bluuebunny/crossref_metadata_2025_split huggingface dataset:
Activation Status of Integrin αIIbβ3 in Essential Thrombocythemia with Calreticulin Mutation
Authors: Yamanouchi Jun , Hato Takaaki , Matsubara Etsuko , Azuma Taichi , Nakanishi Hideyuki , Fujiwara Hiroshi , Yakushijin Yoshihiro , Yasukawa Masaki 
Published: October 2019

Abstract
Mutations in the calreticulin (CALR) gene were recently discovered in patients with essential thrombocythemia (ET) lacking the JAK2V617F and MPLW515 mutations. It has been reported that CALR is a potential regulatory protein of integrin activation based on the interaction between CALR and a conserved sequence of GFFKR in the integrin α cytoplasmic tails. Recent studies suggest that calreticulin activates β1 integrin and modulates integrin-associated signaling. In this study, we examined the effect of CALR mutations on integrin αIIbβ3 activation. We first identified mutations of JAK2, MPL, and CALR genes in 37 patients with WHO defined ET and explored clinical characteristics of patients with CALR mutation. The patients with JAK2 V617F were 22 (59%), MPL W515L was 1 (3%), and CALR mutations were 10 (27%). The two types of CALR mutations were found; deletion (52-bp deletion; c.1092_1143del) and insertion (5-bp insertion; c.1154_1155insTTGTC) mutations. The patients with CALR mutations had lower hemoglobin and leukocyte count compared with JAK2 V617F patients, but platelet count did not have a difference between the CALR and JAK2 mutation groups. Nine (41%) of 22 patients with JAK2V617F had a thrombotic event while 1 (10%) of 10 patients with CALR mutation did (p<0.05), suggesting that patients with CALR mutation had a lower risk of thrombosis than JAK2 V617F patients. Two patients with CALR mutations developed myelofibrosis while no patient with JAK2V617F did. One patient with CALR mutation developed acute myeloid leukemia, with persistence of the CALR mutation in his leukemic cells.To see if the CALR mutation affects activation status of αIIbβ3, we examined the binding of PAC1, a monoclonal antibody recognizing the active conformation of αIIbβ3, to platelets from 5 patients with CALR mutation and 12 patients with JAK2V617F in the presence or absence of ADP. Platelets from all the 5 patients with CALR mutation showed the same level of PAC1 binding as platelets from healthy subjects. Overexpression of recombinant CALR proteins in CHO cells expressing αIIbβ3 by transfection of a protein-expression vector containing wild-type, deletion, or insertion mutant CALR had no effect on PAC1 binding. On the other hand, platelets from 4 of 12 patients with JAK2V617F had an increase in PAC1 binding in the presence and absence of ADP compared with platelets from healthy subjects. All 4 (100%) of 4 patients with increased PAC1 binding had a thrombotic event while 4 (30%) of 13 patients with normal PAC1 binding did. Our study suggests no activation of integrin αIIbβ3 by CALR mutation, which may explain a clinical finding of a low risk for thrombosis in patients with CALR mutation.

Disclosures
No relevant conflicts of interest to declare.
Ultra-Deep Sequencing Defines Stem Cell-Specific Diversity Patterns in Acute Myelogenous Leukemia
Authors: Mencia-Trinchant Nuria , Roboz Gail J. , Carroll Martin P. , Guzman Monica L. , Hassane Duane C 
Published: October 2019

