bluuebunny/crossref_metadata_2025_split huggingface dataset:
Clinical Significance of FLT3 Internal Tandem Duplication in Patients with Acute Myeloid Leukemia Who Underwent Allogeneic Bone Marrow Transplantation.
Authors: Lee Je-Jung , Lee Yu-Ra , Kim Hee-Nam , Kim Nan-Young , Park Kyeong-Soo , Yang Deok-Hwan , Kim Yeo-Kyeoung , Kim Hyeoung-Joon 
Published: April 2020

Abstract
Since the introduction of high-dose cytarabine therapy, there are still controversies for the role of allogeneic or autologous stem cell transplantation (SCT) in acute myeloid leukemia (AML), but the SCT, especially allogeneic, has a benefit in AML patients with intermediate risk. However, it is needed to investigate for a new prognostic factor, in addition to the cytogenetics, to the further stratifying approach. Internal tandem duplications (ITD) within FLT3 are present in 20–30% of patients with AML and have a prognostic implication in the disease. However, there was no report in AML patients who underwent allogeneic bone marrow transplantation (BMT). In this study, we evaluated the prognostic relevance of FLT3/ITD in 42 patients with AML who underwent allogeneic BMT. FLT3/ITD mutations were studied on bone marrow samples at diagnosis using PCR. As a baseline study, we firstly analyzed the incidence of FLT3/ITD in BM samples at diagnosis of 214 patients with AML. Of the patients, FLT3/ITD were found in 68 patients (31.8%). In this study, 64% of the patients who underwent allogeneic BMT were positive for FLT3/ITD mutations. Methologically, FLT3/ITD detections by melting curve analysis showed higher sensitivity (66.7%) than those by gel electrophoresis (61.9%). There were no significant differences in the engraft periods and the incidence of acute and chronic GVHD according to the presence of FLT3/ITD mutations. When we analyze patient’s survival according to the presence or absence of FLT3/ITD, the probability of overall survival (OS) at 3 years in the AML patients with FLT3/ITD tended to be shorter than those lacking FL3/ITD (53.9 ± 9.9% vs. 60.6 ± 14.7%, P=0.46). In addition, the probability of disease free survival (DFS) at 3 years in the AML patients with and without FLT3/ITD was 57.3±10.3% and 85.7±13.2%, respectively (P=0.046). Among the cytogenetic risk group, low-risk and high-risk groups were no significant differences according to FLT3/ITD despite of the limited number of patients studied. However, patients with intermediate-risk were significantly shorter DFS in the presence of FLT3/ITD than those in the absence of FLT3/ITD (P=0.048). These findings suggest that the presence of FLT3/ITD mutations is a poor prognostic factor for disease free survival in AML patients, especially with cytogenetically intermediate-risk, who underwent allogeneic BMT and melting curve analysis to detect the presence of FLT3/ITD mutations is a useful tool with high sensitivity.
Fusion of SFQP to ABL in a Patient with a T(1;9)(p34;q34) and Acute Lymphocytic Leukemia.
Authors: Cross Nicholas C.P. , Chase Andrew J. , Drachenberg Milton , Roberts W. Mark , Finklestein Jerry Z. , Grand Francis H. 
Published: April 2020

Abstract
We have investigated a child who presented with pre-B ALL and an acquired t(1;9)(p34;q34). BCR-ABL was not detected by RT-PCR or FISH analysis, however FISH did indicate that the ABL gene at 9q34 was disrupted. To identify the putative partner locus in this case, a modified 5′RACE strategy was employed that selected against normal ABL transcripts. Several clones were recovered in which ABL was fused to SFPQ (also known as PSF), a gene mapping to 1p34 that encodes a polypyrimidine tract-binding protein-associated splicing previously identified as a fusion partner of the helix-loop-helix transcription factor TFE3 in papillary renal cell carcinomas. Both SFPQ-ABL and reciprocal ABL-SFPQ transcripts were detectable by RT-PCR, and disruption of these two genes was further confirmed by amplification and sequencing of the forward genomic breakpoint. SFPQ-ABL, the likely oncogenic product, is predicted to encode a protein that retains the coiled coil domain of SFPQ and the entire tyrosine kinase domain and C-terminal sequences of ABL. The breakpoint in ABL was downstream of that seen for other ABL fusion genes and the chimeric protein is predicted to lack the ABL-encoded SH3 domain and part of the SH2 domain. The patient was treated according to the Children’s Cancer Group Protocol 1961 and subsequently received augmented BFM therapy with doxorubicin and double delayed intensification. He achieved complete remission but suffered extramedullary testicular relapse at 4.5 years. Following orchiectomy and intensive chemotherapy he remains in complete remission more than 6 years after diagnosis. We conclude that SFPQ-ABL is a novel fusion gene associated with ALL. Although the patient here responded to conventional chemotherapy, SFPQ-ABL is likely to be sensitive to imatinib and use of this agent might be considered in further cases.
