bluuebunny/crossref_metadata_2025_split huggingface dataset:
Zoledronic Acid Inhibits Protein Prenylation in Plasmacytoma Tumors In Vivo and Enhances Survival in the INA-6 SCID Mouse Model.
Authors: Guenther Andreas , Gordon Sharon , Bakker Frank , Burger Renate , Green Jonathan R. , Rogers Michael , Gramatzki Martin 
Published: October 2019

Abstract
Bisphosphonates such as zoledronic acid (ZOL) are effective at preventing osteolytic bone disease in patients with multiple myeloma. ZOL inhibits bone resorption by inhibiting FPP synthase and preventing the prenylation of small GTPases in osteoclasts. In vitro studies have demonstrated previously that ZOL can also directly affect the growth and viability of myeloma cells by inhibiting protein prenylation and therefore could, potentially, have a direct anti-tumour effect in vivo in addition to effects on osteoclasts. To examine this further, the effect of ZOL on six myeloma cell lines, including the IL-6 dependent INA-6 line, was investigated. ZOL caused cell cycle arrest and concentration-dependent growth inhibition in all six cell lines, with varying sensitivity (IC50= 30–285 μM).
The potential anti-tumour effect of ZOL on INA-6 cells in vivo was studied in a SCID mouse xenograft model, in which mice injected intraperitoneally with INA-6 cells develop plasmacytomas but do not develop significant osteolytic lesions. In several experiments involving more than 50 mice, ZOL was administered subcutaneously (sc) or intravenously (iv). ZOL, at a dose of 8 μg or 2 μg three times per week for two weeks after inoculation of INA-6 cells, significantly reduced tumour burden and increased survival of the mice (p= 0.002). The effect of ZOL on protein prenylation in plasmacytomas dissected from the mice was measured by western blotting to specifically detect the unprenylated form of the small GTPase Rap1A. Unprenylated Rap1A was virtually absent from tumour samples of untreated animals, while a single iv injection of 8 μg ZOL induced a marked accumulation of unprenylated Rap1A in the tumours after 24–72 hours.
These studies are the first to demonstrate that ZOL can inhibit protein prenylation in plasmacytomas in vivo. Together with the decreased tumour burden and increased survival, the data suggest that ZOL may have direct anti-tumour effects in this animal model. Thus, nitrogen-containing bisphosphonates such as ZOL may have therapeutic potential in multiple myeloma beyond the prevention of osteolytic lesions.
High FLT3 mRNA Expression in Acute Myeloid Leukemia May Be Functionally an Alternative to Mutational Activation of the Receptor.
Authors: Kuchenbauer Florian C. , Kern Wolfgang , Schoch Claudia , Kohlmann Alexander , Hiddemann Wolfgang , Haferlach Torsten , Schnittger Susanne 
Published: October 2019

Abstract
Mutations of the FLT3 gene are detectable in approximately 30% of all adult AML. These mutations lead to an autoactivation of the receptor inducing increased proliferation of the leukemic clone. An alternative mechanism of FLT3 activation might be mRNA overexpression. In the presented study the FLT3 expression level in 208 adult AML patients and 8 healthy donors was assessed by real time PCR and correlated to several parameters. In all patients cytomorphology, cytogenetics, and FLT3 mutation status was assessed. Significant differences of FLT3-expression levels were found in certain AML subgroups. The highest expression levels were found in FAB subtypes M5 and the lowest in M3. In total, increasing levels were shown in the following order: M3 <M3v <M6 <M2 <M4eo <M4 <M0 <M1 <M5a <M5b. Independent analysis of FLT3 expression in different cytogenetic AML subgroups showed the lowest median in the t(15;17) group, followed by t(8;21), inv(16), normal, complex karyotype and the highest median in the t(11q23) group. No difference was observed between the group of secondary AML following MDS, therapy related AML and the de novo AML (p=0.868, p=0.562, and p=0.570, respectively). Compared to clinical parameters, FLT3 expression correlated with high percentage of bone marrow blasts (p=0.0005) and high leukocyte count (p<0.001). In contrast to previous studies no difference in FLT3 expression levels was detected between AML with (n=74) and without (n=130) any FLT3 mutation. Assessment of FLT3 RNA by microarray analysis and FLT3 receptor surface expression (CD135) detected by flow cytometry correlated significantly with FLT3-expression as assessed by real time PCR (p<0.001, each). To analyze whether high FLT3 expression is a prognostic parameter 118 AML cases with normal karyotype were devided into two groups. Group 1 (n=75) was defined to have less and group 2 (n=43) more than the median of the FLT3 expression level found in the total group. No impact on OS and EFS could be shown (608 vs. 311 days, p=0.1283 and 398 vs. 208 day, p=0.3056). In conclusion, these data support the hypothesis that FLT3 activation through mRNA overexpression is an alternative mechanism to FLT3 mutations. Especially as it was found extremely high in 11q23 AML, that rarely reveal FLT3 mutations.
