bluuebunny/crossref_metadata_2025_split huggingface dataset:
Generation of Resistance Cell Lines to AMN107, a New Inhibitor of BCR-ABL and Its Effects on Cell Lines Sensitive and Resistant to Imatinib.
Authors: Mahon Francois-xavier , Lagarde Valerie , Manley Paul W. , Pasquet Jean-Max , Turcq Beatrice , Reiffers Josy 
Published: April 2020

Abstract
Targeting the tyrosine kinase activity of Bcr-Abl is an attractive therapeutic strategy in Chronic Myelogenous Leukemia (CML) and in Bcr-Abl positive Acute Lymphoblastic Leukemia. Imatinib is a selective inhibitor of Bcr-Abl tyrosine kinase and is now used in frontline therapy for CML. However clinical resistance is the main concern using this treatment, mediated by mutations within the kinase domain of Bcr-Abl, amplification of the BCR-ABL genomic locus or other as yet unknown mechanisms. AMN107 (Novartis Pharma AG, Basel, Switzerland) is a synthetic, second generation inhibitor of Bcr-Abl tyrosine kinase. In the current study, we tested AMN107 against different Bcr-Abl positive cell lines such as K562, LAMA84, AR230 or murine Ba/F3 cells transfected with BCR-ABL (BaF/BCR-ABL). In a 4 day proliferation assay (MTS) the dose of AMN107 that inhibited 90% of the cells (IC90) was 0.01 μM which was 100-fold lower than the IC90 of imatinib. In addition, proliferation of imatinib resistant cell lines which exhibited amplification of BCR-ABL was inhibited by 75% in the presence of 0.01 μM AMN107. However, Ba/F3 cells expressing the imatinib resistant BCR-ABL T315I mutant were only inhibited with 10μM of AMN107. Furthermore, K562-R, an imatinib-resistant cell line exhibiting a new mechanism of imatinib resistance (modification of chaperone proteins such as heat shock proteins) was also insensitive to AMN107; the IC90 for AMN107 at day 4 was 1μM versus 4μM for imatinib.
Finally, we investigated potential resistance to AMN107 in Bcr-Abl positive cells. Resistant cell lines were generated after long-term (2 month) gradual dose-escalation exposure to the inhibitor. Up to now, we have obtained four cell lines, AR230-ra, K562-ra, LAMA 84-ra, and BaF/BCR-ABL-ra resistant to 8, 10, 10, and 100 nM of AMN107, respectively. Resistance was defined as the capacity to survive in the continuous presence of doses of AMN107 that kill in three days more than 90% of the parental cells in liquid culture.
Preliminary investigations of AMN107 resistance using western blot and cytometry have shown that only BaF/BCR-ABL-ra overexpressed wildtype Bcr-Abl as we have already reported for imatinib resistance. Studies with these resistant cell lines investigating cross resistance with imatinib and looking for additional mechanisms of resistance are in progress.We conclude that in vitro, AMN107 is more powerful than imatinib in inhibiting the proliferation of BCR-ABL positive cell lines. In addition, we have demonstrated that it is possible to develop resistant cell lines to this new inhibitor of Bcr-Abl.
Effect of the Combination of Imatinib Mesylate (Glivec) and Curcumin in Chronicm Myeloid Leukemia Cell Line.
