bluuebunny/crossref_metadata_2025_split huggingface dataset:
Absence of JAK2 V617F Activating Mutations in Children with Acute Megakaryoblastic Leukemia with and without Down Syndrome.
Authors: Zwaan Christian M. , Broekhuis Mathilde J.C. , Langebrake Claudia , Goemans Bianca F. , Kaspers Gertjan J.L. , Pieters Rob , Reinhardt Dirk , den Boer Monique L. 
Published: April 2020

Abstract
Activating mutations at codon 617 of the Janus-2 tyrosine kinase (JAK2 V617F) have recently been described in hematological malignancies. In adult acute myeloid leukemia (AML), the reported frequencies vary, and JAK2 V617F mutations have mainly been detected in secondary AML following a myeloproliferative disorder. In adult de novo AML, the mutation was less frequent, and detected in 2/11 (18%) acute megakaryoblastic leukemia (FAB M7) samples (Jelinek et al., Blood 2005), and occasionally in other FAB-types.
This prompted us to analyze a cohort of pediatric AML FAB M7 samples for this particular mutation. In children, at least 3 different subsets of AML M7 can be identified: infants with AML M7 characterized by t(1;22)(p13;q13), older children with random cytogenetic aberrations, and myeloid leukemia of Down syndrome (DS ML). DS ML is often preceded by transient myeloproliferative disease (TMD), hence we also screened TMD samples to detect whether JAK2 V617F mutations would be involved in clonal evolution from TMD to DS ML. To exclude germ-line mutations in DS, we tested normal mononuclear bone marrow cells (NBMC) from children with DS. These NBMC were obtained from a sternal aspirate from children undergoing cardiac surgery, after informed consent was obtained. Genomic DNA was harvested from leukemic cells, and JAK2 exon 12, including the intron-flanking regions, was amplified and sequenced to screen for the JAK2 V617F mutation. As a positive control for the JAK2 V617F mutation, we used HEL 92.1.7 cells (an erythroleukemic cell line). In a dilution experiment we could still detect the mutation, using direct sequencing, if 10% HEL/JAK2 mutated cells were mixed with 90% wild-type control cells. We tested 49 samples, comprising of 9 NBMC, 11 TMD, 14 DS-ML M7, 11 non-DS AML M7 and 4 relapsed non-DS AML M7 samples (including 2 initial diagnosis-relapse pairs). The median age of the TMD cohort was 3 days, for DS-ML children 1.9 years (range 0.9–3.8 yrs), and for non-DS AML 1.5 years (range 1.2–13.7 yrs). The median white blood cell count for TMD was 25.8x109/l, for DS-ML 13.8x109/l, and for non-DS AML 12.4x109/l. Cytogenetic data were available in 5/11 non-DS AML cases only, which showed no cases with a t(1;22). No JAK2 V617F mutations were detected in any of the clinical samples. We conclude that the role of JAK2 V617F mutations in pediatric DS and non-DS acute megakaryoblastic leukemia is limited at best. However, we were not able to screen the subgroup of non-DS AML cases with t(1;22).
Tumor Marker CA 15-3 in Hematological Malignancies.
Authors: Saouli Zoi , Papadopoulos Athanasios , Kaiafa Georgia , Girtovitis Fotios , Charisopoulos Georg , Kontoninas Zisis , Zantidis Anestis , Andreadis Charalambos 
Published: April 2020

Abstract
Introduction: CA 15-3 is a glycoprotein expressed in several adenocarcinomas, especially of the breast. It is used to detect recurrent or metastatic disease. Elevated levels can also be found in adenocarcinomas of the ovary, lung, pancreas, and colon, and are also related to benign breast or ovarian disease, endometriosis, hepatitis, pregnancy and lactation. To our knowledge with the exception of multiple myeloma, there are no references for the significance of the CA 15-3 in hematological malignancies.
