups except the vitronectin levels of standard risk group, new studies are needed.

Disclosures:

No relevant conflicts of interest to declare.
Growth Inhibition and Induction of Apoptosis in K562 Human Chronic Leukemic Cells by abnobaVISCUM® flaxini, Mistletoe Extracts From Ash Tree: Involvement of Activation of Caspases, Mcl-1 Down-Regulation, and ER Stress
Authors: Do Young-Rok , Park Yu-Kyung , Jang Byeong-Churl , Kim Jin Young , Kwon Ki Young
Published: October 2019

Abstract 4418

Evidence suggests that extracts from mistletoe (Viscum album L.) have anti-cancer effects on many human cancers. Little is known about the relationship between mistletoe extracts and hematological malignancies. In this study, we investigated the effect of abnobaVISCUMÒ Flaxini, a commercially available mistletoe extracts from ash tree, on growth of K562 human chronic leukemic cells. Results of cell count and DNA fragmentation assays demonstrated that there was strong growth suppression and induction of apoptosis in K562 cells after treatment with abnobaVISCUMÒ Flaxini at 2 mg/ml for 8 h. Western blot and RT-PCR analyses revealed that abnobaVISCUMÒ Flaxini treatment also led to activation of caspase-9 and -3 and PARP cleavage, but had no effect on expression of death receptor (DR)−4 and −5 in K562 cells. Expression of Bak and Bax was not changed, but that of Mcl-1 protein was strongly repressed in abnobaVISCUMÒ Flaxini-treated K562 cells. Moreover, while treatment with abnobaVISCUMÒ Flaxini decreased phosphorylation of ERK-1/2 and PKB, it stimulated phosphorylationof JNK-1/2 and p38 MAPK in K562 cells. Importantly, pharmacological inhibition studies demonstrated that treatment with z-VAD-fmk (a pan-caspase inhibitor), but not SP600125 (a JNK-1/2 inhibitor) or SB203580 (a p38 MAPK inhibitor), suppressed abnobaVISCUMÒ Flaxini-induced loss of survival and apoptosis in K562 cells, and z-VAD-fmk also blocked abnobaVISCUMÒ Flaxini-induced Mcl-1 protein down-regulation. Induction of ER stress was also seen in abnobaVISCUMÒ Flaxini-treated K562 cells, as judged by increased phosphorylation of eIF-2a and expression of GRP78 and CHOP, three known ER stress sensors. Collectively, these findings provide the first evidence of abnobaVISCUMÒ Flaxini-mediated anti-survival and apoptosis inducing effects on K562 cells via activation of caspases, Mcl-1 down-regulation, and triggering ER stress.

Disclosures:

No relevant conflicts of interest to declare.
Alantolactone Induces Apoptosis in Imatinib-Sensitive and -Resistant CML Cells by Inhibiting NF-KappaB Signaling and Degrading Bcr/Abl Protein
Authors: Yan Hua , Wei Wei , Zhao Shan , Liu Chuanxu , Wu Ying-Li , Chen Guo-Qiang
Published: October 2019

Abstract 4419

Chronic myelogenous leukemia (CML) is characterized by the t(9;22) chromosomal translocation, which results in the generation of Bcr-Abl oncogene that encodes the chimeric Bcr-Abl protein with constitutive tyrosine kinase activity. Despite the great success of imatinib therapy in CML, not all patients, especially those in the accelerated or blastic phase, experience an optimal response to this drug or even to the more potent, second-generation TKIs. The emergence of resistance to TKIs has also become a significant problem implying that physiopathologically relevant cellular targets should be validated, such as NF-kappaB (NF-kB).

Here we describe the activity of alantolactone, a small natural compound and see its effect on K562-sensitive (K562s, sensitive to imatinib) and K562-resistant (K562r, resistant to imatinib) cells.