Authors: Kim Young-Ju , Choi Yong-Hee , Kim Chang Up , Bae Eun Kyung , Kim Byung-Su , Ahn Kwang-Sung , Lee Jong-Seok , Park Seonyang , Kim Byoung Kook , Yoon Sung-Soo 
Published: April 2020

Abstract
Imatinib mesylate (Glivec, Gleevec, STI571) is highly effective in patients with chronic myeloid leukemia (CML). Despite high rates of hematologic and cytogenetic responses to Glivec therapy, the emergence of resistance to Glivec has been recognized as a major problem in the treatment of CML. Those problems leaded us to search for novel therapy, combining curcumin with Glivec. Recently we reported that curcumin induced apoptosis via repression of p-AKT and activation of caspase-3. To investigate whether combination of curcumin with Glivec is more effective than Glivec alone, the expression of apoptosis related genes, such as Bcl-2 family, caspase-3, and its cell cycle proteins, such as p53, p21, cyclin D, cyclin A, cyclin B1 and PCNA were examined by western analysis in K562 chronic myeloid leukemia cell line. Our results showed that Bcl-2, -XL, and Bid were more down-regulated with combined treatment compared with single treatment controls (either Glivec or curcumin alone). Also, the activity of caspase 3 was 2-fold higher than compared with control groups. In western analysis, procaspase 3 was dramatically reduced with combined treatment. Also, cell cycle proteins except p53 were more down-regulated in combined treatment. For further investigation of the genes affected by combination therapy, DNA chip analysis was performed. The expression of candidate genes, which we selected in chip analysis, was confirmed using real-time PCR. Our result indicated that apoptosis related genes were up-regulated and cyclin D1 and cyclin A were down-regulated. Also, several genes (protein phosphatase 2C, ZNF272, protein-tyrosine phosphatase nonreceptor 7 (PTPN7), SOCS2) were more down-regulated in combined treatment, showing 2-fold difference. Those genes may be associated with the progression of chronic myeloid leukemia. We suggest that their down-regulation by combined treatment was effective mainly via the induction of apoptosis. Taken together, our data suggested that combined treatment with curcumin could be effective to overcome the problems of Imatinib mesylate (Glivec, STI571) therapy.
Myeloid Nuclear Differentiation Antigen (MNDA) Protein Expression in Myelodysplastic Syndrome (MDS) Is Reduced in Maturing Myeloid Cells.
Authors: Shults Keith , Head David R. , Flye Leanne , Jagasia Madan H. , Goodman Stacey A. , McClintock-Treep Sara A. , Boulos Fouad I. , Jacobberger James W. , Stelzer Greg , Briggs Robert C. 
Published: April 2020

Abstract
Expression of the human MNDA protein is restricted to hematopoietic cells, and occurs at uniformly high level only in the normal granulocyte/monocyte lineage. Myeloblasts and all later stages of the lineage express MNDA, while stem cells do not (Hum Pathol1999;30:1040). Gene expression analyses of MDS cases have shown that MNDA mRNA is significantly down regulated in familial MDS and in high-risk sporadic MDS cases (Blood2000;100:3553 and Leukemia2004;18:449). We evaluated MNDA protein expression in patients with MDS.
Methods: We directly conjugated Alexa 488 (Molecular Probes) to a rat monoclonal antibody specific for MNDA for use in multiparameter cytometric analysis with CD45/CD34 and DRAQ5 (DNA content). Patients’ (n=17) clinical histories, laboratory data, cytogenetics, and morphology were reviewed for confirmation of diagnoses and blinded to study results. Patients lacking primary marrow disease (n=9) were used as controls. Analysis was performed using Winlist 5.0 software (Verity Software) with DDE links to ModFitLT 3.0 using modifications of published methods along with Esoterix Center for Innovation generated algorithms. Analyses were blinded to clinical results.
Results: The normal dataset revealed MNDA expression in expected cell populations, with only a small subset of CD34+ cells exhibiting MNDA (7.5% on average). Expression of MNDA protein in maturing myeloid forms in normals was consistently greater than in MDS samples (MFI=802 vs. 569 in MDS; MFI=mean fluorescence intensity defined by the linear mean of each population). Six of 17 MDS cases displayed greater than 10% maturing myeloid forms lacking MNDA protein expression (never observed in normals). The % MNDA+/CD34+ cells was identical in the two datasets (7.5% in normal vs. 7.6% in MDS).