Hypergranular Non-M3 Acute Myelogenous Leukemia: A Potential Pitfall in Flow Cytometric Diagnosis.
Authors: Gorczyca Wojciech , Dong Henry , Weisberger James , Liu Zach 
Published: April 2020

Abstract
Background
Flow cytometry (FC) is an important tool in the diagnosis, subclassification and clinical management of acute leukemias. CD45 vs. orthogonal or side scatter (SSC) gating criteria allows easy identification of non-M3 blasts, which form a coherent population in the low SSC/moderate CD45 blast gate. The vast majority of acute myeloid leukemias (AML) of non-M3 type express pan-myeloid antigens CD13 and CD33, HLA-DR, blastic markers CD34 and CD117. AML M3 (acute promyelocytic leukemia, APL) often lacks CD34 and HLA-DR expression, and has characteristically high SSC/moderate CD45 expression, which overlaps with the granulocyte gate, but is CD117-positive. We report nine cases of non-M3 AML (hypergranular AML) with increased side scatter, moderate CD45 expression, and lack of expression of blastic markers, which mimic APL.
Design
Multiparameter 4-color FC analysis was performed on peripheral blood and/or bone marrow aspirate samples using the following antibody panel: CD2, CD3, CD4, CD5, CD7, CD8, CD10, CD11b, CD11c, CD13, CD14, CD16, CD19, CD20, CD33, CD34, CD38, CD45, CD56, CD64, CD79a, CD117, TdT, and HLA-DR. Fresh bone marrow aspirate and/or peripheral blood films and cytospin preparations from FC samples were stained with Wright Giemsa and reviewed in each case.
Results
Instead of a coherent population of blasts in the low SSC/moderate CD45 blast gate, the blasts in all 9 cases displayed increased granularity, which placed them in the granulocyte gate on FC display. The immunophenotype was as follows: HLA-DR+ in 5/9, CD4+ in 6/9, CD10+ in 1/9, CD11b+ in 4/9, CD11c+ in 8/9, CD13+ in 8/9, CD14+ in 0/9, CD16+ in 0/9, CD19+ in 0/9, CD33+ in 9/9, CD34+ in 0/9, CD38+ in 9/9, CD56+ in 5/9, CD64+ in 9/9 and CD117+ in 0/9 cases. The cytologic preparations revealed type II and type III cytomorphology in all 9 cases; none contained Auer rods.
Conclusion
Hypergranular non-M3 AML is difficult to diagnose by FC by pattern recognition because the blasts overlap with maturing granulocytes by SSC/CD45 gating criteria, and are not located in the blast gate. In this respect, they mimic APL and regenerative marrows, including those following G-CSF therapy. Correlation with cytomorphology and the expression of CD10, CD11b, CD16 and CD56 helps to differentiate between hypergranular AML, benign processes, APL, and MDS. Specifically, benign maturing myeloid cells show graduating positivity for CD10, CD11b and CD16, and are negative for CD56. This is the inverse pattern of the majority of cases of hypergranular AML. These cases differ from APL by expression of HLA-DR (in a majority of cases), lack of CD117, and different cytomorphology. Myelodysplastic syndromes, which may display similar aberrant myeloid expression of CD10, CD11b, CD16 and CD56, are most often low in SSC, and blastic cells will be &lt;20% cytologically on smear review.
Interactions between the Protein Kinase C (PKC) and the Calcineurin-Phosphatase (CP) Pathways in Differentiation of HL-60 Promyeloblasts into Dendritic Cells (DC).
Authors: Li Qian , Ozer Howard , Kharfan-Dabaja Mohamed A. 