Aim of our study was to evaluate the levels of CA 15-3 in patients with various hematological malignancies.
Material and Methods: 84 patients with hematological malignancies were tested: MDS 27 pts, non Hodgkin lymphoma 12 pts, Hodgkin’s lymphoma 3 pts, chronic lymphocytic leukemia 13 pts, acute leukemia: 7 pts, multiple myeloma 8 pts, chronic myeloid leukemia 3 pts and other chronic myeloproliferative disorders 11 pts. 55% of the patients were men with average age of 55 (17–87) and 45% were women with average age of 52 (35–64). A group of 45 healthy volunteers’ blood donors was also tested. Immunoradiometric assay (IRMA) has been used to determine the circulating levels of the CA 15-3 marker.
Results:
None of the healthy volunteers had elevated Ca 15-3 levels. Among the 84 patients, 31 (36,9%) had elevated levels of CA 15-3. In MDS 10/27 pts, in Non Hodgkin Lymphoma 5/12 pts, in Hodgkin Lymphoma 1/3 pts, in Chronic Lymphocytic Leukemia 5/13 pts, in Multiple Myeloma 4/8 pts, in acute leukemia 1/7 pts and in other myeloproliferative disorders 4/11 pts. The CA 15-3 levels in hematological patients were higher than the ones in the healthy group. The difference was statistically significant (p &lt;0.0001), (Table 1). 31 patients who where either untreated or had recurrent disease (subgroup a), had elevated CA 15-3 levels (36,8 ± 8,9 U/ml) while the rest 53 patients who were under therapy or were in remission (subgroup b) had normal levels (14,3 ± 5,2 U/ml). A statistically significant difference between the two subgroups was observed, (p &lt;0.0001), (Table 2). The low risk MDS patients (RA, RARS) had normal level of CA 15-3, while the high risk MDS patients (RAEB, RAEB-t, CMML) had high levels of CA 15-3.
Conclusions: CA 15-3 can be an indicative marker of the activity of a hematological malignancy. Also, it might be of value in monitoring hematological diseases and response to therapy.
Table 1 Ca 15-3 (U/ml) Patients (total, n=84) 23 ± 12,3 Normal Subjects (n=45) 14,1 ± 4,6 p &lt; 0.0001 Table 2 Ca 15-3 (U/ml) Subgroup a 36,8 ± 8,9 Subgroup b 14,3 ± 5,2 p &lt; 0.0001
ASPP2 Haploinsufficiency Promotes Tumor Formation in a Mouse Model.
Authors: Kampa Kerstin M. , Acoba Jared D. , Chen Dexi , Beemer Kelly , Gay Joel , Zhu Zhiyi , Padiernos Emerson , Fan Alice , Felsher Dean , Corless Christopher , Naumovski Louie , Lopez Charles D. 