Discussion: Our results suggest that MNDA protein expression in MDS patients follows a pattern similar to that in gene expression analyses. We additionally demonstrate that MNDA protein expression at the cellular level is variable in MDS, versus uniform expression in normals; parallel data for MNDA message expression at the cellular level is not available. The finding of MNDA-negative myeloid forms in MDS may have diagnostic utility, and may provide insight into both impaired myeloid differentiation in MDS and the biologic role of MNDA.
A Nation-Wide Survey of Primary Chronic Myelofibrosis in Japan.
Authors: Shimoda Kazuya , Okamura Takashi , Tanimoto Tetsuya , Mizoguchi Hideaki , Omine Mitsuhiro , Niho Yoshiyuki , Harada Mine ,   Reseach Committee for Idiopathic Hematopoietic Disorder
Published: April 2020

Abstract
We retrospectively analyzed 336 patients with primary chronic myelofibrosis from 203 medical institutes in Japan. Average age at onset was 60.7 years. Men were affected 1.4 times more frequently than women. The median survival in 298 patients that could be evaluated was 10.0 years. The factors associated with shorter survival included anemia, leukocytosis/leukocytopenia, thrombocytopenia, and increased blasts in the peripheral blood, and sex (male), age (&gt;60), and the presence of symptoms. Chromosomal abnormalities were found in 58 of 154 patients (38%), although these did not influence survival. In this retrospective study, the median survival in Japanese patients is much longer than that in French patients reported by Dupriez (Blood 1996) (120 mo in Japan versus 42 mo in France). Then we are now doing prospective analysis of newly diagnosed myelofibrosis patients from 1999. In these 5 years, 175 patients were diagnosed as myelofibrosis. Median age was 64 yr (range: 28–86), and men/women ratio is 1.66. Sixty-six patients (38 %) presented anemic symptoms, 19 patients (11 %) showed bleeding tendency. Chromosomal abnormalities were found in 67 of 115 patients (58%). In these five years, 11 of the patients have died. The most frequent causes of death were pneumonia and disseminated infections (7 patients). Three patients developed leukemia.
Investigation of Platelet Functions in Chronic Myeloproliferative Disorders by Optical and Luminesance Method.
Authors: Akay Meltem O. , Mutlu Fezan S. , Gulbas Zafer 
Published: April 2020

Abstract
Background: Bleeding and thrombosis are common causes of morbidity and mortality in patients with myeloproliferative disorders (MPD). Qualitative platelet abnormalities are frequently found in these patients and range from platelet hypofunction as demonstrated by defective invitro platelet aggregation, acquired storage pool disease and/or platelet membrane defects, in addition to enhanced platelet aggregation, increased plasma beta thromboglobulin levels or shortened platelet survival. In this study we aimed to perform platelet aggregation studies by optical method (on platelet rich plasma=PRP) and luminesance method (on whole blood) in chronic myeloproliferative disorders.
Methods:A total of twenty-five patients with MPD (17 chronic myeloid leukemia, 6 polycythemia vera, 2 essential thrombocytosis) were enrolled. Median age was 54,4 (29–76). Platelet aggregation was measured using the optic and luminesance method. The agonists used were adenosine diphosphate (ADP), arachidonic acid (AA) ristocetin and collagen. Platelets were considered to be hyperactive if at least one result (i.e. aggregation or ATP release with one agonist) was above the reference range, and hypoactive if at least one result (i.e. aggregation or ATP release) was below the reference range. Mixed hypo- and hyperactive platelets were considered present when at least one result (i.e. aggregation or ATP release) was below and above the reference range respectively.
Results:Platelet aggregation test results by two methods in myeloproliferative disorders were shown in Table 1. The percent for detection of platelet function abnormality by luminesance method was found to be higher than the optic method and a significant difference was shown between two methods (p&lt;0,05).
Conclusion Our findings suggest that;1. Luminesance platelet aggregation study is more valuable than optic platelet aggregation study for invitro assessment of platelet function in patients with MPD. 2. The use of luminesance platelet aggregation study appears useful to select patients for antiplatelet therapy.