Abstract
Acute myelogenous leukemia (AML) is a fatal disease with dismal outcomes. Despite aggressive chemotherapy regimens, most patients relapse and ultimately succumb to their disease. Relapse is thought to be driven by a surviving residual population of leukemia stem cells (LSCs) that remain quiescent and survive chemotherapy. Moreover, relapsed disease has been shown to be frequently more genetically complex, acquiring new mutations not seen in de novo disease. Since the acquisition of new mutations and survival of LSCs are both evident with relapse, we hypothesized that genetic diversity of LSCs is a likely contributor to disease outcomes. To assess diversity, we performed ultra-deep targeted next generation sequencing at up to 15000x non-duplicate depth of coverage, constructed local haplotypes, and performed subclonal variant analysis on AML samples across varying molecular and cytogenetic risk strata. The resulting high resolution molecular portrait enabled us to examine the clonal composition of both lymphocytes and leukemia, including LSCs.
Consistent with previous reports, we detected established pre-leukemic mutations in DNMT3A(R882H) evident in both patient-matched sort-purified lymphocytes and leukemic cells. In this patient, the pre-leukemic acquisition of DNMT3A(R882H) observed in lymphocytes was followed by additional acquisition of NRAS(G13D) seen exclusively in the LSC and non-LSC compartments, but not the sorted lymphocyte compartment. Interestingly, we found that LSC populations frequently exhibited more complex clonal architecture and nucleotide diversity (p < 10-18) compared to non-stem matched patient counterparts. Because LSCs inherently resist chemotherapy, genetic lesions occurring in the surviving LSC pool are capable of supporting sustainable outgrowths of new drug resistant clones. Indeed, within this complex stem cell architecture, we identified deeply subclonal lesions in clinically relevant AML genes including FLT3. For example, in one high risk patient, the drug-resistant tyrosine kinase domain mutant FLT3(D835Y) was evident at below 0.25% allele frequency in the LSC compartment. This FLT3 allele demonstrated penetrance into the bulk tumor, but not into normal lymphocytes, indicating the AML-specific context of this subclonal variant.
The data suggest that, in addition to surviving chemotherapy, LSCs represent a diverse reservoir of surviving residual cells that can support emergence of drug resistant mutants and refractory relapse. Moreover, ultra-deep sequencing of LSCs and bulk disease will enable improved precision medicine approaches to enhance AML outcomes via early detection of emergent and clinically impactful mutant alleles.

Disclosures
No relevant conflicts of interest to declare.
Identification of a Novel and Efficacious Oral Proteasome Inhibitor As a Potential Clinical Development Candidate
Authors: Won Kwang-Ai , Kim Byung Gyu , Jung Cheol-Kyu , Park Eok , Baek Eunbok , Park Jonghoon , Kang Seung-Wan , Kim Tae Hun , Jung Youngjoo 
Published: October 2019

Abstract
The proteasome is a validated anti-cancer target and various small-molecule inhibitors are currently in clinical development or on the market. However, adverse events and resistance associated with those proteasome inhibitors call for a new generation of drugs.
LC53-0110 and its analogues were identified as novel, reversible, selective, potent, and orally bioavailable proteasome inhibitors which have physicochemical profiles that are distinct from bortezomib, carfilzomib or ixazomib. As compared to those inhibitors, LC53-0110 is far more selective for the chymotrypsin-like proteolytic (β5) site of the 20S proteasome, with an IC50 value of less than 10 nM over the trypsin-like site and the caspase-like site. LC53-0110 treatment showed accumulation of ubiquitinated proteins and inhibited cell viability with a low nM range potency in various hematologic and solid tumor cell lines including multiple myeloma (MM), lymphoma, colon tumor, breast tumor, and lung tumor.
When a single dose was administered orally to tumor-bearing mice, LC53-0110 showed both greater maximum and sustained tumor proteasome inhibition as compared with ixazomib in MM and solid tumor xenograft models. The robust pharmacodynamic responses in tumor translated to the efficacy studies. In the RPMI8226 (MM) efficacy model, oral administration twice a week at 50 mg/kg inhibited tumor growth 68% by three weeks. In the MM.1S (MM) efficacy model, oral administration twice a week at 100 mg/kg completely abolished measurable tumor mass by one week and prevented any tumor re-growth during the three-week study period.
In the initial studies, LC53-0110 also showed potent activity on freshly isolated CD138+ cells from MM patients (pts) who are resistant/refractory (r/r) to current FDA-approved drug treatment. Further studies, including comparison of the naïve pts vs. thalidomide r/r pts and bortezomib r/r pts as well as their potential clinical implications will be discussed.

Disclosures
Won: LG Life Sciences: Employment. Kim:LG Life Sciences: Employment. Jung:LG Life Sciences: Employment. Park:LG Life Sciences: Employment. Baek:LG Life Sciences: Employment. Park:LG Life Sciences: Employment. Kang:LG Life Sciences: Employment. Kim:LG Life Sciences: Employment. Jung:LG Life Sciences: Employment.
Risk of Wernicke’s Encephalopathy in Patients with Myeloproliferative Neoplasm
Authors: Wu Jasmanda , Mirza Nadeem , Partisano Angela M. , Gallagher Christina , Vaze Anjali , Zhang Ling , Gallagher Meghan , Lin Stephen , Juhaeri Juhaeri 
Published: October 2019