Published: April 2020

Abstract
ASPP2 interacts with the tumor suppressor protein p53 and promotes damage-induced apoptosis in part through stimulation of p53-mediated apoptosis. We have previously demonstrated that low ASPP2 levels correlate with poor clinical outcome in patients with diffuse large B-cell lymphoma treated with anthracycline-based chemotherapy. Moreover, reduced ASPP2 expression has been demonstrated in other tumor types. These findings led us to hypothesize that ASPP2 may function as a tumor suppressor. To further explore this, we targeted the ASPP2 allele in a mouse by homologous recombination using a knockout vector that replaced exons 10–17 with a neoR gene. Two separate ES clones were used for blastocyst injections to generate several chimeras that were used to generate ASPP2 heterozygous mice. ASPP2+/− mice appear developmentally normal and reproduce. However ASPP2−/− mice could not be generated. Genotype analysis as early as Ed 6.5 did not detect ASPP2−/− embryos---which implies an early embryonic lethal defect in the homozygote. ASPP2+/− (n=135) and ASPP2+/+ (n=63) sibling mice were observed for spontaneous tumor formation. Overall median tumor-free survival was 117 weeks in the ASPP2+/− mice verses 125 weeks in the ASPP2+/+mice (p = 0.035 log-rank test). Overall tumor incidence (at 115 weeks) for ASPP2+/− and ASPP2+/+ mice was 43% and 22%, respectively. The incidence of tumor types, from all tumors detected, was similar between ASPP2+/− and ASPP2+/+ mice: 34% versus 33% (lymphoma), 18% versus 14% (sarcoma), and 47% versus 52% (carcinoma), respectively. Compound p53+/−;ASPP2+/− mice did not exhibit accelerated tumor formation relative to p53+/−;ASPP2+/+ mice. Additionally, a tet-Myc:ASPP2+/− lymphoma mouse model did not exhibit accelerated lymphomagenesis. However, preliminary data suggests that ASPP2+/− mice may have an increased incidence of irradiation-induced leukemia/lymphoma when compared to ASPP2+/+ mice, and confirmatory studies are ongoing. In response to ionizing radiation, doxorubicin, or serum-starvation, preliminary analysis reveals a G0/G1 checkpoint defect in ASPP2+/− MEFs compared to ASPP2+/+ MEFs. Our results provide in vivo evidence that ASPP2 can function as a tumor suppressor. Further studies are underway to determine the mechanism of this observation.
Synergistic Anticancer Effects between Curcumin and Doxorubicin in Drug Sensitive and Resistant Human Pediatric Leukemic Cell Lines.
Authors: Ramachandran Cheppail , Nair P.K. Raveendran , Escalon Enrique , Melnick Steven J. , Khatib Ziad 
Published: April 2020

Abstract
Curcumin (diferuloyl methane), the yellow colored chemopreventive agent from turmeric has anti-tumor, anti-inflammatory and anti-oxidant effects. The synergistic effect of curcumin and doxorubicin on cytotoxicity in leukemic cell lines was investigated preclinically in vitro for their eventual use in combination therapy in pediatric patients. Curcumin is cytotoxic to both sensitive (CEM) and resistant (CEM/VLB) cell lines with an IC50 value of 12 μM. Multi-drug resistant (MDR) cell line (CEM/VLB) showed 36.4-fold doxorubicin resistance based on DOX IC50 values of 0.14 μM and 5.1 μM for CEM and CEM/VLB cell lines. Analysis of cytotoxicity data by CalcuSyn program showed that curcumin and doxorubicin combination treatments are synergistic with combination index values less than 1 at IC50 (CEM=0.82; CEM/VLB =0.81) and IC75 (CEM=0.60; CEM/VLB=0.83) levels. Curcumin and doxorubicin induces G2/M arrest and apoptosis in tumor cells and the apoptosis data correlated with cytotoxicity values in both sensitive and resistant cell lines. The combination index values on the percentage of apoptosis induced by curcumin and doxorubicin combination treatments were less than 1 indicating the synergistic effect of both these agents on apoptosis. Doxorubicin treatment up regulated NF-κB activity in both CEM and CEM/VLB cell lines, although the level of up regulation is higher in CEM (2-fold) than CEM/VLB (1.5-fold) cell lines. Curcumin, on the other hand, is anti-inflammatory because of its significant inhibitory effect on NF-κB activity. Curcumin inhibited 80% and 50% of NF-κB activity in CEM and CEM/VLB cell lines in 24 h. These results showed that curcumin can be used either as a single or adjuvant agent along with doxorubicin because of the synergistic effects on apoptosis and cytotoxicity.
Study on RNA Interference Reversing Multidrug Resistance in K562/A02 Resistant Leukemia Cell.
