bluuebunny/crossref_metadata_2025 huggingface dataset:
An Open-Label Study of Pegylated Liposomal Doxorubicin, Vincristine and Reduced-Dose Dexamethasone Combination Therapy in Newly Diagnosed Multiple Myeloma Patients in Chinese Population.
Authors: Shen Yang , Shen Zhixiang , Jiang Bin , Hou Jian , Zhan Rong , Qiu Lugui , Wu Yongji , Jin Jie , Li Juan , Meng Fanyi , Zou Ping , Liu Ting , Li Jianyong , Wang Chun , Wu Depai , Ma Jun
Published: October 2019

BACKGROUND: Pegylated liposomal doxorubicin (CAELYX®) is a liposomal formulation of doxorubicin sterically stabilized by the grafting of segments of polyethylene glycol (PEG) onto the liposomal surface. Given the demonstrated efficacy of VAD (vincristine and doxorubicin and oral dexamethasone) in Multiple Myeloma (MM) patients and the potential for CAELYX® to extend the duration of bone marrow exposure to therapeutic levels of doxorubicin, a combination regimen of CAELYX®, vincristine, and reduced-dose dexamethasone (DVD) has been actively investigated in MM patients. Studies showed that substituting CAELYX® for doxorubicin in the VAD regimen and reducing the dose of dexamethasone in MM patients improves the safety profile and convenience of the treatment regimen without compromising efficacy. Due to potential differences in metabolism of these patients, safety and efficacy results may vary. Thus, we carried out this study in 82 newly diagnosed MM patients in China, in order to demonstrate the efficacy and safety profiles of DVD.

METHODS: Patients (n=82) from 15 sites were recruited in this study. CAELYX® (40mg/m2) was infused intravenously over 60-minutes, administered every 28 days. Vincristine (2.0mg) was administered intravenously on Day 1 of each cycle. Dexamethasone (40 mg) was administered from Day 1- Day 4 of each cycle orally or intravenously. The treatment was repeated every 28 days for 4 cycles.

RESULTS: Upon ITT analysis, the overall response rate was approximately 68% (56/82); 11% of the patients achieved complete remission (CR), 40% achieved partial response (PR), 17% achieved minimal response; 15% had stable disease (SD), and 12% o had progressive disease (PD) after the treatment. The cumulative 4-month progression-free survival (PFS) was 88%. The incidence of all the adverse events was 46%. The most common non-hematological toxicities were palmar-plantar erythrodysesthesia (13.4%) and stomatitis (6.1%), respectively.

CONCLUSION: Pegylated liposomal doxorubicin, vincristine and reduced dose dexamethasone combination (DVD) regimen is an effective and safe regimen in newly diagnosed multiple myeloma patients in Chinese population.
Rb Intrinsically Promotes Erythropoiesis by Coupling Cell Cycle Exit with Mitochondrial Biogenesis.
Authors: Sankaran Vijay G. , Walkley Carl R. , Spiegelman Bruce M. , Orkin Stuart H.
Published: October 2019

Regulation of the cell cycle is intimately linked to erythroid differentiation, yet how these processes are coupled at the molecular level is not well understood. In order to gain insight into this coordinate regulation, we examined the role that the retinoblastoma gene product (Rb), a central regulator of the cell cycle, plays in erythropoiesis. A conditional (floxed) Rb gene was specifically deleted in the erythroid compartment by means of EpoR-GFPcre, or in other hematopoietic compartments (with LysM-Cre and Mx1-Cre) in mice. We found that Rb serves a cell-type intrinsic and autonomous role in erythropoiesis. Deletion of Rb in the erythroid compartment results in an anemia due to ineffective erythropoiesis. A partial block in differentiation occurs at the transition from early to late erythroblasts. Unexpectedly, we found that in addition to a failure to properly exit the cell cycle, mitochondrial biogenesis fails to be upregulated at this transition point. The disturbance in mitochondrial activities was established through gene expression analysis, measures of respiratory function, as well as measurements of mtDNA content in sorted cell populations. The failure to upregulate mitochondrial biogenesis appears to contribute to the block in differentiation. The link between erythroid differentiation and mitochondrial function was validated by chemical inhibition of mitochondrial biogenesis and knockdown of the mitochondrial transcriptional coactivator PGC-1beta in cultured erythroid cells. The ineffective erythropoiesis seen in erythroid cells lacking Rb closely resembles the features of the refractory anemia seen in the context of the myelodysplastic syndromes (MDS). Prior work has demonstrated the presence of defects in cell cycle regulation as well as mitochondrial function in MDS. Our work demonstrates how these seemingly disparate pathways may play a role in coordinately regulating cellular differentiation.
Results of a Prospective Phase II Trial of Limited and Extended Rituximab Treatment in Nodular Lymphocyte Predominant Hodgkin’s Disease (NLPHD).
Authors: Horning Sandra J. , Bartlett Nancy L. , Breslin Sheila , Advani Ranjana H. , Hoppe Richard T. , Ekstrand Bradley C. , Lucas Jennifer B.
Published: October 2019

Background: NLPHD represents a unique clinical entity characterized by limited peripheral nodal disease, an indolent behavior with a frequent relapse pattern following radiotherapy or chemotherapy, and late treatment-related effects. Distinct from classical HD, the malignant cells of NLPHD are universally CD20+ and we therefore postulated that rituximab may have activity with fewer adverse effects.

Methods: In this prospective phase II trial, patients (pt) with untreated or relapsed CD20+ NLPHD and measurable disease were eligible to receive four weekly doses of rituximab at 375 mg/m2. Restaging studies after completion of all treatment were performed at 1, 3 and 6 months (mo), and every 6 mo thereafter to progression. Endpoints were estimated response rate, freedom from progression (FFP) and safety. We previously reported that FFP was less than 1 year despite the fact that all patients responded to a limited, weekly × 4 treatment (Ekstrand et al. Blood101:4285, 2003). The protocol was then modified to repeat 4 weekly 375 mg/m2 doses at 6 mo intervals for 2 yr. One patient received an extended rituximab course after progression on the limited course.

