bluuebunny/crossref_metadata_2025_split huggingface dataset:
Outcome of Pediatric Allogeniec Hematopoietic Stem Cell Transplantation for Nonmalignant Disease: A Single Center Experience
Authors: Abosoudah Ibraheem , Lashin Asem , Yassin Fawwaz , Al trabolsi Hassan , Bayoumy Mohamed 
Published: October 2019

Abstract
Abstract 4558

Background:
The aim of this study was to determine the outcome of pediatric allogeneic hematopoietic stem cell transplantation (Allo-HSCT) for nonmalignant disease in our center.

Method:
Data were retrospectively collected for all patients (aged 0–18 years) who received allogeneic HSCT between May, 2005 and December, 2011. Outcomes according to the type of transplant, diagnosis, and transplant-related complications are reported.

Result:
Allo-HSCT was performed in 17 patients, (11 male; 6 female). Bone marrow Failure (7;41%) was the commonest nonmalignant disease followed by B-thalassemia major (4;24%) and others (6;35%). Only HLA-matched family donors were used. Most patients were conditioned with Busulfan/Cyclophosphamide/ATG. GVHD prophylaxis comprised mainly of cyclosporine and methotrexate. The median time to neutrophil engraftment was 20 (9–27) days. Two patients experienced graft failure. Four patients had Grade (I–III) aGVHD and no patient had grade (IV). Four patients (23%) had chronic GVHD. No patient had sinusoidal obstruction syndrome (SOS). Five patients had reactivation of CMV infection, which was treated and resolved in all of them. Only one patient died of sepsis. Overall survival (OS) and event free survival (EFS) were 94%, and 88% respectively.

Conclusion:
In this cohort of patients, bone marrow failure was the main reason for transplantation. The early results of HSCT were promising and consistent with published international data. Similarly, graft failure remains a concern.

Disclosures:
No relevant conflicts of interest to declare.
Absence of Spred, a Negative Regulator of Tyrosine Kinase Activity, in Acute Myeloid Leukemia Patients
Authors: Rosso Valentina , Panuzzo Cristina , Bracco Enrico , Carturan Sonia , Campia Valentina , Favole Alessandra , Signorino Elisabetta , Niparuk Pimjai , Calabrese Chiara , De Luca Luciana , Caivano Antonella , Morano Annalisa , Frassoni Francesco , Saglio Giuseppe , Cilloni Daniela 
Published: October 2019

Abstract
Abstract 4616

Background:
The Sprouty family proteins, including Spred, were originally identified in Drosophila melanogaster as an antagonist of Breathless, the ortholog of Fibroblast Growth Factor Receptor (FGFR) in mammals. These proteins are inducible inhibitors of signalling induced by receptor tyrosine kinases. Their main function is to deregulate the RAS/MAPK and RAS/RAF/ERK signal pathways by physically interacting. The role of Spread proteins in haematological malignancies is still not been clarified.

Aim:
The aim of this study was to investigate a possible involvement of Spred in maintaining the aberrant TK signalling in patients affected by acute myeloid leukaemia (AML).

Methods:
Bone marrow (BM) cells were collected from 82 AML patients at diagnosis and 10 BM samples from healthy donors as control. In addition 20 patients were analyzed during follow-up at the time haematological remission. All the patients included had been characterized at the cytogenetic level by conventional karyotyping, and screened by reverse transcriptase-PCR for the presence of the most frequent fusion transcripts or mutations. We analysed Spread mRNA expression by RQ-PCR and the protein by Western blot and immunofluorescence assay. In addition gain of function experiments were performed by transfecting with Spred coding sequence different leukemic cells lines lacking Spred activity.

Results:
We found that Spread mRNA levels were significantly decreased in AML compared to healthy subjects (2−DDct = 0,1 in AML compared to 0,6 in controls) (p<0,001). Western blot and immunofluorescence confirmed the Spred downregulation at the protein level. Spread expression did not correlate to FAB subtypes or cytogenetic risk. Interestingly, when patients obtained the complete hematological remission (HR) the levels of Spred mRNA and protein were significantly upregulated to a level similar to healthy subjects (p=0.1 between HR and normal controls). Transfection experiments of Kasumi cell line with Spred coding sequence resulted in a decrease of proliferation of 53%, a colony growth reduction of 50% and a significant increase of apoptosis (p<0.001). Finally, in order to establish the molecular basis for Spred1 deregulation we analyzed the expression levels of miRNA-21 which is a known regulator of Spred and we found a correlation between the expression of miRNA-21 and Spred.