Results:

Firstly, we used CCK-8 assay to determine the cell viability and found that K562s were inhibited up to 64.5% after 48 hours of incubation of 10uM concentration of alantolactone, Annexin V+ cells were up to 68% which were confirmed by morphological observation, and apoptosis-related proteins were activated during the treatment, suggesting its inducing-apoptosis ability. Secondly, incubation of alantolactone could inhibit NF-kB signaling activity, proved by inhibition of IkBa phosphorylation and blockade of nuclear translocation of p65, while the luciferase assay further revealed the inhibited transcription of NF-kB dependent reporter gene, which was confirmed by immunofluorescence staining. Of note, Bcr-Abl protein was specifically and rapidly down-regulated as early as 6 hours after 10uM of alantolactone treatment. And alantolactone-induced Bcr-Abl down-regulation could be partially reversed by the autophagy inhibitor chloroquine but not by the proteasome inhibitor MG132 or bortezomib, indicating that autophagy may contributes to Bcr-Abl reduction. Furthermore, similar concentration of alantolactone could also induce apoptosis in K562r cells and inhibited the growth of primary CD34+ leukemic cells from CML patients.

Collectively, our results primarily demonstrated that alantolactone could induce apoptosis in CML cells, regardless of imatinib-sensitive or -resistant, by inhibiting NF-kB signaling and depleting Bcr/Abl through a unique mechanism. Alantolactone could be used as a probe for deciphering the mechanism of Bcr-Abl destruction and NF-kB inhibition, and this agent may be a promising candidate for treating CML patients.

Disclosures:

No relevant conflicts of interest to declare.
Busulfan Exposure Decrease CCyR Rate On Imatinib Therapy - Impact On Recent Pretreatment
Authors: Dyagil Irina , Lomaia Elza , Abdulkadirov Kudrat , Goryunova Elena , Lazorko Natalia , Romanova Ekaterina , Sbityakova Eugenia , Abdulkadirova Alla , Martinkevich Irina , Udalieva Vera , Usacheva Elena , Zotova Irina , Shuvaev Vasiliy , Machulaitene Elena , Poznyak Ekaterina , Ivanova Maria , Ilina Natalia , Koryagina Elena , Shneider Tatiana , Stepanova Svetlana , Dmitrenko Irina , Babkina Natalia , Perekhrestenko Tatiana , Masliak Zvenyslava , Zaritskey Andrey
Published: October 2019

Abstract 4452

Imatinib (IM) is now used world-wide as a first line chronic myeloid leukemia (CML) treatment. Although some time lack may exist between diagnosis and IM treatment.

Earlier (Blood 2009, 114: Abstract 4278) we have shown that in CML chronic phase (CP) pts with very long history of the disease(more than five years)the pretreatment by Busulfan was the adverse prognostic factor on Imatinib therapy.

Now we extend our study by a population of pts recently and for rather short time pretreated by busulfan.

Aim.

To investigate the effect of busulfan pretreatment on survival and responses to imatinib in CML pts in late CML CP.

Materials and methods.

In retrospective study 85 pts with CML CP from St-Petersburg, Leningrad region (Russian Federation) and several Ukrainian centers were included. The main inclusion criteria were: CML late CP (the duration of the disease more than 6 mos before IM start), IM therapy in routine clinical practice at least 12 months. The median time of IM therapy was 42,9 mos (12–97 mos), the median age of pts at the IM start was 49,5 years (19–83), male/female ratio 31/54.

23 patients were pretreated with busulfan (the study group) and 62 were not (control group). These groups were equal by age, sex, the median time from diagnosis to the IM start (28,3 mos in the study group and 23,9 in the control group), Sokal risk groups. Median time of busulfan pretreatment was 3,9 mos (1–62 mos). Statistical analysis was performed with SPSS 17.

Results.

In the whole group of patients frequency of complete cytogenetic response (CCyR) was 60% (51/85), estimated overall survival (OS) by 5 years from IM start was 87% (death rate 7% - 6/85). In the study group CCyR rate was significantly lower, than in the control group: 34,8% (8/23) and 69,35% (43/62), respectively, p=0,038. Estimated OS by 5 years was 72% (death rate 17% - 4/23) for busulfan-pretreated pts and 95% (death rate 3% - 2/62) for the control group, p<0,01.

Interestingly, that in the group of shortly busulfan-pretreated pts (the duration of pretreatment ≤6 mos), the lower CCyR rate has also been observed – 31% (4/13), although all other parameters were seemed equal to the control group (median time before IM start 28,3 mos).

Conclusion.