Results: Pt received limited (n=23, 12 previously untreated) or extended (n=16, 9 previously untreated) rituximab. Median age at treatment was 44 yr (17–71) with 26 males. Among 21 previously untreated pt, stages were I (6), II (8), or III (7). Median follow-up is 60 months (mo) for all pt, 72 mo for limited and 30 mo for extended treatment pt. Treatment-related adverse events were minimal and no grade 3 or 4 toxicities were reported. All but 1 pt responded (OR 97%) with CR/CRu in 27 (69%), and PR in 11 (28%) and SD in 1 (3%). CR/CRu was achieved in 56% limited and 88% extended rituximab pt (p=0.08). No difference was observed in previously treated v untreated pt. To date, 17 progressions (15 limited, 2 extended rituximab) have been recorded. Median FFP was 24 mo for limited and not reached for extended rituximab (p=0.03). FFP at 30 months was estimated at 52% for limited rituximab and 88% for extended rituximab pt. Four pt transformed to large cell lymphoma (one after multiple relapse treatments). Three deaths occurred, one each due to colon cancer, acute leukemia (pt heavily pre-treated with radiation and MOPP), and large cell lymphoma. For all 38 pt, median follow-up from diagnosis is 7.5 yr (range (<1–38) and survival from diagnosis is estimated at 97% at 10 yr and 85% at 20 yr.

Conclusions: Rituximab is an effective treatment in de novo or recurrent NLPHD that may be associated with fewer adverse effects than conventional therapy. FFP is prolonged with an extended treatment but additional follow-up is needed to better assess the duration of benefit.
The Effect of von Willebrand Factor (VWF) on Site-Specific Factor VIII (FVIII) Expression in Hemophilia A Mice.
Authors: Shi Qizhen , Fahs Scot A. , Kuether Erin L. , Montgomery Robert R.
Published: October 2019

von Willebrand factor (VWF) is a carrier protein for factor VIII (FVIII) and protects plasma FVIII from protease degradation. Our laboratory has had a longstanding interest in the association of FVIII with VWF both in vitro and in vivo. Our in vitro studies have demonstrated that FVIII stores together with VWF in both endothelial cells and megakaryocytes if FVIII is made in these cells. Furthermore, we demonstrated that FVIII and VWF are both releasable by agonist stimulation. To investigate the association of VWF and FVIII in vivo, we generated two lines of transgenic mice that express FVIII either in endothelial cells or in platelets using either the endothelial cell-specific Tie2 promoter or the platelet-specific αIIb promoter, respectively. When the platelet-specific FVIII (2bF8) transgene is bred into the FVIIInull mouse, FVIII can only be detected in platelets, with a level of 0.76 ± 0.27 mU/108 platelets in heterozygous and 1.53 ± 0.14 mU/108 platelets in homozygous 2bF8 mice. When the endothelial cell-specific FVIII (Tie2F8) transgene is bred into the FVIIInull mouse, homozygous Tie2F8 mice maintained normal plasma FVIII levels (1.15 ± 0.16 U/ml) and 50% levels in heterozygous mice (0.56 ± 0.16 U/ml). Both 2bF8trans and Tie2F8trans phenotypes effectively abrogate the bleeding diathesis in hemophilic mice. When 2bF8 transgene was bred into a FVIII and VWF double knockout background, the level of platelet-FVIII significantly decreased, but this platelet-derived FVIII was still stored in a-granules and still maintained clinical efficacy. In contrast, when the Tie2F8 transgene was bred into the double knockout background, plasma FVIII dropped to undetectable levels. This is in contrast to the situation in VWFnull mice in which normal endogenous murine FVIII is synthesized with about 10% of normal FVIII activity persisting in plasma. This could be due to a difference in survival between human FVIII and murine FVIII. All Tie2F8trans/FVIIInullVWFnull mice (n=15) survived tail clipping even though there is no FVIII:C detected in the plasma. To investigate the effect of murine VWF on the levels of plasma FVIII, plasma from FVIIInull mice was infused into Tie2F8trans/FVIIInullVWFnull mice to restore VWF levels to 25% of normal. As expected, the endothelial cell-derived plasma FVIII was stabilized by the infused VWF and was detected within 1 hour after infusion, with a peak (25% level) at 4 hours. The level of plasma FVIII at 24 hours was still about 20% of normal while the level of remaining VWF was only 5% of normal. These results demonstrate that VWF is important for site-specific FVIII expression. Co-expression with VWF in platelets is important for optimal platelet-specific FVIII expression and endothelial cell-derived plasma FVIII is VWF-dependent.
Melphalan-Prednisone-Thalidomide to Newly Diagnosed Patients with Multiple Myeloma: A Placebo Controlled Randomised Phase 3 Trial.
Authors: Waage Anders , Gimsing Peter , Juliusson Gunnar , Turesson Ingemar , Fayers Peter
Published: October 2019

Previously untreated patients with multiple myeloma were included in this double-blind randomised trial. Patients not eligible for high dose treatment in Norway, Sweden and Denmark were recruited. The study started in 2002 and accrual of patients stopped 1st of May 2007. Date of analysis was 1st of June 2007. The treatment was randomized between melphalan/prednisone/thalidomide and melphalan/prednisone/placebo. Starting dose of thalidomide was 200 mg escalating to 400 mg. There was no recommendation for prophylaxis for venous thromboembolism. Endpoints were overall survival, event free survival, response, time to progression and quality of life. Interim analysis for safety was performed in 2004 by an independent commitee. Altogether 362 patients with mean age 75 (49–92) years were included, 55% males. The incidence of venous thromboembolism in the blinded treatment arms was 7%. The study will be unblinded and results will be available September 2007.
Human AML Stem Cells Remain Quiescent and Exhibit Chemotherapy Resistance within the Bone Marrow Endosteal Region.
Authors: Saito Yoriko , Yoshida Shuro , Saito Noriyuki , Fukata Mitsuhiro , Ozawa Hidetoshi , Doi Takehiko , Miyamoto Toshihiro , Ohshima Koichi , Uchida Naoyuki , Taniguchi Shuichi , Akashi Koichi , Harada Mine , Shultz Leonard D. , Ishikawa Fumihiko
Published: October 2019