Conclusions:
A decreased expression and activity of Spred, a negative regulator of TK activity through Ras pathway, is implicated in sustaining the oncogenetic signalling and abnormal proliferation induced by tyrosine kinase proteins in acute myeloid leukemia patients.

Disclosures:
No relevant conflicts of interest to declare.
Tissue Factor Pathway Inhibitor 2 (TFPI2) Is Commonly Methylated in Multiple Myeloma
Authors: Hatzimichael Eleftheria , Dasoula Aggeliki , Kounnis Valentinos , Katsenos Andreas , Crook Tim , Briasoulis Evangelos 
Published: October 2019

Abstract
Abstract 4617

Background:
Tissue Factor Pathway Inhibitor 2 (TFPI2) is a member of the Kunitz-type serine proteinase inhibitor family that regulates extracellular tissue homeostasis by inhibiting matrix metalloproteinases. Transcriptional silencing of TFPI2 due to DNA methylation of the CpG island of its promoter has been reported in several tumors and has been suggested to facilitate tumor cell invasion and angiogenesis. We investigated the DNA methylation status of TFPI2 in multiple myeloma (MM) patients as part of a program of epigenetic profiling of multiple myeloma.

Methods:
Genomic DNA extracted from two human MM cell lines (U-266 and RPMI-8226) and bone marrow aspirate samples from a well chracterized series of MM patients was modified by sodium bisulphite using the EZ DNA methylation kit, ZymoResearch. Bisulfite modified DNA was used as a template for Methylation Specific PCR with primers specific for unmethylated and methylated alleles. Control methylated (CpG Genome™ Universal Methylated, Chemicon International) and unmethylated genomic DNA was included in each experiment. Chi-square test, logistic regression analysis and unpaired t-test were used to investigate associations between gene methylation and ISS stage, presence of extramedullary disease, bone disease, anemia (hemoglobin ≤10 mg/dL), renal failure, serum albumin and beta (2)-microglobulin level. Kaplan-Meier curves were used to estimate the probabilities of survival and the Log-rank test to assess the statistical significance of differences in event rates.

Results:
Forty six patients with MM (26 male, 20 female; mean age 64) and two human MM cell lines were studied. According to the International Staging System (ISS) 21 patients were classified as stage I, 10 stage II and 15 as stage III disease. Methylation in the CpG island of the promoter of TFPI2 was detected in 30 patients (65%, 95%CI= 52,6–78,7%) and in both cell lines (100%). Patients with methylated TFPI2 had significantly elevated beta 2 microglobulin levels (5553 vs 3787 mg/L mean values, p=0.0002) but we did not detect a statistically significant difference in overall survival among patients with methylated versus non methylated TFPI2. No other relevant correlations were detected.

Conclusion:
TFPI2 is commonly methylated in MM patients. Transcriptional silencing of TFPI2 may play a role in the mechanism of myelomatogenesis and its methylation status warrants further evaluation as a diagnostic co-marker for this disease.

Disclosures:
No relevant conflicts of interest to declare.
Identification of Molecular Mutations Causing Haemophilia B From China: A Single-Center Experience
Authors: Zhou Rong-Fu , Zhang Xian , Ouyang Jian , Yang Yonggong , Shao Xiao-Yan , Xu Jingyan , Zhang QiGuo 
Published: October 2019