The pretreatment with busulfan impaired negatively the efficacy of imatinib treatment in CML late CP patients. Even short pretreatment (less than 6 mos) had adverse effect on CCyR. The mechanism is unclear. Busulfan pretreatment before imatinib therapy should not be used.

Disclosures:

No relevant conflicts of interest to declare.
Clinical Relevance of Pre- and Post-Transplantation Residual Disease, Hematogones and Natural Killer Cells in Adult Patients with Acute Leukemia
Authors: Li Chi-Cheng , Tang Jih-Luh , Lin Chien Ting , Ko Bor Sheng , Yao Ming , Loken Michael R.
Published: October 2019

Abstract 4490

Purpose

Allogeneic hematopoietic stem cell transplantation (allo-SCT) may rescue patients with acute myeloid and lymphoblastic leukemia (AML/ALL) even in high risk disease. Little has been investigated about prognostic impact of pre- and post-transplant residual disease (RD) status, hematogones (HG) level and natural killer (NK) cells level in the adult patients with acute leukemia.

Materials & Methods

We retrospectively studied 45 consecutive adult patients, including 31 AML and 14 ALL, receiving allo-SCT at National Taiwan University Hospital. Based on disease status at transplant, 18 patients were in standard risk; 16 in intermediate risk; and 11 in high risk disease. 28 patients underwent myeloablative conditioning and 17 received reduced-intensity SCT. Bone marrow samples were obtained both before conditioning therapy (pre-SCT) and at hemogram recovery after transplantation (post-SCT). RD, HG level and NK cells level were evaluated by multidimensional flow cytometry. Leukemia-free survival (LFS) was estimated by using Kaplan-Meier method.

Results

Pre-SCT RD-negative (RD-) was shown in 26 patients and RD-positive (RD+) in 19 patients; whereas post-SCT RD- was found in 39 patients and RD+ in 6 patients. Level of pre-SCT RD status ranges from 0–90% and level of post-SCT RD status ranges from 0–3%. Estimated one-year LFS were 71.8% in pre-SCT RD- compared to 0% in pre-SCT RD+ group (P=0.008, Fig. 1). In post-SCT flow cytometric analysis, one-year LFS were 33.2% in post-SCT RD- versus 25.0% in post-SCT RD+ group (P=0.002, Fig. 2). After combining pre- and post-SCT RD status, three groups of patients were identified; pre-SCT RD−/post-SCT RD- (n=26), pre-SCT RD+/post-SCT RD- (n=13), and pre-SCT RD+/post-SCT RD+ (n=6). The classification further stratify the patients into different groups with distinct outcome (P=0.003, Fig. 3). We also found the median HG 0.2% (range 0–12%) and 0.1% (range 0–2.1%) and median NK cells 1.8% (range 0.2–20%) and 1.8% (0.02–15.5%) pre-SCT and post-SCT, respectively. The patients with higher NK cells, either pre-SCT or post-SCT, had longer LFS (P<0.001 and 0.004, continuous variables, respectively).

Conclusion

Our data suggest both pre- and post-SCT RD status and NK cells level could predict the transplant outcome. Further large-scale studies may be needed to confirm this point.

Disclosures:

No relevant conflicts of interest to declare.
Expression of CD56 in Acute Myeloid Leukemia (AML) Is Associated with Poor Outcome When Patients Treated with Stem Cell Transplant in Second Remission but Not in the First Remission
Authors: Chaudhri Naeem A. , Almhareb Fahed , Walter Claudia Ulrike , Nounou Randa , Khalil Salem , Bakshi Nasir , Owaidah Tarek , Al Zahrani Hazzaa , Mohamed Said , Rasheed Walid , Alsharif Fahad , Aljurf Mahmoud , Albitar Maher
Published: October 2019

Abstract 4880

Background:

The CD56 is a neural cell adhesion molecule (N-CAM) expressed in natural killer cells. CD56 expression on the surface of leukemic cells in acute myeloid leukemia (AML) has been reported to correlate with higher rate of relapse and to be associated with poor outcome when patients are treated with conventional intensive chemotherapy. The role of allogeneic hematopoietic stem cell transplantation (HSCT) in improving outcome in this subgroup of AML patients is not known. We investigated whether the outcome of CD56-positive AML patients differ from average AML patients when treated with allogeneic HSCT after intensive chemotherapy and its prognostic relevance in patients transplanted in the first (CR1) or second complete remission (CR2).