AML is characterized by the clonal expansion of immature myeloblasts initiating from rare leukemic stem cells (LSCs). We developed a primary human AML xenotransplantation model using newborn NOD/SCID/IL2rg−/ − mice. From 9 patients, 4x106 T cell-depleted BMMNCs were injected iv into sublethally irradiated recipients. At 3 months, newborn NOD/SCID/IL2rg−/ − recipients demonstrated superior engraftment efficiency compared with adult NOD/SCID/IL2rg−/ − and newborn NOD/SCID/b2m−/ − recipients (37.8%, 11.9%, 12.9%, respectively; p<0.005). When sorted primary AML hCD34+CD38− cells from 6 patients were injected iv, as few as 103 cells engrafted dose-dependently. Transplanted hCD34+hCD38− cells gave rise to hCD34+hCD38−, hCD34+hCD38+ and hCD34− cells. In serial transplantation, injection of 103 purified hCD34+hCD38− cells resulted in long-term engraftment, giving rise to hCD34+hCD38− LSCs and hCD34+hCD38+/hCD34− blasts, demonstrating self-renewal and differentiation capacities of primary LSCs in vivo. We next examined homing and localization of LSCs using this model. In recipient femoral sections, 94.0% and 78.4% of hCD34+ cells were found in the endosteal region at day 3 and at 4 months after iv injection, respectively (10,065 cells and 24,973 cells examined, respectively; p<0.001 for each), demonstrating that human primary LSCs preferentially home to and engraft in the BM endosteal region. Dual FISH analysis using mouse and human pan-centromeric probes confirmed the human origin of these cells. When AML-engrafted mice were treated with cytosine arabinoside (Ara-C) to examine the effect of anti-leukemia drugs in vivo, PB AML burden became markedly reduced by day 8, followed by AML relapse by days 16–28. Apoptosis analysis of day 3-post-Ara-C BM demonstrated that hCD34+hCD38− LSCs are more resistant to Ara-C induced apoptosis compared to hCD34+hCD38+ and hCD34− cells (Annexin V-7AAD-: 77.1%, 33.3%, 27.5% respectively; n=3 for each; p<0.05 for each). TUNEL staining using day 3 post-Ara-C femoral sections demonstrated that TUNEL-negative cells were preferentially located at the endosteal surface while the cells in the central BM cavity were apoptotic. The cells abutting the endosteum were hCD34+hCD38− and were adjacent to osteopontin+ osteoblasts. % of cells in the G0 phase of cell cycle was significantly greater in the hCD34+hCD38− compared with the hCD34+hCD38+ and hCD34− fractions (n=5 for each; p<0.005 for each), suggesting a potential mechanism underlying relative chemo-resiatance of LSCs. In summary, primary human LSCs preferentially engraft in the BM endosteal areas where they are protected from AraC-induced apoptosis. Cell cycle quiescence of LSCs may be one of the mechanisms underlying chemoresistance of LSCs.
Donor-Derived Squamous Cell Carcinoma after Allogeneic Bone Marrow Transplantation.
Authors: Janin Anne , Murata Hideyuki , Lebeuf Christophe , Gluckman Eliane , Soulier Jean , de Thé Hugues , Bertheau Philippe , Socié Gerard
Published: October 2019

Long-term survivors after allogeneic hematopoietic stem cell transplantation (HSCT) develop solid tumors. Chronic graft-versus-host disease (GvHD) increases the risk of squamous cell carcinoma (SCC). Five of 24 patients with HSCT and SCC, had sex-mismatched HSCT. In 3 of them, tumor sex karyotype studied by combined XY FISH and double immunostaining on the same SCC section, with 3D Z-stack analysis, was compatible with a cancer originating from donor hematopoietic cells. After control by short tandem repeats analysis (PCR-STR) of laser micro-dissected tumor and normal epidermal cells from the recipient and donor or PCR-STR and FISH analysis of a tumoral cell line from one SCC, a recipient origin was found in one case but in the two other cases the donor genotype of the tumor was confirmed. In a third step, PCR-STR of laser micro-dissected tumor in 2 cases after sex-matched transplant proved SCC to be of the donor origin. We thus obtain conclusive evidence that 4 epithelial tumors among 8 studied arose from the engrafted donor cells. Tissue stem cells were proposed to exhibit an unexpected level of plasticity, although issues on cell fusions have lead to some controversies. Only transplantation experiments using genetically distinct recipients and donors can unequivocally demonstrate these changes in cell fate. We have analyzed a unique set of squamous cell carcinomas arising in long terms survivors of allogeneic stem cell transplantation. In these patients, chronic graft-versus host disease greatly favors development of solid tumors, possibly as a consequence of mucosal inflammation and immunosuppression. Thus, donor-derived hematopoietic stem cells recruited to sites of chronic inflammation yielded epithelial tumors.
MOZ and MOZ-CBP Cooperate with NFκB To Induce Expression of NFκB-Dependent Genes.
Authors: Chan Edward M. , Chan Rebecca J. , Comer Elisha M. , Goulet Robert J. , Crean Colin D. , Brown Zachary D. , Fruehwald Amy M. , Yang Zhenyun , Boswell Howard S. , Nakshatri Harikrishna , Gabig Theodore G.
Published: October 2019