Abstract
Background :Wernicke’s encephalopathy (WE) is an acute neuropsychiatric syndrome resulting from thiamine deﬁciency. This neurological deterioration is potentially dangerous because it predisposes the patient to hypothermia and decreased immunity. Cancer patients are at high risk of WE due to chronic malnutrition, chemotherapy-induced nausea and vomiting, and consumption of thiamine by rapidly growing tumors. Very little published research addresses the risk of WE in patients with myeloproliferative neoplasm (MPN).
Objectives: To estimate the incidence of WE among patients with MPN, and compare it with those without MPN.
Methods: Patients with a diagnosis of MPN, including polycythemia vera (PV), essential thrombocythemia (ET) and primary myelofibrosis (PMF) were identified from 1/1/2003 to 03/31/2013 using the US MarketScan database. The control cohort consisted of a random sample matched to the MPN cohort by age and gender in a 1:1 ratio. Diagnosis of WE was ascertained using ICD-9-CM 265.0, 265.1, 291.1, 294.0 or J code J3411. The two study cohorts were followed for WE from the index date. Cox proportional hazards modeling was used to compare the rates between two cohorts and also control for confounding variables.
Results: A total of 39,761 MPN patients, including PV (51%), ET (42%), post-PV MF (0.2%), post-ET MF (1%), and PMF (2.8%) were identified. Approximately 27% of them were > 65 years of age, and 51% were male. Patients with MPN had higher rates of WE, compared to those without MPN (MPN vs. non-MPN: 1.09 vs. 0.39/1000 person-year, hazard ratio = 2.19, 95% confidence interval 1.43-3.34). The incidence rate of WE was higher in males (male vs. female: 0.93 vs. 0.55/1000 person-year). No specific pattern was observed in age subgroups for both cohorts (e.g. MPN: age <18 vs. 18-39 vs. 40-49 vs. 50-59 vs. 60-69 vs. >70 = 0 vs. 0.85 vs. 1.09 vs. 1.16 vs. 0.72 vs. 0.86/1000 person-year).
Conclusions: Patients with MPN had higher incidence rates of WE, compared to those without MPN. Given the potentially dangerous outcomes associated with WE, physicians who care for patients with MPN should be aware of the risk of WE in this population.

Disclosures
Wu: Sanofi: Employment. Mirza:Sanofi: Employment. Partisano:Sanofi: Employment. Gallagher:Sanofi: Postdoctoral Fellow Other. Vaze:Sanofi: Employment. Zhang:Sanofi: Employment. Gallagher:Sanofi: Employment. Lin:Sanofi: Employment. Juhaeri:Sanofi: Employment.
Biosimilar Erythropoietin Alpha Is Effective As Originator Erythropoietin Alpha Plus Liposomial Iron and B12 and Folates in Patients with Refractory Anemia: A Retrospective Real-Life Approach
Authors: Giordano Giulio , Mondello Patrizia , Tambaro Rosa , Perrotta Nicola , D'amico Fabio , Berardi Giuseppe , Carabellese Bruno , Equitani Francesco 
Published: October 2019

Abstract
Background
Biosimilar drugs have a similar, but usually not inferior although not hidentical effects of old registred drugs. Safety is hidentical to old registred drug.
Aims
Aim of this study is to verify if in MDS patient with refractory anemia biosimilar erythropoietin alpha is not inferior to erythropoietin alpha in terms of safety, efficacy and costs.
Methods
This study is a dicentric, nonrandomized, retrospective study.
Between july 2008 and june 2012, 92 patients affected by refractory anemia were studied.
Median follow-up was 22 months (R3-34). Patients received in group A erythropoietin alpha 40000 IU sc/weekly. In group B patient received biosimilar erythropoietin alpha 40000 IU sc/weekly. In both groups patients received liposomal iron (Sideral®) 14 mg, 2 tablets orally/day calcium levofolinate 7.5 mg/day orally + Vitamin B12: 400 mg/day orally.
In group A median age was 70 years (R63-75), M/F: 18/28; in group B median age was 64 years (R60-70), M/F: 25/21. IPSS was low in 30 patients and int-1 in 13 patients in group A, and low in 32 patients and int-1 in 11 patients, in group B.
Patients with 5q- were excluded from this study.
Median level of haemoglobin was 8.5 g/dl in group A (R8-11) and 9.2 g/dl (R8.5-11.5) in group B.
Results
Group A patients increased Hb level of 1 g/dl after a median time of 5 weeks (R4-9) and after a median time of 4 weeks (R3-8) in group B. No relevant side effects were observed in both groups.
Erythropoietin alpha was reduced in group A because Hb achieved a level > 12g/dl after a median of 12 weeks (R 4-18). Biosimilar erythropoietin alpha was reduced in group B because Hb achieved a level > 12g/dl after a median of 10.5 weeks (R 3-16). In group A maintenance dose was administered with a median of every two weeks (2-4), In group B maintenance dose was administered with a median of every three weeks (2-5). Median cost for every month of erythropoietic therapy was 1536 euros/month (R1240-1850) in group A and 1354 euros/month (R954-1550) in group B. Five patients need transfusion support in group A and seven patients need transfusion support in group B
Summary/conclusion
Biosimilar erythropoietin alpha plus liposomal iron (Sideral®) and B12 and folates support seems to be safe, feasible, probably equally cost-effective and substantially not inferior to classical erythropoietin alpha support in patients affected by refractory anemia. This study needs confifrmation on a larger cohort of patients.