Methods:

Immunophenotyping data of 106 patients with AML who were treated with allogeneic HSCT was reviewed. The expression of the CD56 on the blast population as determined by flow cytometry was correlated with clinical data and outcome. The median age of patients was 24 (range: 14–57) and included 72 patients in the intermediate cytogenetic group and 30 in the adverse cytogenetic group. Of the 106 patients, 71 (67%) were treated with HSCT in their first remission (CR1) and the remaining patients were treated in CR2.

Results:

Twenty nine (28%) of all studied patients expressed CD56 on the surface of the blasts at diagnosis. When we consider only patients treated with HSCT in CR1, 17 (24%) were CD56 positive. The overall survival (OS) and event free survival (EFS) for patients who were positive for CD56 were not significantly different from the rest of the patients in the CR1 group. In contrast, in the group of patients who were treated with HSCT after relapsing (CR2), patients with CD56 positivity had significantly shorter OS (P=0.03) and DFS (P=0.006). In this group, 11 of the 35 (31%) were CD56+ and their median survival was 8.85 months while the median has not been reached in the CD56 negative patients.

Conclusion:

Our data suggests that the expression of CD56 in AML is a significant poor prognostic factor if patients are treated with HSCT after relapsing in their second remission. Although this should be confirmed in larger studies, the CD56 expression in AML should be considered as an adverse risk factor and favors treatment with allogeneic HSCT in the first remission and not after relapse in second remission. AML patients with CD56 positive blasts have higher incidence of relapse after chemotherapy and they might lose their opportunity of good outcome if HSCT is delayed and performed only in the second remission.

Disclosures:

No relevant conflicts of interest to declare.
Serum Free Light Chains Identify Patients With Monoclonal Gammopathy At Increased Risk Of Developing Renal Disease
Authors: Johnson Lawrence R. , Miller Gerald C. , Stout Spencer R.
Published: October 2019

Background

Monoclonal gammopathy (MG) is frequently associated with kidney disease (monoclonal gammopathy of renal significance; MGRS). International guidelines recommend the use of the serum free light chain (FLC) assay when screening for monoclonal gammopathies; an abnormal FLC ratio and an elevation of at least one FLC isotype indicates an Ig LC clone. However it is not known if, in patients with normal kidney function, the presence of a FLC clone predisposes to the future development of kidney disease.

Patients and Methods

A 22485 patient database with multiple laboratory tests and up to 15-year follow up (Regional Medical Laboratory, Tulsa, OK, USA) was analysed. There were 425 confirmed MG patients with no progression to multiple myeloma. Median age was 72 years (22-97 years) and M/F ratio was 177/248. FLC kappa and lambda were measured nephelometrically with Freelite® (The Binding Site Ltd, Birmingham, UK). Baseline was defined on the first available FLC measurement; this was available at inception in 258 (61%) patients and during follow-up in 167 (39%) patients. 128 patients with MG and an eGFR <60ml/min/1.75m2 and/or a clinical diagnosis of renal disease (ICD9 code) prior to and up to 100 days after baseline were excluded from the analysis. Statistical analyses, including Receiver Operating Characteristics (ROC) to identify monoclonal FLC (iFLC) cut-off were carried out using SPSS v21.

Results

297 (of 425) patients had a MG with normal renal function at baseline; 118 (40%) patients had an abnormal FLC ratio (88 kappa, median: 35.5 (7.7-2675) mg/L; 30 lambda, median: 83.6 (0.9-3552) mg/L). 21/297 (7%) patients developed renal impairment during the course of follow up (median: 852 (114-2388) days); 15/21 (71%) had monoclonal FLC production. In these patients there was no association between iFLC (median: 47.8 (0.9-3552) mg/L) and creatinine (median: 0.9 (0.5-1.5) mg/dL) levels (p=0.3966). Time to renal impairment (TTRI) was significantly shorter for patients with an abnormal v normal FLC ratio (75% TTRI: 1261 days v not reached (NR), hazard ratio (HR): 5.31; p<0.001). Age between groups was similar (median: 68 (22-97) v 72 (39-92) years; p=0.0544). In 118 patients withan abnormal FLC ratio, Cox regression analysis identified iFLC concentrations as being associated with the development of impaired renal function (p=0001). Furthermore, patients with iFLC>100mg/L (n=25) were at a significantly increased risk of developing renal impairment compared with patients with iFLC<100mg/L (n=93) (75% TTRI: 874 days v NR, respectively; HR: 6.10; p<0.001).