Although monocytic zinc finger protein (MOZ/MYST3) maintains normal hematopoietic stem cells, its fusion to the coactivator CREB-binding protein (CBP/CREBBP) induces acute myeloid leukemia (AML). Since leukemic stem cells in AML often exhibit excessive signal-dependent activity of the transcription factor NFκB, we hypothesized that cooperation between NFκB and MOZ-CBP represents an alternative mechanism for enhancing NFκB transcriptional activity. In reporter assays, MOZ and CBP separately induce transcription from the NFκB-dependent interleukin-8 promoter; however, these two proteins together markedly activate its expression. Although MOZ has less potent transcriptional activity than MOZ-CBP, both proteins cooperate with steroid receptor coactivator-1 (SRC1/NCOA1) to activate transcription. Since cooperation between MOZ-CBP and SRC1 is strongly reminiscent of the interaction between MOZ-TIF2/SRC2 and CBP required to induce AML (Deguchi et al, 2003), these findings suggest that MYST family fusion proteins may assemble a conserved leukemogenic transcriptional complex composed of a MYST protein (MOZ, MORF), molecular integrator (CBP, p300), and nuclear receptor coactivator (SRC1, TIF2/SRC2). MOZ also induces multiple NFκB-dependent viral promoters. Importantly, MOZ associates in a protein complex with the DNA-binding p65/RELA subunit of NFκB in vivo and interacts directly with p65 in vitro. Whereas deletion of the leukemia-associated protein (LAP/PHD) or MYST domains decreases the transcriptional activity of MOZ, deletion of either the acidic domain or C-terminal domain completely abrogates its activity. Since the C-terminal domain is absent from MOZ-CBP, these results indicate that the potent transcriptional activity of MOZ-CBP represents a gain-of-function property derived from the retained portion of CBP. Collectively, these studies not only demonstrate that MOZ and MOZ-CBP cooperate with NFκB to enhance expression of NFκB-dependent promoters but also suggest that aberrant interaction between MOZ-CBP and NFκB may play an important role in the pathogenesis of certain acute myeloid leukemias.
Novel Concentration-Dependent Roles for Heparin in Modifying LPS-Induced Activation of Mononuclear Cells in Whole Blood.
Authors: Hochart Helene , Jenkins Vince , Smith Owen P. , White Barry , O’Donnell James
Published: October 2019

Background: In addition to their etsblished anticoagulant activity, unfractionated heparin (UFH) and low molecular weight heparin (LMWH) are known to possess clinically important immuno-modulatory properties. However different studies have reported conflicting pro- and anti-inflammatory effects in association with heparin. Moreover, the molecular basis for these heparin effects on inflammation remains unclear. In view of the wide and diverse clinical indications for heparin, it is clearly of direct translational relevance to define how UFH and LMWH differentially regulate inflammatory responses to LPS in-vivo.

Objectives: To determine how UFH and LMWH regulate lipopolysaccharide (LPS)-induced activation of human mononuclear cells in whole blood, and define the role of lipopolysaccharide binding protein (LBP) in mediating this effect.

Methods: Whole blood was pre-treated with UFH or LMWH (0.1–200 IU/ml), prior to stimulation with LPS (10ng/ml). After 6 hours, monocyte pro-inflammatory cytokine (interleukin (IL)-1b, IL-6, IL-8, and TNF-a) secretion was determined by plasma ELISA. Parallel experiments using THP-1 cell line and primary monocytes were performed under serum-free conditions, in the presence or absence of varying doses of recombinant human LBP (range: 50–100nM).

Results: Under serum-free conditions, heparin demonstrated dose-dependent anti -inflammatory effects, significantly reducing secretion of pro-inflammatory cytokines (IL-1b, IL-6, IL-8, and TNF-a) in response to LPS-stimulation of THP-1 cells and primary monocytes. In contrast, in the presence of LBP, both UFH and LMWH demonstrated dose-dependent pro-inflammatory effects at all heparin concentrations. In ex-vivo whole blood experiments, pro-inflammatory effects (increased IL-1b and IL-8 following LPS-stimulation) of heparin were also observed, but only at supra-therapeutic doses (10–200IU/ml).

Conclusion: In keeping with previous reports, we have demonstrated that both UFH and LMWH can significantly down-regulate cytokine (TNF-a, IL-1b, IL-6 and IL-8) secretion in response to LPS-activation in-vitro. However our novel data demonstrate that the effect of heparin on monocyte activation by LPS is significantly more complex in the setting of whole blood. Firstly, in contrast to the anti-inflammatory effects observed under serum-free conditions, we found that in whole blood, high concentrations of heparin exerted marked pro-inflammatory effects. Secondly we have also demonstrated that the effects of heparin in whole blood are entirely dependent upon heparin concentration and LBP concentration.
Platelets and Heparin Induced Thrombocytopenia Antibodies Do Not Influence the Inhibitory Activity of Argatroban on Thrombin Generation.
Authors: Gerotziafas Grigoris T. , Roman Marie-Paule , Verdy Elisabeth , Hatmi Mohamed , Samama Meyer M. , Elalamy Ismail
Published: October 2019