Published: April 2020

Abstract
Continuing success in identifying tumor-specific antigens supports further implementing and refining immunologic strategies to target cancer more specifically. Intracellular signals responsible for differentiation and lineage commitment of pluripotent hematopoietic stem cells remain largely undefined. We have previously reported that blockade of PKC with 2-[1-(3-Dimethylaminopropyl)-1H-indol-3-yl]-3-(1H-indol-3-yl)-maleimide, also known as bisindolylmaleimide- I inhibits morphologic, phenotypic and functional differentiation of HL-60 myeloid blasts into DC by calcium mobilization with the calcium ionophore A23187. To further analyze the interdependence between the PKC and the CP pathways, we evaluated whether blockade of the CP pathway with cyclosporine A (CSA) affects the ability of the PKC agonist phorbol 12-myristate 13-acetate (PMA) to induce DC differentiation of HL-60 myeloblasts. We pre-treated the HL-60 myeloblasts with the CP inhibitor CSA at a dose of 50 ng/ml x 24 hrs, followed by the PMA at 10 ng/ml x 24 hrs or 48 hrs. On a pilot dose evalaution, we noted that a majority of HL-60 myeloblasts pre-treated with CSA x 24 hrs died upon exposure for ≥48 hrs PMA afterwards; therefore, PMA x 24 hrs was selected. Controls consisted of media, CSA (50 ng/ml x 24 hrs) and PMA (10 ng/ml x 24 hrs). We noted that pre-treatment of HL-60 myeloblasts with CSA→PMA, failed to inhibit development of DC-morphologic features; and on phase-contrast microscopy they were similar to those treated with PMA alone. Media and CSA (alone) treated myeloblasts did not develop DC morphology. Addidtionally, we evaluated the ability of HL-60 derived DC to activate allogeneic T-cells by CD69 activation. We noted that blasts treated with PMA upregulated expression of CD69 in allogeneic T-cells suggesting T-cell activation. Interestingly, pre-treatment of HL-60 blasts with CSA→PMA did not prevent T-cell activation (similar to PMA alone). Media and CSA (alone) treated promyeloblasts failed to induce T-cell activation. Our findings suggest that inhibition of the calcineurin-phosphatase pathway with CSA does not inhibit morphologic and functional differentiation of HL-60→ DC by direct PKC signaling with PMA.
Coagulopathy in HLA-DR Antigen-Negative Acute Myeloid Leukemia.
Authors: Uchiumi Hideki , Matsushima Takafumi , Yamane Arito , Handa Hiroshi , Irisawa Hiroyuki , Doki Noriko , Sakura Tohru , Tsumita Yuki , Jinbo Takahiro , Tsukamoto Norifumi , Karasawa Masamitsu , Murakami Hirokazu , Miyawaki Shuichi , Nojima Yoshihisa 
Published: April 2020

Abstract
Background: HLA-DR antigen is present on hematopoietic progenitors and granulocyte/monocyte, erythrocyte and megakaryocytic precursors but absent at the promyelocytic stage during myeloid cell maturation. In accordance with this, majority of promyelocytic leukemia (APL) cells were negative for HLA-DR. Meanwhile, some of non-APL acute myeloid leukemia (AML) cells is found to express HLA-DR. However, the clinical significance of HLA-DR antigen on AML cells is currently unclear.
Purpose: We sought to determine the prevalence and clinical characteristics of negativity in HLA-DR expression by retrospectively analyzing 181 consecutive patients with de novo adult AML.
Patients and Methods: AML patients examined in the current study (aged 15–86 years) had been diagnosed between August 1995 and July 2004, and categorized to M0 (8 patients), M1 (35), M2 (74), M3 (20), M4 (25), M5 (15), and M6 (4), based on the FAB classification. Median follow-up time was 19.3 months. Phenotypic analyses of leukemic cells were performed using CD45 gating methods. HLA-DR-negative AML was defined as HLA-DR expression less than 20% of cells in the CD45 leukemic cell gate.
