bluuebunny/crossref_metadata_2025_split huggingface dataset:
Eradication of Minimal Residual Disease with Alemtuzumab in Chronic Lymphocytic Leukemia Is Associated with Prolonged Survival and Is an Appropriate Theraputic Endpoint for Relapsed CLL.
Authors: Sayala Hazem A. , Moreton Paul , Kennedy Ben , Lucas Guy , Leach Michael , Rassam Saad M.B. , Haynes Andrew , Tighe Jane , Oscier David G. , Fegan Christopher , Rawstron Andy C. , Hillmen Peter 
Published: October 2019

Abstract
Eradication of minimal residual disease (MRD) in chronic lymphocytic leukemia (CLL) is emerging as a desirable therapeutic end point predicting for better outcome. The monoclonal antibody alemtuzumab (Mabcampath) is approved for patients with fludarabine refractory CLL. We previously published 91 patients with relapsed CLL (74 men and 17 women, median age 58 years [range, 32 to 75 years]; 44 fludarabine-refractory) who received a median of 9 weeks (range 1 to 16) of alemtuzumab, 30mg 3x a week after dose escalation, between 1996 and 2003. 84 patients had i.v. alemtuzumab and 7 received it subcutaneously. Responses to alemtuzumab according to NCI-WG criteria were complete remission (CR) in 32 patients (36%), partial remission (PR) in 17 (19%) and no response (NR) in 42 (46%). Detectable CLL to a level of less than one CLL cell in 10,000 leucocytes, assessed by four-color MRD flow cytometry, was eradicated from the blood and marrow in 18 patients (20%). 8 of these 18 patients were fludarabine refractory. We report here the results of long term follow up of this cohort of patients after a median follow up of 77 months (range 5 to 123 months). Median survival was significantly longer in patients achieving MRD negative responses compared with those with detectable CLL at the end of therapy. The median survival for all 18 MRD negative responders has not been reached but was 87 months for the 8 fludarabine-refractory patients achieving MRD negativity. Overall survival for the 18 patients with MRD-negative remissions was 66% at 72 months (see Figure). MRD positive CR patients had a median survival of 56 months, MRD positive PR patients a median survival of 42 months and non-responders a median survival of 14 months. The median treatment-free interval prior to alemtuzumab for the 18 MRD negative patients was 8 months (range 4 to 35). Excluding planned stem cell transplantation performed in CR, the median time to next treatment for the 18 MRD negative patients was 114 months and 72% (13/18) have required no further therapy. Therefore alemtuzumab can induce MRD negative remissions in CLL resulting in a clear survival advantage with 66% of MRD negative patients alive 6 years after alemtuzumab. The markedly increased treatment-free survival and excellent survival for MRD negative patients strongly suggests that achieving an MRD negative remission is an appropriate therapeutic end-point in relapsed CLL.
Figure Figure
AT7519, a Potent CDK Inhibitor, Is Active in Leukemia Models and Primary CLL Patient Samples.
Authors: Squires Matthew S. , Feltell Ruth E. , Lock Victoria , Smith Donna M. , Lewis Jon E. , Higgins Jacqueline , Yule Murray , Thompson Neil T. , Cooke Laurence , Croce Kimiko D. , Qi Wenqing , Lyons John F. , Mahadevan Daruka 
Published: October 2019