Argatroban is a synthetic, reversible, direct thrombin inhibitor (DTI) used in patients with heparin induced thrombocytopenia (HIT). Argatroban as other DTIs prolongs PT, aPTT and ecarin clotting time. Argatroban added in vitro in normal platelet poor plasma (PPP) inhibits tissue factor (TF) triggered thrombin generation (TG) in a concentration dependent manner. We studied the influence of platelets, HIT antibodies and residual heparin, on the inhibitory effect of argatroban on TG. Argatroban (0 to 2 μM) was added in normal platelet rich plasma (PRP) and PPP, in pool PPP from three consecutive HIT patients (HIT-PPP) and in HIT-PRP prepared by mixing normal PRP with HIT-PPP (v/v). HIT-PRP and HIT-PPP were containing residual heparin (0,035 and 0,07 anti-Xa IU/ml respectively). All experiments were repeated 3 times. TG was triggered in the presence of TF (Dade-Behring Innovin; 1/1000 final dilution in plasma) and assessed with Calibrated Automated Thrombinoscope®. TG in PPP was triggered by adding PPP reagent (Thrombogram-Thrombinoscope®). Lag time (LT), time to peak (ttP), peak (P), endogenous thrombin potential (ETP) and mean velocity index (MVI) of thrombin generation were measured. The concentrations of argatroban which prolonged 2-fold the LT and the ttP and which inhibited 50% (IC50) the P, ETP and MVI were calculated. In the presence of low argatroban concentrations (0,1 an 0,2 μM) an artifactual increase of TG was observed. This is probably due to the interference of alpha2macroglobulin-bound thrombin with the fluorogenic substrate (as shown by Hemker’s group for other reversible DTIs). Argatroban at 1 μM inhibited TG by 50% in both normal PRP and PPP. Argatroban at 1 μM induced a 2-fold prolongation of aPTT (Table 1). HIT antibodies did not modify the inhibitory activity of argatroban on TG in HIT-PRP and HIT-PPP. The presence of traces of heparin in plasma from HIT patients had a synergistic effect with argatroban on the inhibition of TG (Table 1). Our in vitro study shows that argatroban at concentration achieved in clinical practice (about 1 μM) induces 50% inhibition of TG. The inhibitory activity of argatroban on TG is not modified by the presence of platelets or HIT antibodies. In contrast, traces of residual heparin in plasma from HIT patients amplify the inhibition of thrombin generation induced by argatroban. This finding has to be confirmed in a prospective clinical study since it implicates an increased vigilance during the switch from heparin to argatroban in acute HIT patients.

Table 1. Inhibitory activity of argatroban on thrombin generation. normal PRP normal PPP HIT-PRP (0,03 aXa/ml) HIT-PPP (0,07 aXa/ml) Lag-time x2 0,7 μM 0,7 μM 0,9 μM 0,1 μM ttPeak x2 1 μM 1 μM 1 μM 0,2 μM Peak IC50 1 μM 1 μM 0,7 μM 0,3 μM ETP IC50 1 μM 1 μM 0,7 μM 0,3 μM MRI IC50 1 μM 1 μM 0,5 μM 0,3 μM PTx2 - 1 μM - - aPTT x2 - 1 μM - -
The Reliability of Point-of-Care (POC) Prothrombin Time Testing - A Comparison of CoaguChek XS® INR Measurements with Hospital Laboratory Monitoring.
Authors: Ryan Fiona , O’Shea Susan , Ortel Thomas L. , Byrne Stephen
Published: October 2019

Background The development of Point of Care (POC) testing devices enables patients to test their own International Normalised Ratio (INR) at home, and has lead to the development of patient self- testing (PST) programmes. This facilitates more frequent testing of anticoagulation levels which in turn improves time in therapeutic range (TTR) and reduces bleeding and thrombotic complications in patients on oral anticoagulation therapy (OAT).1 External quality control of POC devices is essential to ensure that results are robust. One approach is to collect a venous sample at the same time as POC testing, which is analysed using an appropriate hospital laboratory analyzer. It has been recommended that INR results should be within 0.5 of each other when measured by each method2. However, previous studies have shown that when compared with clinical laboratory values, POC devices may show statistically significant differences between the INR values.3 Aim The aim of this study is primarily to compare TTR and adverse events between two different approaches to OAT management and more specifically in this presentation to examine the intra patient variability of INR measurements performed by two different techniques.

Methods 160 patients routinely attending the anticoagulant clinic of Cork University Hospital (CUH) were enrolled to take part in a prospective randomized controlled, cross-over design trial of supervised PST of warfarin therapy using an internet based expert system (Coagcare®, ZyCare, Inc) and POC testing device (CoaguChek XS®, Roche, UK) vs. rountine medical care in CUH. During the 6 months PST arm of the study a venous blood sample was taken from each patient for laboratory determination of the INR on 3 occasions; at enrolment, 2 months and 4 months. This measurement was then compared with the INR measured on the Coaguchek XS® machine using capillary blood from the patient’s fingertip.

Results 88 patients have being enrolled in the PST arm of the study to date and 454 POC measurements have been compared with laboratory values. Good correlation has been shown between the two methods of INR determination (r2 = 0.8314), however, statistically significant differences did occur (p < 0.005). A Bland and Altman plot showed good agreement between the dual measurements at values between 2.0–3.5 INR units. However there seemed to be increasing disagreement between the two methods as the INR rose above 3.5. Four patients were excluded from the study because the differences in the INR measurements were greater than 0.5 INR units. Conclusion POC testing devices for OAT are reliable and safe. External quality assessment should be inherent to any programme involving these devices. Clinicians involved in the use of POC anticoagulation monitoring should be aware that POC and laboratory values may differ and intervene if there is not adequate correlation between the two INR measurements.
Acquired Genetic Abnormalities in Acute Lymphoblastic Leukaemia in Patients with Down Syndrome.
Authors: Kearney Lyndal , Horsley Sharon W. , Bateman Caroline M. , De Castro David Gonzalez , Young Bryan D. , Chaplin Tracy , Eguchi Minenori , Greaves Mel
Published: October 2019

Children with Down syndrome (DS) have a 30 fold increased risk of developing leukaemia compared to their non-DS counterparts. While it is now known that cooperating mutations in the haemopoietic transcription factor GATA1 occur in all cases of acute megakaryoblastic leukaemia in Down syndrome patients, the additional genetic events which confer an increased risk of acute lymphoblastic leukaemia in Down syndrome (ALL-DS) are unknown. We initiated a search for mutations in the coding region of candidate genes including RAS, B-RAF, FLT3, KIT and JAK2 in a series of ALL-DS cases. No mutations were identified. We then carried out high resolution (Affymetrix 250K Nsp and 250K Sty) SNP array analysis of leukaemic blast cell DNA from 9 cases of ALL-DS, the majority of which had matched remission DNA. Overall, the whole and partial chromosome gains and losses were in agreement with the karyotype, but in all cases these were updated and refined. There were between 1 and 12 additional copy number alterations per case, with small focal deletions comprising 1 or 2 genes being more frequent than gains. The most common of these was a focal deletion of the CDKN2A gene (4 cases), all of which had either a partial deletion or copy number neutral LOH of the whole of 9p. The other most common focal deletions were of 12p13.3 (ETV6 gene) and 9p13.2 (PAX5), found in 2 cases each. Other regions affected included 3q13.2 (BTLA), 5q33.3 (EBF1), 13q14.2 (RB1) and 20p12.2 (including c20orf94), identified in 1 case each. These results indicate that the secondary genetic events in ALL-DS are similar to those for ALL overall (Mullighan et al., Nature446: 758–764, 2007). The number and pattern of submicroscopic genetic abnormalities more closely resembles that of ETV6-RUNX1 positive ALL than high hyperdiploid ALL (which includes acquired trisomy 21).
Epigenetic Repression of the Adaptor Molecule LAT2 by the Leukemic Fusion Protein AML1/ETO.
Authors: Duque-Afonso Jesus , Berg Tobias , Heidenreich Olaf , Luebbert Michael
Published: October 2019