Results: Among 181 patients, HLA-DR antigens were not detected on AML cells from 46 patients; 20 with APL and 26 with non-APL (non-APL/DR(−)), the latter of which included M0 (2 patients), M1 (15), M2 (7), M4 (2). Leukemic cells from other non-APL patients were HLA-DR-positive (non-APL/DR(+)). None of non-APL/DR(−) patients had t(15;17) nor PML/RARa rearrangement on cytogenetic analysis. Twenty out of 26 patients with non-APL/DR(−) had normal chromosome, and 6 had abnormal karyotypes. In the non-APL/DR(−) group, various degrees of nuclear folding, convolution, or lobulation were observed in 9 patients. Although treatment response and overall survival rate were similar in the three groups (APL, non-APL/DR(−), and non-APL/DR(+)), both FDP levels at diagnosis (57.3 vs 13.2, p&lt;0.05) and maximal FDP levels (232.6 vs 43.8, p&lt;0.01) were significantly higher in non-APL/DR(−) compared with non-APL/DR(+). The maximal FDP levels in the non-APL/DR(−) patients were comparable to those in the APL patients. FDP levels greater than 40 mg/ml were significantly more prevalent in the non-APL/DR(−) than in the the non-APL/DR(−) group. Logistic regression analysis demonstrated that low HLA-DR expression was an independent risk factor for FDP &gt; 40 mg/ml.
Conclusion: Our study suggests that AML with negative HLA-DR antigen tend to be associated with abnormality in coagulation and fibrinolysis even if they are genetically non-APL. We propose that more attention should be paied for HLA-DR expression to avoid a devastating coagulopathy which carries a high risk of mortality unless specifically addressed.
Multi-Dimensional Flow Cytometric Analysis of Acute Myelomonocytic Leukemia: Evaluation of Disease Complexity and Effects on Host Hematopoiesis.
Authors: Katz Ben-Zion , Kam Zvi , Deutsch Varda , Bar-On Shoshana , Ben-Tal Ofira , Cohen Nir , Naparstek Elizabeth , Geiger Benjamin , Zamir Eli 
Published: April 2020

Abstract
The study of normal or malignant hematopoiesis requires the analysis of heterogeneous cell populations using multiple morphological and molecular criteria. Flow cytometry has the capacity to provide multi-parameter information of large hematopoietic cell populations, utilizing various combinations of over 200 molecular markers (CDs). However, current flow cytometry analyses are based on serial gating of two-parametric scatter plots - a process that is inherently incapable to discriminate all subgroups of cells in the data. Here we studied the cellular diversity of normal and leukemic bone marrow samples using multi-dimensional cluster analysis of six-parametric flow cytometry data (CD13, CD33, CD34, CD45, forward scatter and side scatter), focusing mainly on the myeloid lineage. Twenty-three subclasses of cells were identified within normal bone marrow samples, many of them inseparable even when examined at all possible two-parametric scatter plots. Seven subclasses of cells expressing the immature CD34 marker were identified. The multi-dimensional analysis could identify the hematopoietic progenitors, and readily distinguish this population from other types of immature cells. We further addressed the compositional variability of bone marrow samples by designing a classifier that assigns cell samples to subclasses, based on robust six-dimensional characteristics. We then applied this classifier to assess the population diversity in nineteen samples of acute myelomonocytic leukemia (AML M4). The myeloid classifier analyzed simultaneously normal cellular populations (e.g. lymphocytes, garnulocytes), and the diverse populations of AML cells. Most of the AML M4 samples contained an immature population assigned to a myeloid immature subclass distinct from the normal hematopoietic progenitors. Nearly 50% of the AML M4 samples contained a combination of immature myeloid cells and a significant population of immature monocytes. The presence of immature monocytic sub-population correlated with lower leukemia cell burden, and higher platelet counts. In summary, our data indicate that AML M4 may be separated into two distinct types, one restricted to an immature myeloid state, and the other with limited monocytic differentiation capacity. The sub-population composition of the leukemia may affect normal hematopoiesis. Multi-dimentional analysis using the myeloid classifier provides a novel method to accurately define both the increasing numbers of hematopoietic cellular markers and sample heterogeneity in AML. This method should further expand our ability to study normal hematopoiesis, and to identify and monitor hematopoietic diseases.
Ligation of CD44 by A3D8 Monoclonal Antibody Induces Terminal Differentiation of THP-1 Leukemia Cells by Promoting Cross-Talk between Extracellular Signal-Regulated Kinase and Phosphoinositide-3 Kinase/Akt Signaling Pathway.