Conclusions

These results indicate that the presence of an Ig LC clone is associated with an increased risk of people with MG and normal kidney function progressing to renal disease.

Disclosures:

No relevant conflicts of interest to declare.
Role Of RECQ1 Helicase In Multiple Myeloma Biology and Drug Resistance
Authors: Viziteu Elena , Klein Bernard , Bruyer Angelique , Hose Dirk , Goldschmidt Hartmut , Grandmougin Camille , Basbous Jihane , Constantinou Angelos , Moreaux Jerome
Published: October 2019

Multiple Myeloma (MM) is a still lethal disease in 2013 characterized by the accumulation in the bone marrow of a clone of malignant plasma cells.

Recent studies have shown that epigenetic modifications play a role by silencing various cancer-related genes in MM. We initiated a microarray-based genome-wide screen for genes responding to DNMT inhibition in MM cells and built a “DNA methylation gene score” that makes it possible identification of myeloma patients that will be sensitive to DNMT inhibitors. Among the genes regulated by DNMT inhibitor and associated with the worst prognostic value in patients, RECQ1 was identified. RECQ helicase are DNA unwinding enzymes involved in the maintenance of chromosome stability. RECQ1 is highly expressed in various types of solid tumors. RECQ1 silencing in cancer cells results in mitotic catastrophe and prevents tumor growth in murine models. In glioblastoma cells, depletion of RECQ1 induces reduction in cellular proliferation, spontaneous γ-H2AX foci formation and hypersensitivity to drugs. Furthermore, it was described that RECQ1 protein could interact with MSH proteins, RAD51 and PARP1 involved in DNA repair pathways.

RECQ1 protein is expressed in human myeloma cell lines (HMCLs) and primary myeloma cells of patients. In four HMCLs (XG2, XG7, XG19 and LP1), RECQ1 was downregulated by conditional shRNA expression through lentiviral delivery. RECQ1 knock down inhibits growth of myeloma cells, induces 53BP1 foci formation and apoptosis. RECQ1 depletion sensitizes myeloma cells to DNA alkylating agent (melphalan) but not to corticosteroid (dexamethasone) or proteasome inhibitor (bortezomib). Using immunoprecipitation of myeloma cell nuclear proteins with anti-RECQ1 antibody, RECQ1 was shown to interact with PARP1 but not RAD51 or MSH2. An increased association of the two proteins was found upon DNA damages induced by melphalan. In agreement, RECQ1 depletion sensitizes myeloma cell lines to the PJ34 hydrochloride hydrate PARP inhibitor.

In conclusion, RECQ1 could represent a biomarker of drug resistance in MM, which is targeted by DNMT inhibitor. This suggests association of alkylating agents and/or PARP inhibitors with DNMT inhibitor may represent a promising therapeutic approach.

Disclosures:

Goldschmidt: Celgene and Janssen: Membership on an entity’s Board of Directors or advisory committees.
Anemia, Shortened Erythrocyte Lifespan and Stomatocytosis In a Flippase Mutant Mouse Strain
Authors: Lucy Coupland , Yabas Mehmet , Cromer Deborah , Winterberg Markus , Teoh Narcissus , Kirk Kiaran , Broer Stefan , Parish Christopher , Enders Anselm
Published: October 2019

Background

Mammalian erythrocytes are anucleate biconcave discs with marked flexibility and deformability, features necessary for efficient function. The cell membrane consists of a lipid bilayer containing proteins and an asymmetric distribution of phospholipids (PL). Phosphatidylcholine and sphingomyelin are predominantly concentrated in the outer layer and phosphatidylserine (PS) and phosphatidylethanolamine are mainly confined to the inner layer of the erythrocyte membrane. Maintenance of PL asymmetry is essential for erythrocyte survival and function as increased externalization of PS results in adherence of erythrocytes to vascular endothelium, activation of plasma blood clotting factors and premature removal from the circulation. Flippases, floppases and scramblases are the enzymes responsible for the establishment and maintenance of PL distribution.