Abstract
Cyclin Dependent Kinases (CDKs) play a central role in the eukaryotic cell cycle. The activation of these kinases is modulated by the expression and binding of their regulatory cyclin partners. Their key role in cell cycle progression, coupled to evidence that pathways leading to their activation are deregulated in a number of human cancers makes them attractive therapeutic targets. More recently the role of CDKs 7, 8 and 9 in the regulation of transcription has been explored. CDK9 has been shown to play a role in the regulation of transcription via phosphorylation of RNA polymerase II. The outcome of transcriptional inhibition via CDK9 exhibits significant variation between cell lines. Many leukemic cell lines, which are dependent upon the expression of short half-life transcripts such as Mcl-1 for survival, undergo apoptosis following transcriptional inhibition and the cell cycle effects of such inhibitors are masked. AT7519 is a potent inhibitor of cyclin dependent kinases 1, 2 and 9 and is currently in early phase clinical development. These studies profile the mechanism of action of AT7519 in leukemia cell lines (HL60 and MOLT-4). AT7519 causes rapid induction of apoptosis (within 6 hours of initial exposure) in the absence of cell cycle arrest. This induction of apoptosis occurs in parallel with a reduction in the levels of anti-apoptotic proteins such as Mcl-1. In HL60 xenograft models anti-tumour efficacy is observed following 2 cycles of daily dosing of 15 or 10mg/kg for 5 days followed by 2 days off treatment. We observe a 50% cure rate (4/8 mice) at the 15mg/kg dose and a 30% cure rate at the 10mg/kg dose level 40 days following dosing. Pharmacodynamic biomarker studies demonstrate that a greater extent and duration of Mcl-1 knockdown and apoptosis induction are associated with efficacious doses. B-Cell lymphoproliferative disorders, including CLL, rely on the expression of transcripts with a short half-life such as Mcl-1, Bcl-2 and XIAP for survival. In vitro studies have demonstrated that compounds with transcriptional inhibitory effects are effective pro-apoptotic agents in models of this disease. Therefore, we also characterised the transcriptional effects of AT7519 on CLL cells isolated from patients. AT7519 was shown to induce apoptosis (by MTS, morphology and PARP cleavage) in these samples at concentrations of 100–300nM. These effects were correlated with the cytogenetic background of individual patients and the data supports further clinical investigation of AT7519 in B-Cell lymphoproliferative disorders where survival proteins play a pivotal role.
VWF Concentrates - Does Size Really Matter - Von Willebrand Factor Function under Physiological Flow Conditions.
Authors: Schwartz Bruce A. , Fuchs Birte , Fisseau C. , Kessler Craig M. , Kannicht Christoph 
Published: October 2019

Abstract
Introduction: The Von Willebrand Factor (VWF), is a very large glycoprotein present in human plasma essential for normal thrombus formation at sites of vascular injury. Blood coagulation is initiated by exposure of vessel wall components, e.g. collagen, followed by platelet tethering, activation and adhesion leading to the formation of a stable clot. All steps of blood coagulation occur in flowing blood under various conditions depending on vessel size. It is important to assess the function of a VWF concentrate under physiologic conditions. Under static or low shear conditions, platelets can bind directly to collagen without assistance of VWF, while the VWF is essential for mediation of platelet adhesion under high flow occurring in the arterial circulation. It has been assumed that multimer size plays an important role in this binding and in subsequent platelet adhesion. In this study we evaluated the binding of VWF of different multimeric structures to collagen and determined VWF-mediated platelet binding under low to high shear rates in a flow chamber model.
Methods: A flow-chamber coated with human collagen was developed to mimic physiological flow conditions. A high purity VWF/factor VIII (FVIII) concentrate (Wilate®) and two other VWF/FVIII concentrates were tested at shear rates of 400 1700 and 4000 s−1 reproducing shear rates occurring in veins, small arteries and capillaries. Collagen-bound VWF was characterized by antigen determination (VWF:Ag) and multimer (MM) analysis. Binding of labeled platelets was visualized by a fluorescence microscope and surface coverage was quantified.
Results: All VWF MMs independent of MM size were found to bind to collagen even under high shear rates. The amount of collagen-bound VWF:Ag and VWF-mediated platelet adhesion at 1700 s−1 differed significantly between the tested concentrates, when equal VWF:Ag amounts were applied.
Conclusion: Binding of VWF to collagen does not depend on VWF MM size in this model even when measured under high shear rates. The differences in collagen-bound VWF:Ag and VWF-mediated platelet binding do not seem to depend on the VWF MM distribution of the concentrates. Other structural features than VWF MM size may likely be caused by the differences seen in the binding levels of the different concentrates and should be further explored. Possible differences between VWF activity assays performed under static or flow conditions and their appropriate use for VWD diagnosis and quantification of in-vivo activity need to be further investigated.
Microparticle Levels Are High in Patients with Hemophilia A and Can Be Further Increased by DDAVP Administration.
Authors: Artoni Andrea , Santagostino Elena , Mancuso Maria E. , Lecchi Anna , Mannucci Pier M. 
Published: October 2019