The chromosomal translocation (8;21) fuses the hematopoietic transcription factor AML1 (RUNX1) with ETO, resulting in the leukemia-specific chimeric protein AML1/ETO. This fusion protein represses transcription by recruiting a nuclear co-repressor complex containing HDACs and DNMT1 to its target promoters. Previously, we have identified a novel in vivo AML1/ETO target gene, LAT2 (NTAL/LAB/WBSCR5), which is involved in FcεR I, c-Kit, B cell- and T cell receptor signalling. Notably, LAT2 is strongly repressed in AML1/ETO positive cells including primary AML blasts, which was confirmed by others in several large AML cohorts. We have now addressed the molecular mechanisms of AML1/ETO-mediated LAT2 repression. AML1/ETO was induced by Ponasterone A in an ecdysone-inducible system in U937 cells (9/14/18 cell line). To deplete AML1/ETO in t(8;21)-positive cells, we electroporated Kasumi-1 cells with AML1/ETO siRNA. To interfere with epigenetic modifications more directly, cells were treated with the DNMT inhibitor decitabine (DAC) and 4 different HDAC inhibitors. LAT2 expression was determined by Northern Blot, qRT-PCR and Western Blot. HDAC occupation and the histone status of the LAT2 promoter was examined by chromatin immunoprecipitation (ChIP). LAT2 mRNA was downregulated already after 4 hours of conditional expression of AML1/ETO in 9/14/18 cells, and constitutively repressed in the AML1/ETO-positive Kasumi-1 and SKNO-1 cells. siRNA-mediated AML1/ETO depletion caused a 9-fold upregulation of LAT2 in Kasumi-1 cells, suggesting a possible direct mechanism of repression. To address this question, we performed ChIP assays for the LAT2 promoter after AML1/ETO induction in 9/14/18 cells. AML1/ETO inhibited acetylation of histone H3, H3K9 and H4, but did not affect trimethylation of H3K4. These changes were associated with the recruitment of HDAC2, but not HDAC1 and HDAC3, to the LAT2 promoter. The HDAC inhibitors MS-275, SAHA, TSA and valproic acid induced LAT2 mRNA in a dose-dependent manner in AML1/ETO-expressing Kasumi-1, with MS-275 being the most efficient inhibitor. MS-275 induced LAT2 expression also in t(8;21)-positive SKNO-1, but not in AML1/ETO-negative HL60 and U937 cells. LAT2 mRNA was also upregulated in a dose-dependent manner after DAC treatment in Kasumi-1 cells. The combination of DAC and MS-275 had a synergistic effect on inhibition of cell growth, acetylation of histones H3 and H4, and re-expression of LAT2 mRNA. MS-275-mediated re-expression of LAT2 was associated with an increase in acetylation of histone H3, H3K9, H4 and trimethylation of H3K4. The increase of activating histone modifications was associated with the release of HDAC1, HDAC2 and HDAC3 from the LAT2 promoter. In conclusion, the epigenetic changes of the LAT2 promoter caused by AML1/ETO could be pharmacologically reverted by inhibition of histone acetylation.
Genetic and Clinical Spectrum of Osteopetrosis.
Authors: Schulz Ansgar S , Moshous Despina , Debatin Klaus-Michael , Villa Anna
Published: October 2019

Abstract 1087

Poster Board I-109

Background

Osteopetrosis (OP) is a multi-systemic inborn disorder characterized by heterogeneous clinical presentations on the basis of a variety of mutated genes. We performed a retrospective analysis in patients with OP from 28 European centers on behalf of the ESID and the EBMT. Main objectives of this analysis were survival and quality of life with respect to genetic background, initial clinical presentation, and treatment modality.

Methods

A set of clinical parameters, genetic findings, transplant information and follow up data were collected using specific questionnaires. Questionnaires were sent at several time points to European centers treating patients with OP and the collected data were stored in a central OP database. DNA sequencing of candidate genes involved in human OP was done predominantly at laboratories in Milan, Paris and Ulm.

Results

Data of 173 patients and of 134 hematopoietic stem cell transplantations (HSCT) were obtained during a time frame between 1983 and 2008. In the whole cohort, mutations in the following genes were detected: 47% biallelic TCIRG1, 9% biallelic CLCN7, 6% monoallelic CLCN7, 5% OSTM1, 2% RANK, 1% RANKL; 30% of patients have no mutations in these genes or could not be tested completely. In 23% of patients HSCT was not considered because of severe neurological problems or other complications mostly in the context of OSTM1 or CLCN7 mutations, or because of less severe phenotypes mainly due to monoallelic CLCN7 mutations (autosomal dominant OP, ADO).

Follow up data of 103 transplanted patients were available. With respect to donor type, the probability of survival at 2 years was 88% for matched family donors, 80% for matched unrelated donors, 68% for HLA-haploidentical donors and 54% for mismatched unrelated donors. There was no significant difference in survival of patients with different gene mutations but a trend to a worse long term outcome in CLCN7 patients. Most notably, several major sequelae were present in the majority of surviving patients. Visual impairment or dwarfism were found in about 2/3 of children showing no significant influence of the affected genes. About 10% of patients are suffering from significant persistent neurological problems after HSCT, which were found in patients with TCIRG and CLCN7 mutations. The quality of life were judged as normal by the parents in about 2/3 of surviving patients.