Platelet aggregation test results in myeloproliferative disorders (n=25)
Normal Hypofunction Hyperfunction Mix Total abnormality Luminesance method 1(%4) 4(%16) 8(%32) 11(%44) 24(%96) Optic method 9(%36) 11(%44) 2(%8) 3(%12) 16(%64)
Zap-70 on Bone Marrow Biopsy Is a Good Prognostic Indicator in B-CLL.
Authors: Deliliers Giorgio Lambertenghi , Vener Claudia , Gianelli Umberto , Bosari Silvano , Savi Federica , Cortelezzi Agostino , Somalvico Franco , Soligo Davide , Fracchiolla Nicola S. 
Published: April 2020

Abstract
Bone marrow biopsies (BMB) taken at the time of diagnosis of 108 patients with chronic lymphocytic leukemia (B-CLL) (median age: 62 years, range: 36–81; M/F ratio: 2/1) were collected between 1985 and 2004, and tested immunohistochemically for the expression of ZAP-70 (zeta-associated protein 70), “Anti-ZAP-70 (human), clone 2F3.2 (mouse monoclonal IgG2a)”. There were degrees of ZAP-70 expression: “negative (a)” (58 patients), “weak (b)” (20 patients) and “intense (c)” (30 patients)” in comparison with natural killer (NK) and T-cell ZAP-70 expression (BMB positive when 20% of B-CLL cells express ZAP-70). The cases were analysed separately by three different experts.
In terms of BMB morphology, proliferative centres were 36% in (a) 35% in (b) and 47% in (c); a diffuse pattern: 8.6% in (a), 10% in (b) and 13% in (c); a nodular pattern: 66% in (a), 55% in (b) and 37% in (c): (a) vs (b+c) (p=0.024), (a+b) vs (c) (p=0.014). The different degrees of ZAP-70 expression correlated with two groups of patients according to the Rai and Binet criteria: “0-I-II, A” and “II-III-IV, B, C”. Cases (a) correspond more frequently to the first, and (b+c) to the second (p=0.013); moreover, cases (a+b) correspond more frequently to the first group, and (c) to the second (p=0.0026).
A lymphocyte doubling time (LDT) of less than six months and less than one year was most frequent in (c) (p=0.00044) (p=0.0065). An LDT of less than one year was more frequent in (b+c) vs (a) (p=0,01). Patients receiving no therapy were more frequent in (a) or (a+b) (p=0.00011) (p=0.00045). Patients receiving no therapy or only one line of treatment were more frequent in (a) or (a+b) (p=3.2x10–5) (p=0.0073). Cytogenetic abnormalities were more frequent in (a) or (a+b) (p=0.017) (p=0.022), and complex caryotypes more frequent in (a+b), (p=0.016). The incidence of death was lower in (a+b) than (c) (8% vs 27%) (p=0.041).
Mann-Whitney analysis showed a statistical difference between (a+b) and (c) for the following variables at diagnosis: PLT (p=0.02); WBC (p=0.05); Ly (p=0.055); LDH (p=0.015); B2m (p=0.0005); splenomegaly (p=0.03); and a statistical difference between (a) vs (b+c) for the following variables at diagnosis: PLT (p=0.044); LDH (p=0.006); B2m (p=0.003); percentage of bone marrow infiltrate (p=0.055); CD 38 flow-cytometry (p=0.0009). Kruskall-Wallis analysis of (a) vs (b) vs (c) showed a statistical difference for the following variables at diagnosis: LDH (p=0.01); B2m (p=0.0017); CD 38 flow-cytometry (p=0.0051). These data confirm the prognostic significance of ZAP-70 expression in patients with B-CLL, and its good correlation with clinical and cytogenetic parameters and the Rai-Binet classification.
Normal and Chronic Lymphocytic Leukemia (CLL) Lymphocytes Can Be Distinguished in Flow-Cytometry-Based Automated Differential Cell Counts by Their Positional Parameters.