Abstract
Abstract 4627

Objective:
To identify F9 gene mutations in patients with hemophilia B registered in Nanjing Drum Tower Hospital Hemophilia Registeration Center. Methods: One stage method was used to detect APTT, PT, TT, Fg and the activities of endogenous coagultation factors. Correction testing was employed to exclud the existence of inhibitor with mixed normal plasma. Genomic DNA was extracted from blood samples of 19 unrelated haemophilia B patients and their traceable family members. All exons and their flanking sequences of the F9 gene were amplificated by PCR and subsequently, the products were purified and sequenced directly. Results: APTT was significantly prolonged for all 19 cases of hemophilia B patients, but could be corrected by mixed normal plasma. According to the serial number, FIX:C was 3.7%, 3.5%, 1.9%, 1.9%, 2.2%, 2.0%, 1.9%, 3.2%, 3.5%, 10.8%, 7.8%, 2.2%, 3.8%, 2.3%, 1.6%, 1.4%, 3.7%, 7.8% and 3.5%, respectively. Thirteen different mutations of F9 gene were identified, including C 20518 T, T 6427 C, C 6460 T, C 31008 G, C 17761 T, A 17759 G, G 30150 A, G 31093 C, T 30930 C, G 20565 A, G 30987 A, A 6473 G and C 9 G, respectively. The mutations were composed of 10 missense mutations, one nonsense mutation, one a donor splice site mutation and one promoter mutation. Among them, mutations sites nt6460, nt17761, nt20518, nt30150 and nt31008 were located in CpG islands, belonging to mutation hot-spots. Mutations including C 9 G, C 31008 G and G 31093 C were firstly reported. Conclusions: No inhibitors are detected in the plasma of all patients. The F9 mutations are heterogenous and the missense mutations are the most prevalent gene defects in Chinese HB patients.

Disclosures:
No relevant conflicts of interest to declare.
Refractory Atypical Hemolytic Uremic Syndrome
Authors: Chao Ju-Hsien , Riley Beth C. , Redman Rebecca 
Published: October 2019

Abstract
Abstract 4635
Atypical hemolytic uremic syndrome is a rare but often lethal disease if not diagnosed and treated early. The mainstay of treatment has been plasma exchange, though eculizumab has recently been approved for treatment.
We report a 24-year-old Caucasian male whom presented from a rehab facility with weakness. Three months prior, he had sustained polytrauma from a motorcycle accident. At discharge, labs were normal. At admission, he was found to have anemia, thrombocytopenia, and elevated creatinine and bilirubin. Further testing revealed evidence of hemolysis. ADAMTS13 level was greater than 100% to assess for thrombotic thrombocytopenic purpura (TTP). Flow cytometry for CD59 was negative to assess for paroxysmal nocturnal hemoglobinuria (PNH). Shiga toxin was negative. He was diagnosed with atypical hemolytic uremia syndrome (aHUS). Patient was initially treated with steroids then underwent daily plasma exchange for 2 weeks with improvements in lab parameters and no evidence of hemolysis. Exchanges were changed to every other day for 2 weeks with plans to start eculizumab as outpatient therapy. After discharge, one dose of eculizumab 900 mg was given without further exchanges. Three days later, lab check showed decrease in hemoglobin and platelets, concerning for aHUS exacerbation. Patient was admitted for daily plasma exchange with eculizumab 100 mg boost after each procedure for about 2 weeks. Once his lab parameters were consistent with resolution, exchanges were spaced out to every other day for another 1 week then discontinued. Then he was started on monotherapy with eculizumab 900 mg once weekly with surveillance lab monitoring. He had no further exacerbation of his aHUS and was discharged back to rehab with weekly eculizumab and lab surveillance.
Early detection and treatment of aHUS is important to minimize the damages from the underlying thrombosis and prevent disease related mortality. Plasma exchange remains the standard treatment for this condition with close monitoring of lab parameters, though no set treatment schedule has been established. The recent approval of eculizumab offers another option in the management of this disease and appears equally efficacious.

Disclosures:
No relevant conflicts of interest to declare.
Treatment of Congenital Fibrinogen Deficiency; Global Development Plan for a Double Virus Inactivated Fibrinogen Concentrate
Authors: Schwartz Bruce A. , Knaub Sigurd 
Published: October 2019