Conclusion

HSCT in infantile OP results in acceptable survival rates even after HLA-nonidentical transplants. However, outcome is influenced by a set of specific disease related problems and long term sequelae. The heterogeneity of OP necessitates an individualised therapeutic strategy respecting the genetic background, the clinical presentation and the type of the available donor.

Disclosures

No relevant conflicts of interest to declare.
Second Autologous Stem Cell Transplant (ASCT) as Salvage Therapy in Patients with Relapsed Multiple Myeloma.
Authors: Mikhael Joseph R. , Zadeh Sahar , Stewart A. Keith , Chen Christine , Trudel Suzanne , Kukreti Vishal , Winter Andrew , Franke Norman , Reece Donna E.
Published: October 2019

Abstract 1217

Poster Board I-239

Background

Single autologous stem cell transplant (ASCT) is considered the standard of care after induction therapy for younger multiple myeloma (MM) patients (pts). However, it is not curative and virtually all patients will ultimately relapse. As more options are available to treat relapsed disease, the role of a second ASCT as salvage therapy is unclear.

Method

Retrospective review of all MM pts who received a 2nd ASCT as salvage therapy at Princess Margaret Hospital.

Results

Between February 1997 and July 2009, 79 MM pts received a second ASCT for relapsed MM at our institution. Median age was 60 yrs (range 39-72) at second transplant. 48 pts (61%) were male. Immunoglobulin subtype included IgG (42), IgA (21), light chain (11), nonsecretory (3), IgM(1) and IgD (1). Transplant conditioning regimen for first transplant was high dose melphalan (MEL) 140-200 mg/m2 in 67, busulphan and cyclophosphamide in 1, and combinations of MEL with etoposide (E) or TBI in the rest. 2nd ASCT conditioning consisted of MEL alone in 76, the remaining 3 had MEL with either TBI or E. The median time to relapse after the first transplant was 2.72 years (0.81-8.26), with a median interval between transplants of 3.61 years (1.63-9.59). Response to first transplant in 78 evaluable patients was 13 CR/nCR (17%), 56 PR/VGPR (72%), 9 MR/SD (12%). Nineteen pts received maintenance therapy between transplants. Two transplant-related deaths occurred following 2nd ASCT. In 73 evaluable patients, response to second transplant was 11 CR/nCR (15%), 57 PR (78%), 6 MR/SD (8%). After 2nd ASCT, with median follow up 5.92 years (71 months), median progression-free survival (PFS) was 18.5 months and median overall survival (OS) was 4.4 years. Long term progression-free status based on the progression-free interval after 1st ASCT is summarized Table 1. PFS based on progression free interval after 1st ASCT is outlined in Figure 1.

Conclusions

2nd ASCT is a feasible and safe salvage therapy in relapsed MM patients. It is effective in providing a median progression free survival of 18.5 months and median overall survival of 4.4 years (52.8 months) after 2nd ASCT. This is comparable if not better than modern salvage chemotherapies. The longer the disease free interval after 1st ASCT the more effective 2nd ASCT is at extending both progression free survival and overall survival. It is reasonable, therefore, to consider a 2nd ASCT if the time to progression is greater than 2 years after first ASCT.

Disclosures

Chen: Celgene: Honoraria, Research Funding; Ortho Biotech: Honoraria. Trudel:Celgene: Honoraria, Speakers Bureau; Ortho Biotech: Honoraria. Kukreti:Celgene: Honoraria.
Improved Survival for Patients Undergoing Autologous Stem Cell Transplant for Primary CNS Lymphoma in First or Later Remission.
Authors: Johnston Patrick B , Micallef Ivana N , Ansell Stephen M , Inwards David J , Porrata Luis F. , O'Neill Brian Patrick
Published: October 2019

Abstract 1222

Poster Board I-244

Background

Survival for patients with primary CNS lymphoma (PCNSL), in general, is poor with patients requiring frequent chemotherapy treatments or receiving whole-brain radiation therapy, which can potentially result in significant neurologic decline and dementia. Because of the improved survival of high risk patients with aggressive lymphoma undergoing autologous stem cell transplant (ASCT), we began ASCT for patients with PCNSL in first or later remission with chemotherapy sensitive disease. We now update on outcomes of patients who have had at least 100 day follow up post ASCT.

Baseline characteristics

Between June, 2000 and January, 2009, 22 patients underwent ASCT for PCNSL. Median age at transplant was 50 years old (range 26-67). Median number of prior treatments 1 (range 1-3). Median time from diagnosis to transplant was 7.2 months (range 2.9 to 75.8). Median International Extranodal Working Study Group Prognostic Score: 2 (range 0-3). Disease status at transplant: First CR 10 patients, later CR or PR 12 patients.

Results

Twenty-two patients underwent ASCT for PCNSL and have a minimum of 100 days follow-up. All patients received BEAM conditioning. Median follow up post-transplant was 30 months (range 3-107 months). Eight patients have relapsed at a median of 217 days (range 40-1349). Of the patients who relapsed, four have died of disease progression and the remaining four are alive after additional therapy. Median overall survival from diagnosis or transplant has not been reached. Median progression free survival from transplant was 70 months.

Conclusions

Although limited by patient selection and retrospective biases, this review suggests that ASCT for PCNSL demonstrates improved overall survival when compared to historical controls with similar PCNSL Prognostic Scores (2 year survival for patients from diagnosis with PS 2-3 was 48% in a prior published study). ASCT in first remission in patients with PCNSL appears promising and may limit the need for additional therapy which can be myelosuppressive or result in neurologic decline secondary to radiation therapy in patients who are appropriate candidates.