Authors: Millá Fuensanta , Barberá Angels , Oriol Albert , Navarro Tomas , Juncà Jordi , Rodriguez Lluis , Ribera Josep-Maria 
Published: April 2020

Abstract
Introduction and objectives. Flow-cytometry based automated blood cell counters measure the volume (V), conductivity (C) and light scatter (S) of cells and make available these values for each subpopulation of white blood cells. Although differences in V, C and S among several lymphoproliferative disorders, reactive lymphocytes and normal lymphocytes have been described, the ability to use this this information to detect lymphoproliferative disorders has not been defined.
Methods. We registered VCS data of all consecutive cases of lymphocytosis (lymphocytes &gt; 3.5x10e9/l) detected in routine laboratory testing during 50 non-consecutive days. Lymphocytes were evaluated morphologically, immunophenotypically and by conventional cytogenetics if necessary and the patients were evaluated clinically to document diagnosis adequately. Discriminant analisys modelling was used to assess the ability to predict diagnosis of VCS data.
Results. 202 cases of lymphocytosis were detected in 120 males and 82 females, mean age 53 years (SD 16). Patients were considered to have normal lymphocytes (N=102, 50%), to present a reactive lymphocytosis (N=43, 21%, 20 of them HIV infected patients) or achieved a final diagnosis of lymphoproliferative disorder (N=57, 28%, 43 of them CLL).
The mean volume of CLL lymphocytes was significantly lower than that of normal and reactive lymphocytes (p=0.009) and so was light scatter (p=0.018). When mean VCS values and their standard deviations were included in a discriminant analysis model, a unique function adequately classified 166/189 cases (87%) as CLL versus reactive or normal. Sensitivity and specificity were 73% and 92% respectively and false positive and false negative cases were 26% and 8% respectively. Inclusion of a second discriminant function did not improve significantly the predictive power of the model.
Conclusion, VCS data from flow-cytometry based automated blood cell counters can be used to differentiate CLL from normal or reactive lymphocytes by simple discriminant analysis classification.
CD5 and CD23 Positive Mantle Cell Lymphoma Detected by Flow Cytometry and Confirmed by FISH Study t(11;14).
Authors: Zhang John , Chin David , Anthony Adam , Bolton Heather , Phillips Cheri , Hii Anselm , Chen Sing-Tsung 
Published: April 2020

Abstract
The differential diagnoses of CD5 positive B-cell lymphoproliferative disorders mainly include chronic lymphocytic leukemia/small lymphocytic lymphoma (CLL/SLL) and mantle cell lymphoma. Occasionally large cell and marginal zone lymphomas may also be CD5 positive. An accurate diagnosis effects patient management.
The classical immunophenotype for chronic lymphocytic leukemia/small lymphocytic lymphoma is CD19/CD5/CD23 positive FMC-7 negative cells with dim CD20 and dim light chain expressions, while mantle cell lymphoma is CD19/CD5/FMC-7 positive with bright CD20 and bright light chain expressions. The diagnosis of mantle cell lymphoma is usually confirmed by either immunostain for cyclin D1 or FISH study for t(11;14). In reality, immunostaining for cyclin D1 can be difficult and may show variable results in different laboratories and FISH study may not be readily available. Generally, when it comes to the diagnosis of lymphoma, immunohistochemical positivity of both CD5 and CD23 is almost pathognomic for chronic lymphocytic leukemia/small lymphocytic lymphoma if no fresh tissue is saved for flow cytometry analysis.
Flow cytometry analysis of 44 FISH-confirmed mantle cell lymphomas was reviewed in our lab. Among these, 37 showed the classical immunophenotype of mantle cell lymphoma. However, 7 cases (16%) were positive for both CD5 and CD23. The expression of CD23 varied from dim to bright. When compared to typical CLL, they showed FMC-7 expression and brighter than dim light chain expression. In one case, the light chain expression was dim.