Abstract
Abstract 4636
Congenital afibrinogenaemia and hypofibrinogenaemia are rare inherited disorders occurring in homozygotic patients with an estimated incidence of 1 in 106. Patients present with frequent severe bleeding episodes since birth or early childhood. Bleeding may occur after a minor trauma or a small surgical intervention, into the skin, mucosa, muscles, gastrointestinal tract, or the brain. Therapeutic substitution with human fibrinogen concentrate can correct the haemostatic defect and arrest the bleeding in patients with these fibrinogen deficiencies. Octafibrin is a highly purified, lyophilized, human plasma fibrinogen concentrate, without added albumin. Octafibrin is double virus inactivated using 2 dedicated virus inactivation/removal steps, solvent/detergent treatment, and nanofiltration. A plan for the global development of Octafibrin has been prepared taking into account discussions with European Regulators and the FDA. This plan also involves discussions with the European pediatric committee (PDCO) which oversees the inclusion of pediatric subjects into drug development under the new EMA guidelines.
The development plan calls for a prospective, randomized, open label, multinational, pivotal PK comparison of Octafibrin to an existing marketed product in 18 adult and adolescent patients, including comparison of a surrogate efficacy endpoint measured by TEG. In a second study the efficacy and safety of the product in bleeding and invasive procedures will be assessed in 24 adult and adolescent patients. Finally a pediatric PK, efficacy and safety study in patients below 6 years will be performed but because of the rarity of these patients this study will not need to be completed before review and approval by the regulatory agencies. Clinical studies are planned to be started later this year.
A double virus inactivated, plasma derived fibrinogen concentrate (Octafibrin) will be globally developed in an ultra rare congenital disease after harmonized discussions with EU and US regulators. Pivotal comparative PK data and interim efficacy and safety data will be available at time of regulatory submissions while the finalization of the pediatric study is deferred.
Effective management of congenital fibrinogen deficiencies in bleeding situations is necessary for the prevention of potentially life-threatening bleeding episodes. This clinical program will help to confirm that fibrinogen substitution is able to successfully control bleeding, increase the fibrinogen plasma levels, and reduce the amount of transfusions needed with allogeneic blood products. In addition, it will give additional information on the tolerability and overall safety profile of fibrinogen replacement therapy.

Disclosures:
Schwartz: Octapharma: Employment. Knaub:Octapharma: Employment.
Acquired Hemophilia After a Spider Bite
Authors: Vinayek Namita , Kloecker Goetz H , Riley Beth C. 
Published: October 2019

Abstract
Abstract 4637
A 59 year old white female with no history of bleeding diathesis presented with swelling of her left forearm two days after a brown recluse spider bite. Hematology was consulted for uncontrolled bleeding from the fasciotomy site after compartment syndrome. The hand surgeons did not see any vascular abnormality at the site of surgery. Due to ongoing bleeding for days from the surgical site, she required 8 PRBCs, 6 FFPs and 6 apheresis platelets units. PT, PTT, and platelet count were normal, ristocetin cofactor was 339% and vWF antigen 306% and vW Factor VIII binding capacity was normal. Her platelet functional assay was normal. Factor levels of II, V, VII, XI, X, XI and lupus anticoagulant were all with in normal range. A Bethesda assay however showed a low titer of antibody against factor VIII (1.25 BU). Factor VIII was low at 6% and remained low at 16% with no significant response to recombinant factor VIII.The bleeding however responded well to recombinant factor VIIa. She was maintained on high dose steroids for few months and tapered. She eventually had a skin graft to the site of injury and is now followed in our hemophilia clinic with no further bleeding complications.
To our knowledge only one case of spider bite resulting in factor VIII inhibitor has been documented in the literature. We hypothesize that direct antigenic stimulation by spider venom can transiently develop factor VIII inhibitor in individuals.

Disclosures:
No relevant conflicts of interest to declare.
Quantification of Immature Platelets with Abbott CD-Sapphire and Sysmex XE-5000 in Patients with Haematological and Malignant Diseases
Authors: Meintker Lisa , Haimerl Maria , Fritsch Juliana , Ringwald Jürgen , Krause Stefan W. 
Published: October 2019

Abstract
Abstract 4639

Background:
Quantification of immature, i.e. RNA-containing platelets (PLT) was introduced into routine diagnostics by Sysmex as immature platelet fraction (IPF) some years ago and recently by Abbott as reticulated platelets (rPLT). In this study, we evaluated the precision and agreement of both analyzers in normal and pathological samples and correlation of IPF or rPLT with clinical situations.

Methods:
Anonymized blood samples of cancer patients after chemotherapy, patients with PLT destruction (immune thrombocytopenia, ITP, or heparin-induced thrombocytopenia, HIT), and normal controls were analyzed with double measurements with both, the Abbott CD-Sapphire and Sysmex XE-5000. Furthermore, longitudinal evaluations after chemotherapy were performed and effects of PLT transfusions were analyzed. Precision and agreement were assessed with Bland-Altman analyses and Pearson correlation.