Authors: Gong Yuping , Gu Ling , Liu Ting 
Published: April 2020

Abstract
Multidrug-resistance (MDR) is a major hindrance to successful chemotherapy. Among different mechanisms responsible for MDR, cellular over-expression of P-glycoprotein (Pgp) plays an important role. Pgp is an energy dependent drug efflux pump that effectively transports a broad of commonly used chemotherapeutic drug out of the cells and leads to resistance to the chemotherapeutic drugs. Use of modulators is main approach to overcome MDR. Although, in vitro and in vivo preclinical studies have yielded encouraging results, the clinical reversal of resistance by common modulators such as verapamil, quinine, and cyclosporine A has been difficult due to the toxic side effects. Antisense technology offers an alternative approach to overcome MDR. RNA interference (RNAi) is the best one of all antisense technologies. RNAi acts through a post-transcriptional targeting of mRNA for degradation, resulting in sequence-specific post-transcriptional gene silencing. Short hairpin RNA (shRNA) vectors can be transfected into the cells to stably express siRNA. In our preliminary study, eukaryotic shRNA expression vectors aimed at mdr1 mRNA target sequences were cloned and transfected into drug resistance cell line K562/A02 by liposome-induced gene transfection. The mdr1 mRNA was identified by real time RT-PCR, the function of P-gp was measured by a daunorubicin (DNR) efflux assay, and the sensitivity of cell lines to doxorubicin (ADM) was detected by an MTT assay. Our results showed that two mdr1-targeted shRNAs could down-regulate mdr1 mRNA and P-gp expression, with mdr1 mRNA reduced by 86% and 88%. The intracellular DNR increased after RNAi treatment, with the daunorubicin efflux ratio at 60min were 13% and 22%, compared with control (40%~45%) (P&lt;0.05). The MTT assay demonstrated the relative reversing efficiency to doxorubicin to be 84% and 77%.
Conclusion: RNA interference can effectively reverse mdr1-mediated multidrug resistance in resistant leukemic cell line.
Assessing DNA Damage Ex Vivo Induced by Chemotherapy Targeting Topoisomerase Inhibitors in Acute Leukemias.
Authors: Halicka Dorota H. , Ozkaynak Fevzi M. , Seiter Karen , Kajstura Malgorzata A. , Liu Delong , Tanaka Toshiki , Traganos Frank , Darzynkiewicz Zbigniew 
Published: April 2020

Abstract
Our previous experiments on leukemic cell lines demonstrated that the intensity of immunofluorescence (IF) that is proportional to the level of the phosphorylated histone H2AX (γH2AX) reveals the frequency of DNA double-strand breaks (DSBs) in chromatin induced by DNA topoisomerase I and II (topo I and II) inhibitors. The aim of the present study was to explore whether ex vivo assessment of the extent of DNA damage in leukemic blasts induced during chemotherapy is feasible. The long term goal is to develop a rapid assay to assess the effectiveness of chemotherapy early in the course of treatment of leukemia. Mononuclear cells were isolated from blood samples of 1 AML and 10 ALL patients with peripheral blasts, 4 of whom were, at diagnosis, high-risk ALL while 4 were in relapse. The median age of the patients was 17 yrs (2–25 yrs) and all were entered into an anthracycline-containing drug regimen. Samples were collected:
prior to drug administration; and, one hour after completion of the drug infusion.
The extent of DNA damage was assessed by analysis of histone H2AX phosphorylation occurring at sites of DSBs, which could be detected immunocytochemically and measured by multiparameter flow cytometry in conjunction with DNA content. In the only patient not responding to treatment, one of 4 relapsed ALL patients, there was no induction of DSBs and no activation of ATM. In all patients phosphorylation of ATM appeared to be strongly linked to induction of DSBs after treatment targeting topo inhibitors. In a pilot study of 3 adult patients with acute leukemia (two AML and one ALL in relapse) treated with the topo II inhibitor Mitoxantrone (MTX), ATM activation was strongly linked to induction of DSBs 1 h post drug infusion. ATM activation and induction of DSBs was several times higher in two adult responders in comparison to one non responder to therapy. A new approach to detect DNA damage in leukemic cell by flow cytometry using gating strategy which combines CD45 expression on the cell surface with side scatter (SS) is being developed and was applied in 5 cases. This approach will be useful when the proportion of blasts in peripheral blood is low. Although the number of patients is very small in this study, the results suggest that a cytometric assay that reveals the extent of DNA damage based on detection of H2AX phosphorylation during induction therapy targeting topo I and II inhibitors may provide an early prognostic marker.