In conclusion, CD23 expression which was thought to be a specific marker for CLL/SLL may also be seen with mantle cell lymphoma. Although FMC-7 expression is seen in all CD23 positive mantle cell lymphomas, bright light chain expression is not universal. We recommend that FISH or immunohistochemical studies for cyclin D1 be performed on CD5/CD19 clonal B cell proliferations with CD23 expression if morphology or immunophenotype is atypical for CLL/SLL.
Establishment of an In Vitro Culture Method for Multiple Myeloma Cells.
Authors: Cohen Yossi , Hardan Yzhar , Nagler Arnon , Zipori Dov 
Published: April 2020

Abstract
In July 2004 we have established an in vitro method for culturing and expansion of human primary myeloma cells in a cytokine free system. This method which was designed to optimize the cell signaling between myeloma cells and their stromal support is easilly reproducible and so far has been applied successfully in 15 of 15 myeloma patients. This progress allows, for the first time, long term maintenance and expansion of the tumor cells with day by day observation of how the myeloma clone propagates, as assessed by morphology, FACS analysis and by the FISH-Bioview system (see figures 1,2 showing a group of myeloma cells after 28 days in culture along with their single FISH signal for known del13q).
The protocol is currently under intensive procedure of standardization for general use.
Applications of this culture system have already yielded several new findings. One, relating to the peculiar nuclear changes appearing in myeloma cells during culture is being presented in detail in another abstract. An additional issue is the ability to manipulate the culture to detect very small numbers of myeloma cells in the range of MRD. Yet another use is in vitro drug assay. Preliminary results have revealed no apparent influence of pamidronate or erythropoietin on myeloma cell expansion up to 28 days in co-culture (4 cases). In contrast, thalidomide and simvastatin had deleterious effect on the stromal support (3 cases) but the former did not eliminate the myeloma cells whereas no myeloma cells were seen with simvastatin after 3–7 days in culture.
Figure Figure Figure Figure
Vascular Endothelial Growth Factor (VEGF) Is a Growth and Survival Factor in Waldenstrom’s Macroglobulinemia.
Authors: Tournilhac Oliver , Le Gouill Steven , Santos Daniel Ditzel , Podar Klaus , Xu Lian , Hatjiharissi Evdoxia , Tai Yu-Tzu , Hunter Zachary , Branagan Andrew , Anderson Kenneth C. , Treon Steven P. 
Published: April 2020

Abstract
Waldenstrom’s macroglobulinemia (WM) is a B-cell disorder characterized by excess of bone marrow (BM) IgM secreting lymphoplasmacytic cells (LPC). In previous studies, we and others have demonstrated that VEGF within the BM micro-environment plays a role in the growth, survival and migration of myeloma plasma cells. Conversely, we demonstrated that BM mast cells (MC), which produce and secrete VEGF (Boesiger, J Exp Med, 1998) support growth and survival of WM LPC (Tournilhac et al, JCO2004; 22:517S). In the present studies we therefore explored the role of VEGF in WM pathogenesis. Using RT-PCR analysis, we detected VEGF receptors R1 (Flt-1) and R2 (flk1/KDR) transcripts in sorted LPC from 4/4 (100%) and 15/16 (94%) WM patients respectively. Moreover, we showed functionality for VEGF as a growth and survival factor in WM. Specifically, recombinant human VEGF (rh-VEGF) at 25 ng/ml induced dose dependent proliferation of sorted LPC from WM patients and prevented serum starvation- induced apoptosis. Inhibition of VEGF signaling pathways both induced apoptosis and blocked mast cell- induced proliferation of sorted WM LPCs in 4 of 6 and 4 of 4 patients, respectively. Besides the known MC-derived WM LPC growth and survival factors CD40 ligand (CD40L) (JCO2004; 22;517S) and B-lymphocyte stimulator protein (BLYS) (Ditzel Santos et al, ASH 2004 submitted), the present study demonstrates also a role of VEGF in WM pathogenesis. We therefore suggest VEGF as an additional therapeutic target in WM treatment regimens.