Abstract
Microparticles (MP) are circulating submicroscopic cell fragments expressing procoagulant phospholipids and haemostatic proteins as tissue factor. Several evidences support a pro-haemostatic role of MP and suggest that the axis VWF-platelet GpIb is critical in their generation. The aim of this study was to measure the levels of platelet-derived MP (PMP) and tissue factor-expressing MP (TFMP) in patients with hemophilia A to evaluate whether or not a correlation with the phenotype exists. The levels of PMP and TFMP were also measured after DDAVP administration in a subset of patients with mild hemophilia A to assess whether MP levels change in response to DDAVP. FVIII:C, VWF:Ag, PMP and TFMP were measured in plasma 30 min, 1, 2, 4, 8 and 24 h after DDAVP 0.3 μg/kg, subcutaneously. PMP and TFMP were analysed in the pellet of platelet poor plasma by flow cytometry and defined as Annexin V and GpIIb/IIIa or Tissue Factor positive events falling in a gate defined by 1 μm beads. MP were enumerated adding a known number of 7 μm beads to each test tube. PMP and TFMP were measured in 50 patients with severe (FVIII:C<1%) and in 37 with mild (FVIII:C 7–34%) hemophilia A after a wash-out period >72 h, and in 20 male controls. Levels of both PMP and TFMP were lower in controls than in all hemophiliacs (p<0.05), severe hemophiliacs (p<0.05) and mild hemophiliacs (p<0.05). No correlation was found between levels of MP and FVIII:C in mild hemophiliacs. A significant increase in the levels of PMP and TFMP (p<0.05 at all time points vs baseline) was observed in 9 patients after DDAVP administration; peak levels were achieved after 2–8 h. Correlations were found between the increase of VWF:Ag and FVIII:C levels and the peak levels of MP (r=0.62 and r=0.7, respectively). This study showing higher MP levels in hemophiliacs than in controls suggests a role of MP in this patient population. Furthermore, increased MP levels in response to DDAVP administration support the hypothesis that VWF of ultralarge size released by the endothelium interacts with platelets generating MP.
All hemophiliacs (#87) Severe (#50) Mild (#37) After DDAVP (#9) Controls (#20) Median PMP/ml (range) 139 (25–709) 150 (12–586) 128 (28–709) 463 (179–3442) 57 (31–288) Median TFMP/ml (range) 185 (36–859) 169 (36–718) 188 (93–859) 240 (91–1571) 143 (55–314)
Prolonged In-Vivo Half-Life of FVIIa by Fusion to Albumin.
Authors: Schulte Stefan , Weimer Thomas , Wormsbaecher Wilfried , Kronthaler Ulrich , Groener Albrecht , Lang Wiegand , Liebing Uwe 
Published: October 2019

Abstract
For the treatment of hemophilia patients with inhibitors, recombinant Factor VIIa (rFVIIa) is available as a therapeutic option to control bleeding episodes with a good balance of safety and efficacy. The short in-vivo half-life of approximately 2.5 h requires multiple injections, which is inconvenient for treaters and patients. Here we describe the generation of a half-life extended recombinant FVIIa molecule based on genetic fusion of FVIIa to human albumin. In this fusion protein the design of the linker sequence is important to optimize the effect of the albumin moiety on FVII activity. The recombinant FVII-albumin fusion protein (rVII-FP) was expressed in mammalian cells and upon activation displayed a FVII activity comparable to wild type rFVIIa. Pharmacokinetic studies in rats and rabbits demonstrated that the half-life of the activated recombinant FVII albumin fusion protein (rVIIa-FP) was 6 to 9 fold extended compared to wild type rFVIIa. The in-vitro and in-vivo efficacy was evaluated and found comparable to commercially available rFVIIa (NovoSeven®). The results of this study demonstrate that it is feasible to improve the attributes of a rVIIa molecule by extending its half life, while retaining a molecule with very similar hemostatic properties to the wild type factor.
Investigating Mature B-Cell Lymphomas by MicroRNA Profiling.
Authors: Basso Katia , Morozov Pavel , Kitagawa Yukiko , Maute Roy L. , Rosolen Angelo , Chen Chang-Zheng , Califano Andrea , Dalla-Favera Riccardo 
Published: October 2019