Authors: Min Yoo-Hong , Kim Ji Yeon , Jeung Hoi Kyung , Lee Seung Tae , Cheong June-Won 
Published: April 2020

Abstract
Ligation of CD44 with anti-CD44 monoclonal antibody A3D8 induces terminal differentiation of human leukemia cells. However, the underlying molecular mechanisms remain largely unknown. In this study, we examined the importance of MEK/ERK, p38 MAPK, and phosphatidylinositol 3-kinase (PI3-K)/Akt pathway in A3D8-induced monocytic differentiation of THP-1 leukemia cells. THP-1 cells displayed cytologic changes typical of mature monocytes and an increased expression of monocyte-specific antigen CD14 (49% ± 4%) and of myeloid-specific antigen CD11b (68% ± 6%) after 3 days of A3D8 treatment. The level of CD15 was also increased. The increase in the expression of these differentiation antigens was dose- and time-dependent. We found that CD44 ligation with A3D8 led to rapid and sustained activation of the essential kinases in the extracellular signal-regulated kinase pathway such as phospho-Raf-1, phospho-MEK1/2, and phospho-ERK1/2. However, the total levels of these kinases were not affected during the course of differentiation. Ser473 Akt phosphorylation was also observed shortly after the cells were exposed to A3D8 and sustained thereafter. In contrast, the phosphorylation of p38 mitogen-activated protein kinase (MAPK) and C-Jun N-terminal kinase were not increased by CD44 ligation. Pretreatment of cells with 20mM of MEK1 inhibitor PD98059 abrogated the A3D8-induced activation of MEK1/2 and ERK1/2 and potently inhibited the THP-1 differentiation. However, a p38 MAPK inhibitor SB203580 did not abolished the CD44 ligation-induced terminal differentiation of THP-1 cells. Pretreatment of cells with PI3-K inhibitor LY294002 resulted in a nearly complete inhibition of A3D8-induced terminal differentiation. Overexpression of dominant-negative Akt also reduced the A3D8-induced THP-1 differentiation, suggesting that A3D8-induced THP-1 differentiation is coupled to PI3-K/Akt activation. Surprisingly, the pharmacological inhibition of PI3-K blocked not only A3D8-induced Akt activation but also A3D8-induced activation of Raf, MEK and ERK pathway. By contrast, pretreatment of cells with PD98059 did not inhibit the A3D8-induced Akt activation, suggesting that PI3-K/Akt is the upstream of Raf/MEK/ERK pathway in A3D8-induced terminal differentiation of THP-1 leukemia cells. Taken together, our findings demonstrated that the cross-talk between the activation of A3D8-inducible PI3-K/Akt pathway and the Raf/MEK/ERK pathway in THP-1 cell exists, and plays a critical role during CD44 ligation-induced THP-1 differentiation.
In Vitro Studies of Bortezomib with Daunorubicin and Cytarabine:Sequence of Administration Affects Leukemia Cell Chemosensitivity.
Authors: Attar Eyal C. , Learner Emily , Amrein Philip C. 
Published: April 2020

Abstract
Induction chemotherapy with cytarabine and an anthracycline has been a standard treatment of newly diagnosed AML for the past 20 years. While the complete remission rate is high, only 20–40% of patients have prolonged leukemia-free survival and thus novel treatments are required. Bortezomib represents a first-in class proteasome inhibitor with activity in a variety of hematologic malignancies. Recent evidence has demonstrated synergistic cytotoxicity between the proteasome inhibitor, MG-132, and the anthracycline, idarubicin, in primitive leukemia cells.(Guzman et al. Proc Natl Acad Sci U S A 99 2002 16220-5) In order to determine if adding bortezomib to the conventional antileukemic therapies daunorubicin and cytarabine enhances cellular cytotoxicity, we conducted in vitro studies on the acute myeloid leukemia cell line, KG-1. Proliferation assays demonstrated that single-agent daunorubicin or single-agent cytarabine impaired proliferation to a greater extent than when either agent was simultaneously combined with bortezomib. Reasoning that treatment with bortezomib resulted in cell cycle arrest and protection from cycle-dependent agents, we treated cells with bortezomib and observed an accumulation of cells in the G2/M phase of the cell cycle, supporting our hypothesis. We then sought to determine whether sequential administration of bortezomib and chemotherapy affected leukemia cell proliferation. Cells were treated with bortezomib either 24 hours before or after idarubicin or cytarabine. Surprisingly, pretreatment of cells with bortezomib followed by chemotherapy resulted in increased proliferation relative to chemotherapy alone while initial treatment of cells with chemotherapy followed by bortezomib resulted in enhanced cytotoxicity. In conclusion, these data indicate that in in vitro leukemia cell cultures the timing of administration of bortezomib relative to conventional anti-leukemic agents critically affects cytotoxicity.