A mouse deficient in the flippase ATP11C was generated through ENU mutagenesis and was found to have reduced hemoglobin in comparison to wild type (WT) littermates. This study was performed to characterize the etiology of anemia in these mice.

Results

ATP11C-deficient mice had significant reductions in erythrocyte numbers and hematocrit compared to WT but higher MCH and MCV. Reticulocyte numbers were comparable to WT as were serum iron parameters. Bone marrow and splenic erythropoiesis was normal in ATP11C mutant mice, however, erythrocyte lifespan was reduced by 35%. A marked increase in the frequency of PS+ erythrocytes was demonstrated in ATP11C mutant mice and was shown to increase with erythrocyte age. Bone marrow and splenic erythroblasts from ATP11C-deficient animals displayed a lower rate of PS translocation in vitro compared to WT confirming defective flippase activity.

Erythrocytes and late-stage splenic erythroblasts in ATP11C mutants were significantly larger than WT based on flow cytometry. SEM revealed the majority of mutant erythrocytes displayed stomatocyte-like morphology, as confirmed on peripheral blood smears. The osmotic fragility of the ATP11C-deficient erythrocytes was comparable to WT erythrocytes as was Na+ and K+ homeostasis.

Discussion

These studies reveal the important role of flippases in maintaining normal erythrocyte function through the maintenance of the asymmetric distribution of PS within the membrane. Perturbation of this process, as seen in the ATP11C-deficient mice, results in (i) increased exposure of PS on the erythrocyte outer membrane, (ii) increased size of late-stage erythroblasts and erythrocytes with stomatocyte formation (iii) reduced erythrocyte lifespan and (iv) normochromic anemia. This study, therefore, reveals a new mechanism for stomatocytosis and raises the question of whether mutations in ATP11C may serve as a previously unclassified cause of anemia in humans.

Disclosures:

No relevant conflicts of interest to declare.
Slit2N Inhibits Transmission Of HIV-1 From Dendritic Cells To T-Cells By Modulating Novel Cytoskeletal Elements
Authors: Shrivastava Ashutosh , Prasad Anil , Kuzontkoski Paula , Yu Jinlong , Groopman Jerome E.
Published: October 2019

During sexual contact, HIV-1 can enter the mucous membranes of the vagina, throat, or rectum. There, the virus encounters potential target cells, including antigen-presenting cells such as dermal dendritic cells (DC), Langerhans cells, and plasmacytoid DCs, which play pivotal roles in both innate and adaptive immunity. All DC subsets are susceptible to HIV-1 infection or internalization of intact virus, and appear to facilitate cell-mediated transmission of HIV-1 in the mucosa and lymphatic tissues.

Once internalized, HIV-1 can be transmitted from DCs to T-cells at a specialized junction termed the infectious synapse (IS). Recent studies reveal that several actin-rich DC membrane extensions, including actin filopodia, membrane sheets, cytonemes, and nanotubes, establish direct contact between cells at the IS and facilitate HIV-1 transmission. Although the virus can infect T-cells directly (cis-infection), transmission of HIV-1 from DCs to T-cells at IS (trans-infection and de novo trans-infection) appears to be some 10 to 100 times more efficient than direct infection; therefore, cell-to-cell transmission is an attractive target for novel HIV therapeutics.

We evaluated the effects of Slit2N, a ligand for the Roundabout (Robo) receptors, on the transmission of HIV-1 from DCs to T-cells. Slit2N inhibits HIV-1 transmission to T-cells and its replication. Binding of HIV-1 to DC-SIGN on the surface of DCs initiates signaling through cellular-Src (c-Src) and cell division control protein 42 (Cdc42), which modulates actin polymerization and filopodia formation via the actin-related protein 2/3 (Arp2/3) complex and mammalian Diaphanous 2 (mDia2). Slit2N inhibited HIV-1-induced signaling through both Arp2/3 and mDia2, decreased filopodial extensions on virus-infected DCs, and inhibited cell-to-cell transmission of HIV-1 at the IS in a Robo1-dependent manner. Of particular note, we identified Flightless-1 (Fli1) as a novel, Robo1-interacting protein, and demonstrated that it is required for efficient cell-to-cell transmission of HIV-1.