Abstract
In order to understand the role of microRNA (miRNAs) in mature B cell function and lymphomagenesis, we analyzed the miRNA expression profile of B cells at different stages of differentiation by cloning of short RNA and by array-based miRNA expression studies. We generated libraries representative of miRNAs expressed in normal germinal center (GC), naïve and memory B cells isolated from human tonsils and in a Burkitt lymphoma cell line (Ramos). Short RNAs were gel purified and linked to adaptor oligonucleotides, reverse transcribed, PCR amplified, cloned into an expression vector, and subjected to sequencing. Candidate miRNAs have been aligned to the human genome to retrieve their potential precursor sequences, whose folding characteristics have been analyzed in order to identify bona fide precursors. The cloned and structurally validated miRNA precursors have been matched to the public miRNA database (miRBase v10.0) to detect previously identified miRNA sequences. The results show that 201 (109 previously known and 92 newly discovered) mature miRNAs were cloned from normal B cells and Ramos cell line. Overall, this analysis led to the discovery of 92 not previously reported miRNAs and allowed us to identify the complete miRNA expression profile of different stages of B cell development. The cloning data show a good degree of concordance with the array-based analysis leading to the identification of a distinct miRNA expression profile for each normal B cell population. Naïve and memory B cells appear to share a large fraction of their miRNA profile while centroblasts display a clearly unique pattern of miRNA expression. We performed a comprehensive analysis of miRNA expression profiles of GC-derived lymphomas (Burkitt lymphoma, diffuse large B cell lymphoma, follicular lymphoma) using the same microarray platform including approximately 500 miRNA sequences. Each tumor type shows a distinct profile that separates them from their normal counterpart. Interestingly, initial results indicate that a set of miRNAs is expressed in normal GC cells, but not in lymphoma samples and cell lines, suggesting that structural and/or functional alterations of miRNAs occur during lymphomagenesis.
Risk Prediction of Heparin Induced Thrombocytopenia and Thrombosis in the Tertiary Care Setting.
Authors: Jooma Nuruddin , Kabir Azad , Allison Paul M. , Bracey Arthur 
Published: October 2019

Abstract
Background: Heparin induced thrombocytopenia (HIT) is a challenging diagnosis; especially post cardiac surgery and patients with cardiac support devices. The diagnosis is based on clinicopathologic findings including pertinent clinical history, and heavy reliance on laboratory testing (Heparin PF-4 ELISA) to detect antibodies immunologically or by functional means (serotonin release assay - SRA). Experience with routine measurement of SRA in combination with HPF4 in this setting is limited.
Aim: To determine the clinical and laboratory findings most predictive of thrombosis in patients with HIT.
Patients and Methods: This is a retrospective analysis where we reviewed data on patients who had a positive heparin antibody thrombocytopenia from 1/1/06 to 5/1/07 at the Texas Heart Institute in Houston. We evaluated their diagnosis, O.D., SRA, Doppler ultrasound and platelet count.
Results: Our study found 52 (12.5%) out of 416 patients with positive heparin antibody. Thirty-two patients (62%) with positive heparin antibody had undergone a cardiac procedure within last 30 days. Sixteen (31%) out of the 52 patients with positive heparin antibody had thrombosis confirmed by Doppler/Ultrasound study. Of those 16 patients, 12 (80%) had undergone a cardiac procedure and 10 (63%) had their platelets drop below 75,000. Mean O.D. in patients with thrombosis was 1.10 versus 0.82 for patients with no thrombosis (p=0.14). Serotonin Release Assay (SRA) was considered positive if the serotonin release was > or = 20%. It was positive in only 5 (10%) patients, however, 3 out of those 5 patients had thrombosis. Mean O.D. for patients with positive SRA were 1.88 as compared to O.D. of 0.81 (P<0.001) in patients without positive SRA. All 5 patients had undergone a cardiac procedure.
Conclusion: Our study found almost one third of patients with positive HPF4 antibody developed thrombosis. Patients with thrombosis were more likely to have a positive SRA, higher O.D. and more likely to have undergone a cardiac procedure. Prospective studies are planned to further delineate the relationship between O.D., SRA and thrombosis in the setting of cardiac procedures.
Exploring Genetic Basis of GVHD by Whole-Genome Association Studies in a Large Series from the Japan Marrow Donation Program (JMDP).
Authors: Nannya Yasuhito , Onizuka Makoto , Kashiwase Koichi , Sanada Masashi , Akatsuka Yoshiki , Satake Masahiro , Chiba Shigeru , Kurokawa Mineo , Yamamoto Ken , Saji Hiroo , Maruya Etsuko , Inoko Hidetoshi , Morishima Yasuo , Kodera Yoshihisa , Ogawa Seishi 
Published: October 2019