Results:
In a first set of experiments, a total of 220 blood samples were analyzed. We studied patients after chemotherapy with beginning thrombocytopenia (30 samples, median PLT count 23/nl) or recovering PLT counts (64 samples, PLT 48/nl), patients with PLT destruction (16 samples, PLT 84/nl) and normal controls (110 samples, PLT 249/nl). Both analyzers demonstrated acceptable overall precision (reproducibility) with lower precision in thrombocytopenia and a non-significant trend towards a better precision with XE-5000. Inter-instrument comparison showed a moderate correlation (Pearson r2=0.38) and a systematic bias of 1.04 towards higher IPF-values with the XE-5000 in the Bland-Altman analysis. Regarding different patient groups, the reference range (percentile 5 – 95) of normal controls was much narrower with the CD-Sapphire, leading to a clear separation of patients with recovering platelet counts after chemotherapy and IPT/HIT, both with increased rPLT as expected (figure). The same trend was visible for the XE-5000, however, with a much broader overlap to the normal range. In longitudinal studies, increased IPF or rPLT were frequently observed already during early thrombocytopenia with a decrease following platelet transfusions.

Conclusion:
Correlation between both analyzers for measurement of IPF and rPLT, respectively, was moderate. Precision of measurements was low in thrombocytopenic samples. Samples with abnormally increased IPF/rPLT could be indentified with both analyzers, however, separation from normal was better with the CD-Sapphire. Prediction of PLT recovery after intensive chemotherapy by the measurement of IPF/rPLT is questionable.

Disclosures:
Krause: Beckman Coulter: Research Funding.
Hyperbaric Oxygen Therapy Improves Post-Transplant Umbilical Cord Blood Engraftment
Authors: Aljitawi Omar S. , Yinghua Xiao , Lin Tara , Eskew Jeff , Parelkar Nikhil , Swink Megan , Radel Jeff , McGuirk Joseph , Li Linheng , Vielhauer George 
Published: October 2019

Abstract
Abstract 4663

Background:
Delayed engraftment following umbilical cord blood (UCB) transplantation is associated with increased transplant-related mortality. Earlier, we have reported that hyperbaric oxygen therapy (HBO) might have altered early UCB engraftment kinetics in a transplant animal model. Herein, we report use of HBO to improve long-term UCB engraftment.

Methods:
Six-to eight-week old NSG mice were sublethally irradiated 24 hours prior to CD34 selected UCB cell transplant. The irradiated mice were separated into a control group (where mice remained under normoxic conditions) and the HBO group (where mice received two hours of HBO therapy; 100% oxygen at 2.5 atmospheres absolute). Six hours after starting HBO therapy, both groups received 1×105 CD34 selected UCB cells via tail vein injection. The infused CD34 selected cells were transduced with a lentivirus carrying luciferase gene for in vivo imaging. Mice (n=4) from each treatment group were sacrificed after 3 and 4 months to assess long-term engraftment. Engraftment was measured as assessed by flow cytometry. Myeloid, B-cell, and T-cell subset engraftment was determined by flow cytometry.

Results:
At 3 and 4 months post-transplant, the mean percentage of human CD45 expressing cells in peripheral blood of HBO mice was significantly higher than controls (p=0.03 and p=.008 respectively). The mean percentage of human CD45 expression was higher in bone marrow of HBO mice at 3 months and 4 months, but only reached statistical significance at 4-month time point (p=0.04). At 3 months, the mean percentage of human CD19 expressing cells was significantly higher in peripheral blood of HBO mice (p=0.03) while no differences were noted between the two groups in terms of CD33 or CD3 expression. At 4 months, the mean percentage of human CD19 expressing cells and human CD3 expressing cells was significantly higher in peripheral blood of HBO mice (p=0.009 and p=0.0009, respectively) while no differences were noted between the two groups in terms of CD33 expression.

Conclusions:
HBO therapy given prior to UCB cell infusion significantly improved UCB's long-term engraftment. At 3 months, engraftment was skewed toward B-cell engraftment, while more balanced engraftment was noted at 4 months. Future studies are planned to confirm these findings and to determine the underlying mechanisms by which HBO improved UCB CD34 cell engraftment.

Disclosures:
No relevant conflicts of interest to declare.