ARRY-429520, a KSP Inhibitor with Efficacy in Animal Models of Solid and Liquid Tumors.
Authors: Woessner Rich , Corrette Chris , Allen Shelley , Hans Jeremy , Zhao Qian , Aicher Tom , Lyssikatos Joe , Robinson John , Poch Greg , Hayter Laura , Cox April , Lee Patrice , Winkler James , Koch Kevin , Wallace Eli 
Published: April 2020

Abstract
Kinesins are eukaryotic microtubule-associated motor proteins. One of these, kinesin spindle protein (KSP) is a mitotic-specific kinesin that plays a key role in spindle pole separation and production of the bipolar spindle, as well as centrosome separation and maturation. As KSP is expressed predominately in proliferating cells and is absent from postmitotic neurons, inhibition of KSP has the potential to provide the antitumor activity of a mitotic inhibitor while avoiding peripheral neuropathy. ARRY-429520 is a member of a series of KSP inhibitors discovered and optimized by structure-based design. It is a potent inhibitor of human KSP (IC50 6 nM), with an EC50 of 1.5 nM for cellular phosphorylation of histone H3 (a pharmacodynamic marker for accumulation of cells in mitosis), and in vitro antiproliferative IC50s ranging from subnanomolar to low single digit nanomolar across a varity of human and murine tumor cell lines. We report here the in vivo characterization of this compound. ARRY-429520 inhibits histone (H3) phosphorylation in tumor xenografts , showing that the compound has pharmacodynamic activity in vivo. As a result, it is a highly efficacious inhibitor of the growth of tumor xenografts. At its maximally tolerated dose in mice of 25 – 30 mg/kg i.p. on a Q4Dx3 schedule, ARRY-429520 caused tumor regressions, with some complete responses, in subcutaneous mouse xenograft models of leukemia: HL-60 (acute promyelocytic leukemia) and K-562 (chronic myelogenous leukemia). The compound was also highly efficacious against other tumor types, including HT-29 (colon), HCT-116 (colon), and A2780 (ovarian). ARRY-429520 also showed activity in intravenous leukemia models. The molecule possess desirable drug-like properties, including high aqueous solubility (&gt; 4 mg/ml at physiological pH), low CYP inhibition (&gt; 25uM for 3A4, 2C9, 1A2, 2D6, and 2C19), and pharmacokinetics favorable for an IV-infused targeted chemotherapeutic.
Preanalytical mRNA Stabilization of Whole Bone Marrow Samples.
Authors: Langebrake Claudia , Guenther Kalle , Lauber Juergen , Reinhardt Dirk 
Published: April 2020

Abstract
Background: Gene expression profiling is a useful tool for the diagnosis and basic research of cancer. One of the major limitations of this approach is that already a short-term storage of native specimens of peripheral blood (PB) or bone marrow (BM) and/or the method of RNA isolation has significant influence on gene expression due to gene induction, repression and RNA degradation. The objective of the current study was to investigate the influence of a newly developed RNA stabilization and preparation system for BM specimens (PAXgeneTM Bone Marrow RNA System), focusing on RNA-yield, RNA-integrity and stability of gene expression profiles over time of BM sample storage.
Patients and methods: 180 RNA samples that have been processed from 45 heparinized BM specimens of unselected children with acute leukemia at initial diagnosis, during treatment or without pathological findings in hematopoiesis were analyzed. Immediately after collection, heparinized BM specimens were divided into two PAXgeneTM RNA tubes and two standard tubes, each. RNA isolation using either the PAXgeneTM protocol (P) or a reference protocol (R) was performed after 2 hours (P0 and R0) or after 48 hours (P2 and R2).