Disclosures

No relevant conflicts of interest to declare.
The First Human Cell Line Recombinant Factor VIII (Human-cl rhFVIII) - Functional Properties and Differences From Currently Available Recombinant Factor VIII Products.
Authors: Schwartz Bruce A. , Sandberg Helena , Agerkvist Irene , Kannicht Christoph , Ramström Margareta , Stenlund Peter , Oswaldsson Ulla , Brunberg Tobias
Published: October 2019

Abstract 1298

Poster Board I-320

All current recombinant factor VIII products produced today are derived from the expression of FVIII in hamster cell lines. This is a report of the functional properties of a newly developed recombinant factor VIII concentrate produced in a human cell line. The possible advantage of using a human cell line for expression of recombinant factor VIII is that a human pattern of post-translational modifications (PTM), such as glycosylation, are obtained. These human like PTMs, may possibly lead to improved function and a reduced immunogenic product profile compared to recombinant factor VIII products from CHO or BHK cells that have hamster-specific PTM. Data are presented on the characteristics of the Human-cl rhFVIII, a B-domain-deleted factor VIII protein produced in HEK293F cells, and how they compare with those of other factor VIII products.

Human-cl rhFVIII is highly pure (>99.9% purity) with a high specific activity of 8000-11000 IU/mg protein, and a FVIII:C to FVIII:Ag ratio close to 1.0. This indicates that the entire factor VIII protein in the product is active. Human-cl rhFVIII showed no significant assay discrepancy between one-stage and chromogenic assay results. Because of the human glycosylation profile of the new Human-cl rhFVIII, there was an absence of the antigenic type of sialic acid, N-glycolylneuraminic acid as well as of antigenic Galα (1,3) Gal epitopes found in the non humane derived products. Another type of PTM of factor VIII is the sulfation of certain tyrosines that are crucial for the biological function e.g. the interaction with von Willebrand factor (vWF). Human-cl rhFVIII was found to be optimally sulfated when compared to endogenous human FVIII. In this context, it is of interest to note that Human cl-rhFVIII showed a 60% higher affinity in binding to vWF compared to other recombinant factor VIII products and the number of molecules in the product able to bind to vWF was significantly higher for Human-cl rhFVIII.

Interaction with thrombin and capacity to generate thrombin were shown to be similar to those of plasma-derived factor VIII. The sensitivity to be inactivated by activated protein C was similar to all currently licensed rhFVIII products. The important properties of Human cell line derived recombinant factor VIII may be very promising for its use in future treatment of Haemophilia A.

Disclosures

Schwartz: Octapharma: Employment. Sandberg:Octapharma: Employment. Agerkvist:Octapharma: Employment. Kannicht:Octapharma: Employment. Ramström:Octapharma: Employment. Stenlund:Octapharma: Employment. Oswaldsson:Octapharma: Employment. Brunberg:Octapharma: Employment.
Survival Among Leukemia Patients Over the Past Three Decades: A Single Institution Based Retrospective Review From 1976 to 2006.
Authors: Sheikh Hassan S , Joshi Monika , Camacho Fabian , Hand Lisa A , Bayerl Michael G. , Rybka Witold B
Published: October 2019

Abstract 1394

Poster Board I-416

Background:

Leukemia is the fifth most common cause of cancer death in men and seventh most common cause in women in the US. Since the advent of multi-drug chemotherapy, studies have shown improvement in survival for specific age groups. However, there have been few studies showing the impact of evolving therapies on survival for the total burden of leukemia patients. We assessed survival for an aggregate population of all patients presenting with leukemia to a regional tertiary university hospital over the past 3 decades.

Goal:

To assess the magnitude of improvement in survival for patients with leukemia over the past 3 decades.

Methods:

We analyzed data from the Penn State Hershey Medical Center Cancer Registry, selecting all cases diagnosed with leukemia by ICD-0-3 codes, excluding chronic lymphocytic leukemia, from Jan 1st 1976 to Dec 31st 2006. Five and ten year (yr) absolute survival rates during five time periods [group (gp) 1: 1976-1980, gp 2: 1981-1985, gp 3: 1986-1990, gp 4: 1996-2000, gp 5: 2001-2006] were obtained using conventional period analysis (PA). In addition, a period Cox Proportional Model (CPH) was fit to the data, allowing for survival risk estimates of 5 yr survival, statistical testing of time periods, and adjustments for age at diagnosis. SAS v 9.1 was used to obtain estimates, with Brenner's PERIOD macro used for PA and PHREG used for CPH.

Results:

Of 1892 patients, chronic myeloid leukemia (CML) accounted for 11%, acute myeloid leukemia (AML) 52% and acute lymphoblastic leukemia (ALL) 37%. Median age was 44 yr and mean was 40 yr with a standard deviation (SD) of 27.2 yr. Median follow up was 1.6 yr and mean was 3.9 yr with SD 5.1 yr. Approximately 13% (N=249) survived beyond 10 yr. CPH adjusted for age demonstrated an improvement in 5 yr survival among all patients with leukemia of 16%, Hazard Ratio (HR)=1.57, p=0.0002, and 16% in 10 yr survival, HR=1.6, p= 0.0001, between gp 5 and gp 1. Consistent improvements in age adjusted 5 yr survival were detected for CML (24%, HR=2.06, p=0.124), AML (17%, HR=1.53, p<0.0015) and ALL (21%, HR=1.8, p=0.01) comparing gp 5 to gp 1. Results for conventional PA were similar. There was a 19% improvement in 5 yr survival between gp 5 and gp1 for all patients with leukemia. There was greater variability by PA in the subgroups, CML(58% 5 yr), AML (7% 5 yr) and ALL (24% 5 yr).

Conclusion:

Our analysis shows that there is a significant improvement of 16% in overall five and ten year survival rates in all leukemia patients over the last 30 years after adjusting for age at diagnosis. Subgroup analysis shows comparable improvements among the three diagnostic groups.

Disclosures:

No relevant conflicts of interest to declare.