Phase 3 Randomized Trial of Fludarabine/Cyclophosphamide Chemotherapy with or without Oblimersen Sodium (Bcl-2 Antisense; Genasense; G3139) for Patients with Relapsed or Refractory Chronic Lymphocytic Leukemia (CLL).
Authors: Rai Kanti R. , Moore Joseph O. , Boyd Thomas E. , Larratt Loree M. , Golenkov Anatoliy K. , Koziner Benjamin , Chanan-Khan Asher , Bociek R. G. , Itri Loretta M. 
Published: October 2019

Abstract
Bcl-2 is an anti-apoptotic protein that has been closely linked to chemotherapy resistance and inferior survival in patients (pts) with CLL. Preliminary studies showed that oblimersen, an antisense oligonucleotide that targets Bcl-2, enhanced apoptosis induced by fludarabine (Flu), dexamethasone, and rituximab in cultured CLL cells. In a Phase 1 study, oblimersen displayed modest single-agent activity in CLL pts who had received extensive prior therapy. We conducted a randomized, multinational, Phase 3 trial of chemotherapy given with or without oblimersen in pts with advanced CLL. Eligibility included: symptomatic disease; intermediate or high-risk Rai stage; ≥ 1 Flu-containing regimen; ECOG PS ≤ 2; adequate organ function. Exclusions: stem cell/allogeneic transplant; autoimmune anemia/thrombocytopenia. Pre-randomization strata: prior treatment (1 or 2 vs. ≥ 3 regimens); response vs. refractory to prior Flu; and response duration to last therapy (> or ≤ 6 mos). All pts received Flu 25 mg/m2/d, plus cyclophosphamide (Cy) 250 mg/m2/d, daily for 3 days. Pts randomized to oblimersen received 3 mg/kg/d x 7 days by IV infusion, beginning 4 days before Flu/Cy and on each day of Flu/Cy treatment. Subsequent cycles were repeated every 28 days, for a maximum of 6 cycles, or to the time of CR, intolerable toxicity, or progression. All pts received prophylactic filgrastim, TMP/SMZ, and acyclovir. Response assessment was conducted at every cycle during treatment, and every 2 months thereafter for up to 36 months. The primary endpoint of the study was to compare the proportion of pts who achieved complete (CR) and nodular partial (nPR) responses. Secondary endpoints included: overall response rates (CR/nPR/PR); durable responses; response duration; time-to-progression; clinical benefit; and safety. Bone marrows in responding pts underwent central blinded review. A total of 241 pts were randomized; the last patient was enrolled in the 2nd quarter 2003. All pts have completed ≥ 1 year of follow-up, and results of this trial will be presented.
Perturbed Endocytosis by CALM-Containing Fusion Proteins: A Leukemogenic Mechanism in AML.
Authors: Chao Mwe Mwe , Walker Ann C. , Pendergast Molly B. , Bohlander Stefan K. , Wechsler Daniel S. 