These results suggest that targeting the Slit/Robo pathway involves the modulation of previously unrecognized cytoskeletal elements in HIV-1 transmission, and thus, offers an innovative strategy to limit viral infection in the host.

Disclosures:

No relevant conflicts of interest to declare.
Biologic Response and Adverse Events To Stimate In Patients With Von Willebrand Disease
Authors: Song Xiangqian , Valentino Leonard A. , Simpson Mindy L. , Boggio Lisa
Published: October 2019

von Willebrand factor (VWF) is a large multimeric glycoprotein secreted from platelet α-granules and Weibel-Palade bodies of endothelial cells. VWF mediates the initial adhesion of platelets at sites of vascular injury and binds to and stabilizes blood coagulation factor VIII in the circulation to protect it from inactivation and clearance. von Willebrand disease (VWD) is the most common hereditary bleeding disorder and results from the deficiency or dysfunction of VWF leading to mucocutaneous bleeding, including epistaxis, menorrhagia, and excessive bleeding after trauma or surgery. Bleeding in patients with VWD is treated with infusion of plasma-derived VWF containing FVIII concentrates or 1-desamino-8-D-arginine vasopressin (DDAVP, desmopression). DDAVP is an analog of vasopressin antidiuretic hormone which can stimulate the exocytosis of VWF from storage sites and increase VWF and FVIII levels. DDAVP can be administrated by intravenous and intranasal routes. There are several reports of the safety and efficacy of intravenous DDAVP, but few with concentrated intranasal DDAVP (Stimate®). Here we report on the efficacy and safety of Stimate® in patients with VWD

Methods

Hospital records for 72 patients with VWD who received Stimate® from 1998 to 2012 were reviewed. The primary endpoint was the patients’ biological response to Stimate®, defined as at least a 3 fold increase compared to baseline of both ristocetin cofactor activity (VWF:RCo), factor VIII procoagulant activity (FVIII:C), or both VWF:RCo and FVIII:C are over 100% after Stimate®. Individuals not meeting the response criteria were deemed to be nonresponse. The adverse events were analyzed between different VWD types, response, age, gender, and race.

Result

Of those 72 VWD patients, 45 have type 1 (62%), 7 have type 2 (10%), and in 20 cases the subtype was unclear (28%). Responsive to Stimate® was observed in 43 (95.6%) of type 1, 4 (57.1%) of type 2, and 19 (95%) in which the diagnosis was unclear. In total, 16 patients (22.2%) had adverse events: 1 – allegoric response, 8 - mild headache, 4 - mild hyponatremia, and 4 - blood pressure (BP) reduced more than 20 mmHg.

Conclusion

Patients with VWD 1 have higher response rates to Stimate® than patients with VWD 2 in our test. The Stimate® is effective and safe for treatment of VWD.

Disclosures:

Valentino: Baxter, Bayer, Biogen Idec, GTC Biotherapeutics, Inspiration Biopharmaceuticals, Novo Nordisk: Consultancy, Membership on an entity’s Board of Directors or advisory committees.
An Antibody Targeted To The Anionic Region Of Human Protease Activated Receptor 4 Inhibits Thrombosis Without Influencing Bleeding
Authors: Mumaw Michele M. , Fuente Maria de la , Arachiche Amal , Nobel Daniel N. , Nieman Marvin T.
Published: October 2019