Results: The overall RNA yield (normalized to 1×107 leukocytes) was not different in all four isolation procedures, however the integrity of the RNA using the PAXgeneTM system was significantly higher at both time-points than that of the reference system: 8.6±0.2 (P0) vs. 6.8±0.4 (R0), p=0.0003 and 8.1±0.2 (P2) vs. 6.7±0.5 (R2), p=0.008. For the stabilized samples, we found very good pairwise correlation in gene expression for either gene at P2 compared to P0: GATA1 89±6%, RUNX1 83±10%, NCAM 69±12% and SPI1 89±6%. However, there were significant differences in two of the analyzed genes using the reference RNA isolation procedure: 40±6%, p&lt;0.001 (GATA1) and 47±8%, p=0.005 (NCAM), respectively. Comparing the two RNA isolation procedures, there are significant differences in the expression levels of the analyzed genes.
Discussion: The PAXgeneTM system is appropriate for the stabilization of gene expression levels in BM samples after short-term storage. However, as in some genes the reference RNA isolation procedure resulted in similar gene expression levels, the use of a stabilization reagent has to be carefully evaluated within the preanalytical handling of the samples. Our analysis has shown that it is not suitable to compare RNA expression levels derived from different isolation procedures. In conclusion, the PAXgeneTM system is able to stabilize RNA from clinical BM samples while being suitable to isolate high quality and quantity RNA.
Study of FLT3 Internal Tandem Duplication (FLT3-ITD) in Patients with Acute Promyelocytic Leukemia (APL).
Authors: Hu Jiong , Zhu Yong-Mei , Liu Yuan-Fang , Zhang Su-Jiang , Shen Zhi-Xiang 
Published: April 2020

Abstract
Objectives To evaluate the prevalence and association with clinical features of FLT3 internal tandem duplication (FLT3-ITD) in patients with newly-diagnosed acute promyelocytic leukemia (APL).
Methods A total of 103 APL patients were screened by polymerase chain reaction (PCR) for FLT3-ITD.
Results FLT3-ITD mutations were identified in 19.4% (20/103) patients. The FLT3-ITD was associated with short/variant form of PML-RARα isoforms (p&lt;0.0001). Among 20 patients with FLT3-ITD, 16 patients presented with short, 2 with variant and 2 with long form of PML-RARα isoforms. The expression level of FLT3 via quantitative real-time RT-PCR was significantly increased in patients with FLT3-ITD (p&lt;0.05). Patients with FLT3-ITD also presented significantly increased initial peripheral white blood cell (WBC) count (p&lt;0.01), especially in patients with short/variant PML-RARα isoforms (p=0.015). For patients with long form PML-RARα, there was no significant difference in initial WBC count. For clinical outcome, 90% (18/20) of FLT3-ITD positive patients obtained complete remission and 16 evaluable patients (2 lost follow-up) remained in first remission with relatively short follow-up (median 26 months, 11-47), which was not significantly different from patients without FLT3-ITD.
Conclusion FLT3-ITDs were frequently identified in patients with newly diagnosed APL. FLT3-ITD was associated with short/variant form of PML-RARα fusion gene and increased initial WBC. No significant impact of FLT3-ITD on treatment outcome was observed with limited follow-up.
Association of FLT3-ITD and PML-RARα isoforms in patients with APL No FLT3-ITD+ FLT3-ITD− Long 57 2(3.5%) 55(96.5%) Short 41 16(39.0%) 25(61.0%) Variant 5 2(40%) 3(60%) Total 103 20(19.4%) 83(80.6%)
FLT3 expression in different patient sample FLT3 expression in different patient sample FLT3 expression in different patient sample FLT3 expression in different patient sample