Published: October 2019

Abstract
Background: We recently identified a novel fusion protein, MLL-CALM, in an infant with fatal AML. Although disruption of normal MLL function clearly contributes to leukemogenesis, perturbed CALM function may also be important. The native CALM protein plays a key role in clathrin-dependent endocytosis (CDE) and intracellular vesicle transport. Interestingly, CALM was first described as a partner for the AF10 gene - itself an MLL partner - in aggressive leukemias and lymphomas. Recently, it was shown that mutations in calm are responsible for hematopoietic abnormalities in inbred fit1 mice. Taken together, these observations suggest an important role for CALM in both normal and malignant hematopoiesis. We hypothesize that disrupted CDE resulting from the presence of CALM-containing fusion proteins interferes with downregulation of growth factor signaling, thereby contributing to leukemogenesis.
Objective: 1) To determine whether expression of MLL-CALM or CALM-AF10 interferes with endocytosis of transferrin (TF), 2) To analyze the importance of distinct CALM domains which affect endocytosis, and 3) To determine whether MLL-CALM expression reduces the IL-3 growth factor dependence of FL5.12 cells.
Design/Methods: The relative ability of leukemia cell lines that harbor MLL and CALM translocations to endocytose AlexaFluor633-conjugated TF was assessed by flow cytometry. The degree of Texas Red (TR)-TF endocytosis in COS7 cells expressing GFP-tagged MLL-CALM and CALM-AF10 was quantitated using densitometry. CALM deletion constructs were prepared and endocytosis of Texas Red-TF was examined in COS-7 cells transfected with these plasmids. Finally, IL-3-dependent FL5.12 cells engineered to stably express MLL-CALM were grown in decreasing amounts of IL-3-containing WEHI conditioned medium and survival was determined.
Results: U937 and P31/Fujioka cells (that harbor the CALM-AF10 translocation) showed reduced endocytosis of AF633-TF, but cells that lack CALM translocations (THP-1, K562, MonoMac6, HL-60) showed robust AF633-TF uptake. COS7 cells expressing MLL-CALM or CALM-AF10 showed 40–50% less endocytosis of TR-TF compared with untransfected cells or cells transfected with empty vector. CALM aa 484–620 appear to be important in inhibiting endocytosis. IL-3-dependent FL5.12 cells transfected with an MLL-CALM expression vector grew better in low concentrations of IL-3-containing WEHI conditioned media in comparison with FL5.12 cells transfected with empty vector.
Conclusions: Expression of MLL-CALM or CALM-AF10 interferes with endocytosis, and MLL-CALM expression renders FL5.12 cells less dependent on exogenous IL-3. The ability of MLL-CALM and CALM-AF10 to interfere with CDE may result in sustained growth factor signaling and contribute to the development of aggressive hematopoietic malignancies.
Identification of Genomic Imbalances in AML with Complex Karyotype Using Matrix-Based Comparative Genomic Hybridization.