Disclosures
No relevant conflicts of interest to declare.
DNA Methylation-Based Subtype Prediction for Pediatric Acute Lymphoblastic Leukemia (ALL)
Authors: Nordlund Jessica , Bäcklin Christofer , Zachariadis Vasilios , Cavelier Lucia , Dahlberg Johan , Öfverholm Ingegerd , Barbany Gisela , Nordgren Ann , Övernäs Elin , Abrahamsson Jonas , Flaegstad Trond , Heyman Mats , Jónsson Ólafur G , Kanerva Jukka A. , Larsson Rolf , Palle Josefine , Schmiegelow Kjeld , Gustafsson Mats G , Lönnerholm Gudmar , Forestier Erik , Syvänen Ann-Christine 
Published: October 2019

Abstract
We present a method that utilizes DNA methylation profiling for prediction of the cytogenetic subtypes of acute lymphoblastic leukemia (ALL) cells from pediatric ALL patients. We used the methylation status of ~450,000 CpG sites from 546 well-characterized patients with T-ALL or B-cell precursor ALL with the cytogenetic subtypes high hyperdiploidy (HeH), t(12;21)(p13;q22)[ETV6/RUNX1], t(1;19)(q23;p13.3)[TCF3/PBX1], 11q23/MLL-rearrangements, t(9;22)(q34;q11)[BCR/ABL1], dic(9;20)(p13.2;q11.2), and intrachromosomal amplification of chromosome 21 iAMP21[RUNX1 x >3], to design and validate sensitive and accurate DNA methylation classifiers. After repeated cross-validation, a final classifier was derived consisting of only 246 CpG sites that accurately predicted the subtype of >96% of the patients with known subtype. The mean sensitivity and specificity across the subtypes was 0.90 and 0.99, respectively. We then used the DNA methylation classifier to screen for subtype membership of 210 patients with undefined karyotype (normal or no result) or non-recurrent cytogenetic aberrations (“other” subtype). Nearly half (n=106) of the patients lacking cytogenetic subgrouping displayed highly similar methylation profiles as the patients in the known recurrent groups. We verified the subtype of 20% of the newly classified patients by examination of diagnostic karyotypes, array-based copy number analysis, and detection of fusion genes by quantitative PCR and RNA-sequencing (RNA-seq). Using RNA-seq data from ALL patients where cytogenetic subtype and DNA methylation classification did not agree, we discovered several novel fusion genes involving ETV6, RUNX1, and PAX5. The CpG sites constituting the classifiers highlight relevant biological characteristics of otherwise unclassified ALL patients and provide clues to the origin and development of leukemic transformation. Together with response to induction therapy, genetic aberrations are among the most important prognostic factors in pediatric ALL. Our findings indicate that DNA methylation profiling can help clarify some of the heterogeneity in cytogenetically undefined ALL patient groups and can potentially be implemented for diagnostics of ALL and other types of hematological cancers.

Disclosures
No relevant conflicts of interest to declare.
Gallstones in Sickle Cell Disease: A Single Institution Experience
Authors: Coats Thomas , Gardner Kate , Thein Swee Lay 
Published: October 2019