Abstract
Allogeneic stem cell transplantation (allo-HSCT) has been established as one of the most effective therapeutic options for hematopoietic malignancies. Its major therapeutic benefits are obtained from allo-immunity directed against patients’ tumor cells (graft-versus-tumor effects, GVT). However, the same kind of allo-immune reactions can also be directed to normal host tissues resulting in graft-versus-host disease (GVHD) and subsequently offset the potential benefit from GVT. Because both reactions are initiated by donor T-cells that recognize minor histocompatibility antigens (mHA) that are incompatible between donor and recipient, the knowledge of these mHA is of particular importance to develop a new measure to prevent GVHD while harnessing the antitumor potential of allo-HSCT. On the other hand, the incompatibility of mHAs, as well as other predispositions to development of GVHD, is basically determined genetically, and thus, are plausible targets of genetic association studies, which have recently been enabled with the advent of large-scale genotyping technology. In the current study, we explored genetic basis of GVHD by whole genome association studies, in which relevant SNPs to GVHD development were explored by genotyping a large cohort of GVHD-positive and negative transplantations performed through the Japan Marrow Donor Program (JMDP). Among 7,800 transplants genotyped for HLA types, we selected 1856 transplants (3712 donors/recipients) for large scale genotyping of more than 500,000 SNPs using Affymetrix® GeneChip® 500K arrays, according to the following criteria:
HLA A, B, C, DR, DQ loci were completely matched at the DNA level Short term MTX (stMTX) + CyA or stMTX + FK506 was employed as GVHD prophylaxis, and Genome DNA from both donor and recipient and record of clinical course was available.
Based on the genotyping data, possible association was tested for each SNP in donors and recipients with regard to the development of GVHD. Association of genotype incompatibilities was also investigated for major HLA subtypes to identify the mHA loci that are relevant to GVHD pathogenesis. In the intermediate analysis, a number of candidate polymorphisms in donors and recipients have been identified together with possible mHA loci.
Selective Depletion of Alloreactive T Cells and Study of Anti-Tumor Activity of Specific T Cell Clones in Patients with Leukemia and Renal Carcinoma.
Authors: Matejkova Eva , Buresova Ivana , Vidlakova Petra , Kyjovska Drahomira , Vitetta Ellen S. , Michalek Jaroslav 
Published: October 2019

Abstract
A major challenge in the field of hematopoietic stem cell transplantation (HSCT) is to prevent the alloreactivity of donor T-cells which leads to acute graft-versus-host disease (GVHD) while preserving a graft-versus-tumor (GVT) effect. Selective depletion using anti-CD25 immunotoxin (IT) can eliminate harmful alloreactive T-cells while preserving other donor T-cells with antileukemic/antitumor reactivity. We have used irradiated peripheral blood mononuclear cells (PBMC) from cancer patients and healthy donor PBMC as responder cells in primary mixed leukocyte reaction (MLR).To prepare GVL/GVT-specific T-cells, alloreactive T-cells in primary MLR were depleted with anti-CD25 IT. The remaining T-cells had insignificant alloreactivity in secondary MLR. Allodepleted donor cells were then repeatedly stimulated using purified leukemia/tumor cells from the same cancer patient. Leukemia/tumor-reactive donor T-cells were purified by immunomagnetic separation on the basis of INF-g production. 23 MLRs (12 with acute myeloid leukemia cells, 3 with acute lymphocytic leukemia cells and 8 with renal carcinoma cells) were performed. Selective depletion of alloreactive donor T-cells with anti-CD25 IT led to more than 2log depletion (99.2–100%, median 99.7%). Graft-versus-leukemia (GVL) effect of donor T-cells was well preserved (4.7% of donor T cells were GVL-reactive) while the graft-versus-host (GVH) reactivation of donor cells was negligible even after repeated stimulation with patient’s non-leukemic PBMC. In the case of renal cell carcinoma, GVT effect was less dominant (1.8% of donor T cells were GVT-reactive clones) and GVH reactivation appeared in 2.4% of donor T cells. In conclusion, it is possible to selectively deplete donor alloreactive T-cells with anti-CD25 IT. In case of patients with leukemia, a strong GVL reactivity was noticed in allodepleted donor T cells while in case of renal cell carcinoma the GVT effect was less dominant and associated with mild GVH-reactivation.