Authors: Rucker Frank G. , Schwanen Carsten , Lipka Daniel , Wessendorf Swen , Frohling Stefan , Kestler Hans , Lichter Peter , Bentz Martin , Dohner Konstanze , Dohner Hartmut 
Published: October 2019

Abstract
Approximately 10 to 15 % of acute myeloid leukemia (AML) cases exhibit complex karyotypes, i.e., three or more chromosome abnormalities without presence of a specific fusion transcript. Using chromosome banding analysis, the majority of such cases cannot be completely described due to the low resolution of this method. Comparative genomic hybridization to microarrays (matrix-CGH) is a novel technique that allows genome-wide screening for genomic imbalances at high resolution and thus may facilitate the identification of novel regions harboring potential disease-related genes. We constructed a microarray consisting of 2799 different human genomic DNA fragments cloned in bacterial artificial chromosome (BAC) or P1-derived artificial chromosome (PAC) vectors. A set of 1502 of these clones covers the whole human genome with a physical distance of approximately 2 Mb. The remaining 1297 clones either contiguously span genomic regions known to be frequently involved in hematologic malignancies (e.g., 1p, 2p, 3q, 7q, 9p, 11q, 12q, 13q, 17p, 18q) (n=610) or contain oncogenes or tumor suppressor genes (n=687). In this study 45 AML cases with complex karyotypes were analyzed. Matrix-CGH detected genomic aberrations in all cases and genomic losses were found more frequently than gains. The most frequent aberration was deletion 5q in 38 of 45 cases. Three consensus regions were delineated mapping to chromosomal bands 5q11 (23 cases), 5q34 (22 cases), and 5q35 (21 cases). Further frequent deletions affected 17p (60%); two consensus regions were identified mapping to chromosomal bands 17p13 (60%), and 17p11-q11 (33%). Other chromosomal losses affected 7q (51%), 16p (31%), 16q, 18q (29% each), 15q (24%), 10p (22%), 14q, 4p (20% each), 11q, 20q (18% each), 3p, and 12p (15% each). The most frequent genomic gain was identified at 8q (40%). Further frequent gains were detected on 11q (38%), 21q (27%), 1p, 2q (24% each), 9p (22%), 6p, 19p (20% each), 3q (18%), 7p, 20p (15% each), 13q, and 20q (13% each). Genomic amplifications were identified on chromosomal band 8q24 including the MYC gene (2 cases), 9p24 (JAK2, 2 cases), 10p15 (PRKCO, 1 case), 11q23 (MLL, 2 cases), 11q23.3 (ETS1 and FLI1, 2 cases each), 12p13 (CCND2 and FGF6, 2 cases), 13q12 (CDX2, 3 cases), 20q11.1 (BCL2L, 1 case), 21p (1 case), 21q (2 cases), and 22q (1 case). These data demonstrate the potential of matrix-CGH with regard to spatial resolution and sensitive detection of genomic imbalances. Analysis of a large series of AML cases with complex karyotype may lead to the identification of novel critical regions and pathogenetically relevant genes.
Triptolide-Induced Apoptosis Is Dependent on Caspase Activation and NFκB Signaling and Mediated by XIAP and Survivin Downregulation through the Mitochondrial Pathway in Leukemic Cells.
Authors: Carter Bing Z. , Schober Wendy D. , McQueen Teresa , Evans Randall L. , Andreeff Michael 
Published: October 2019

Abstract
Triptolide, an immunosuppressor isolated from the Chinese herb, Tripterygium wilfordii Hook. F, has recently shown anti-tumor activities in a broad range of solid tumors. We examined its effects on leukemic cells and investigated mechanisms of apoptosis. Triptolide, at less than 100 nM, arrested cell growth and potently induced cell death in myeloid and lymphoid leukemic cells tested, including OCI-AML3, U937, Jurkat, KBM5, and K562 cells. In OCI-AML3 cells, triptolide induced caspase 3 activation, PARP cleavage and annexin V positivity with an IC50 of about 30 nM, at 24 hrs, all of which were inhibited by a general caspase inhibitor suggesting caspase dependent cell death. However, Triptolide-induced cell growth arrest was not affected by caspase inhibition. Treatment of OCI-AML3 cells with triptolide decreased XIAP and survivin expression, but did not affect Bcl2 and BclXL levels. Forced overexpression of XIAP attenuated Triptolide-induced cell death. Triptolide induced Bid cleavage, but Jurkat cells deficient in caspase 8 were only slightly less sensitive to triptolide than the wild-type counterpart indicating that Triptolide-induced cell death is caspase 8 independent. Jurkat cells deficient in receptor interacting protein (RIP) and therefore deficient in NFκB activation were resistant to Triptolide demonstrating that NFκB signaling is essential for Triptolide-induced cell death. Triptolide treatment induced cytosolic release of cytochrome C and loss of mitochondrial membrane potential, overexpression of Bcl2 effectively suppressed apoptosis induced by Triptolide, and caspase 9 knockout MEF cells were resistant to Triptolide suggesting criticality of the mitochondrial pathway. The antioxidants GSH (5 mM) and vitamin C (150 μM) did not protect from apoptotic cell death induced by Triptolide. In addition, Triptolide-induced apoptosis of blast crisis CML KBM5 cells was independent of their sensitivity or resistance to Imatinib: Triptolide killed Imatinib resistant KBMSTI cells as effectively as Imatinib sensitive KBM5 cells. Ex vivo studies showed that Triptolide also induced cell death in primary AML blasts. Collectively, our studies demonstrate that Triptolide potently induces caspase-dependent apoptosis and arrests cell growth in leukemic cells. Triptolide-induced cell death is dependent on NFκB signaling, and mediated by downregulation of XIAP and survivin through the mitochondrial pathway. The potent anti-leukemic activity of Triptolide in vitro warrants further investigation of this compound for the treatment of leukemia and other malignancies. This drug may also be potentially useful in overcoming Imatinib resistance in CML and Philadelphia chromosome positive ALL.