Abstract
Background: There is an increased incidence of gallstones in patients with sickle cell disease (SCD), due to haemolysis. Complications of gallstones include cholecystitis, pancreatitis and cholangiopathy and gallstones can trigger an acute sickle cell crisis. It is not known whether patients with asymptomatic gallstones would benefit from elective cholecystectomy to avoid such complications.
Method: Electronic patient records of all 767 adult SCD patients attending clinic at King’s College Hospital, London between 1st Jan 2003 and 31st Dec 2013 were retrospectively reviewed to identify cases of gallstones. Medical records and steady state blood values were analysed in all those patients with an ultrasound of the biliary tree during this time period.
Results: Amongst the cohort of 767 patients with SCD, 481 (62.7%) were HbSS, 244 (31.8%) HbSC, 35 (4.6%) HbSB+, 6 (0.8%) HbSB0 and 1 (0.1%) HbSHFPH genotype. 43% were male. Mean age at the end of the study period was 36.6 +/- 12.5 years. 344 patients had an ultrasound scan of the biliary tree during the time period of the study. 38 of the 344 patients scanned had had a cholecystectomy prior to 2003. Of the remaining 306 patients with an ultrasound scan, 134 had gallstones identified within the gallbladder. The 134 patients with gallstones comprised 119 (88.8%) HbSS, 12 (9.0%) HbSC, 2 (1.5%) HbSB+ and 1 (0.7%) HbSB0. 39.6% were male. Mean age at the end of the study period was 35.4 +/- 12.2 years.
Of the 134 patients with gallstones identified during the study, 35 developed serious complications directly relating to cholelithiasis (5 pancreatitis, 4 acute cholangitis, 8 choledocholithiasis and 18 isolated cholecystitis) and 8 of these patients required sphincterotomies +/- stone removal with endoscopic retrograde cholangio-pancreatogram.
34 of the 134 patients with gallstones went on to have a cholecystectomy during the 11 year study period. Of these 34, 3 had recorded surgical complications following cholecystectomy (2 bile leaks, 1 hepatic injury). All 3 cases had gallstone-related complications prior to the cholecystectomy.
Discussion: Our findings of cholelithiasis in 134 of the 306 of sickle cell disease patients scanned, is similar to incidence reported in the literature. Notably, we documented a high incidence of complications associated with cholelithiasis. Furthermore, there were higher than expected rates of surgical complications in cholecystectomy undertaken following the development of a complication relating to gallstones. These findings make routine screening for cholelithiasis followed by elective cholecystectomy for positive cases an attractive approach.

Disclosures
No relevant conflicts of interest to declare.
Rivaroxaban Safety Threshold at 30 Ng/ml: Are a PT and/or a Aptt an Alternative to a Specific Test in the Real World?
Authors: Cauchie Philippe , Dierge Laurine , Isabelle Courtois , Alewaeters Alain , Chatelain Bernard 
Published: October 2019

Abstract
The French Working Group on Perioperative Haemostasis (GIHP) published in March 2013 proposals for the management of major bleeding complications and emergency surgery in patients treated with direct oral anticoagulants, and especially the Rivaroxaban. This Direct Oral Anticoagulant (DOA) induces most of clotting tests, including Prothrombin Time (PT) and APTT. This led the GIHP to a degraded proposition for the institutions who does not propose the specific dosage of the molecule. This proposal assumes that the normality of the PT and aPTT exclude most patients with Rivaroxaban level >30 ng/ml. The aim of our evaluation is to verify the real-life practicability of this proposal.
Samples from Rivaroxaban treated patients have been tested with 4 PT reagents: STA Neoplastine R (NeoR), STA Neoplastine CI+, Recombiplastine and Innovin. Four APTT reagents were also tested: STA-PTT automate, STA Cephascreen, Synthasil, Pathrontin SL. All reagents were tested on their specific analysers (STA-R, ACL TOP500, BCS XP). PT reagents were calibrated following manufacturer’s instructions (Stago, Instrumentation Laboratory, Siemens).
Rivaroxaban prolongs PT but reagent’s sensitivity varies considerably and we observed a major individual variability, and this even at low Rivaroxaban concentrations. The NeoR presents the greatest sensitivity, the Innovin the lowest. For a normal PT (>70%), the NeoR alone can validly identify patients with Rivaroxaban level > 30ng/ml (sensitivity 93%). With the other reagents, from 42 to 51% of our patients with normal PT has Rivaroxaban level > 30 ng/ml. Two patients with Rivaroxaban > 150 ng/ml had normal PT with these 3 reagents. None of the APTT reagents has sufficient sensitivity to validly exclude these patients.
In conclusion, none of the PT reagents can estimate Rivaroxaban concentration. The NeoR is the only reagent that allows distinguishing patients with residual Rivaroxaban level. For all other situations, specific test seems mandatory and the development of point-of-care system should be encouraged.

Disclosures
No relevant conflicts of interest to declare.