Protease activated receptors (PARs) are G-protein coupled receptors which are activated by cleavage of their N-terminus by thrombin. This generates a tethered ligand which is then able to activate the corresponding receptor. Human platelets express PAR1 and PAR4, which both have crucial roles in mediating the response of platelets to injury. Our hypothesis is that PAR4 is an ideal target for new anti-platelet therapies because it is required for stable clot formation and has limited tissue distribution. We have previously determined a region on PAR4 that is required for efficient activation by thrombin. A polyclonal antibody (CAN12) targeted to this region of the PAR4 exodomain does not cross react to PAR1. Initial studies determined that CAN12 is able to block thrombin-induced human platelet aggregation with an IC50 of 10 ng/ml. Control IgG does not inhibit aggregation at 2 mg/ml. In mouse platelets, CAN12 inhibits P-selectin expression and integrin activation. In the Rose-Bengal mouse model of carotid artery thrombosis, CAN12 (1 mg/kg) administered 10 minutes prior to injury was able to completely inhibit the formation of a thrombus in a dose dependent manner. The antibody delayed thrombosis to greater than 90 min; the experiment was terminated at 90 minutes. In contrast, control treatment (2 mg/kg IgG or saline) resulted in complete occlusion at ∼40 minutes. Further, the minimal dose of CAN12 required for complete inhibition of thrombosis (0.5 mg/kg) administered fifteen minutes after injury also delayed thrombosis from ∼50 minutes to ∼80 minutes. This indicates that CAN12 is able to disrupt a thrombus after it has been initiated. Preliminary evidence indicates that CAN12 is able to delay the cleavage of PAR4. Importantly, CAN12 (2 mg/kg) treatment does not increase bleeding time or blood loss in the tail clip assay compared to control IgG (2 mg/kg) treatment. There was also no significant increase in bleeding in the saphenous vein assay. The mice treated with CAN12 (2 mg/kg) had an average bleeding time of 102 seconds for 12 clot formations in 20 minutes compared to the control mice (IgG 2 mg/kg) which had an average bleeding time of 143 seconds for 11 clot formations. These data demonstrate that we are able to inhibit platelet aggregation in vitro and thrombosis in vivo without influencing bleeding time. Overall, these studies provide insight towards the development of new anti-platelet therapies and, specifically, PAR4 as an antiplatelet therapy target.

Disclosures:

No relevant conflicts of interest to declare.
MSC-Derived HSCs Can Reconstitute Hematopoietic Function In Patients With Severe Aplastic Anemia
Authors: Yin James Q , Gao Chunji , Han Bing , Sheng Jianliang
Published: October 2019

Introduction

Naturally-occurring regeneration of cells and tissues is generally involved in four working mechanisms such as directed differentiation, dedifferentiation, trans-differentiation and transdetermination. The better exploring of these mechanisms could be beneficial to develop clinical strategies for regenerative medicine and to reduce the likelihood of immune rejection and relevant complications Recently, “trans-determination” has attracted great controversy, mostly in regards to whether adult stem cells can colonize other tissues after transplantation. More importantly, how to generate large amounts of a particular stem cell type through a transdetermination process remained to be unsolved. Similarly, it is unclear whether mesenchymal stem cells (MSCs) can transdeterminate into hematopoietic stem cells (HSCs).

Methods

Many technologies were used to validate the transdetermination of adipose-derived mesenchymal stem cells (AD-MSCs) into hematopoietic stem cells (HSCs) from different aspects. They include FACS analysis, PCR tests, immunostaining, expansion and repopulating assays, transplantation analysis and others, showing their in vivo and in vitro potentials for long-term self-renewal and differentiation into multi-lineages of blood cells. Moreover, these AD-HSCs can reconstitute hematopoietic function in six patients.

Results

We report firstly here that a huge number of human AD-MSCs that are CD44+,CD29+, CD105+, CD166+,CD133-,CD34- could rapidly transdifferentiate into hematopoietic stem cells (CD49f+/CD133+/CD34+) and their descending blood cells in vitro, after transfected with two small RNAs. The sRNAs were high-effectively delivered into MSCs by a novel peptide means. These adipose-derived HSCs (AD-HSCs) could form different types of hematopoietic colonies as nature-occurring HSCs did. Upon the primary and secondary transplantation into sublethally or lethally irradiated mice, these MSC-HSCs engrafted and differentiated into all hematopoietic lineages such as erythrocytes, lymphocytes, myelocytes and thrombocytes. Furthermore, we demonstrated the first evidence that the transdetermination of MSCs was induced by acetylation of histone proteins and activation of many transcriptional factors. More excitingly, these MSC-derived HSCs can reconstitute hematopoietic function in six patients with severe aplastic anemia.

Conclusion

our findings identify the molecular mechanisms that regulate the directed transdifferentiation of MSCs toward HSCs, create a new source for individual HSC transplantation used for the treatment of blood diseases and cancers, and break the stalemate caused by bone marrow match and graft-versus-host disease.

Disclosures:

No relevant conflicts of interest to declare.