Reoxygenation Induces Premature Senescence of FA Hematopoietic Progenitor/Stem Cells through a Pathway Involving ATM, P53, and P21CIP1/WAF1.
Authors: Zhang Xiaoling , Li June , Sejas Daniel P. , Pang Qishen 
Published: October 2019

Abstract
Hematopoietic cells are often exposed to transient hypoxia as they develop and migrate between blood and tissues. We sought to test the role of continuous cycles of hypoxia and reoxygenation in the pathogenesis of bone marrow failure in Fanconi anemia (FA). We isolated Lin− BM cells from wild-type (WT) and Fancc−/− mice, and subjected them to two cycles of hypoxic (1% O2) then reoxygenation (20% O2). Expansion of Fancc−/− Lin− BM cells was significantly decreased in response to the oxidative stress of reoxygenation, as compared with WT Lin− BM cells (1.3 vs. 4.9-fold by day 6, respectively). This inhibition was attributable to a marked decrease of Fancc−/− progenitor (Lin− ScaI− c-kit+) cells, as well as a slower expansion of Fancc−/− stem (Lin− ScaI+ c-kit+) cells than the WT stem cells following reoxygenation. Fancc−/− Lin− BM cells transduced with FANCC retroviruses exhibited a significant growth advantage to untransduced cells, particularly in hypoxia and reoxygenation conditions where the predominance of FANCC/GFP+ progenitors was observed in both the short-term liquid culture and clonogenic assays. There was no evidence of increased apoptotic death in these reoxygenated Fancc−/− BM progenitor and stem cells compared to their WT counterparts, as assessed by a flow cytometric method for caspase 3 activation (early apoptosis). Interestingly, evaluation of the apoptotic profile and cell cycle of stroma-supported long-term BM culture (LTBMC; two weeks after second reoxygenation) revealed that reoxygenated Fancc−/− LTBMC cells had reduced apoptosis compared to reoxygenated short-term culture. However, a vast majority (70.6%) of reoxygenated Fancc−/− LTBMC cells was residing in the G0 + G1 phases compared with 55.8% in WT LTBMC cells. Fancc−/− LTBMC cells stained intensely for SA-b-galactosidase activity, a biomarker for senescence; this was associated with increased expression of senescence-associated proteins p53 and p21CIP1/WAF1 and to a lesser extent, p16INK4A. Moreover, reoxygenation-induced phosphorylation of p53ser15 was dependent on the ATM kinase but not ATR, as inhibition of ATM signaling by the kinase inhibitor 2-aminopurine or siRNA knockdown of ATM decreased p53ser15 phosphorylation and reduced cell senescence. Taken together, these results suggest that reoxygenation induces premature senescence in Fancc−/− BM hematopoietic progenitor and stem cells by signaling through the ATM to p53, upregulating p21 and causing senescent cell cycle arrest. In addition, reoxygenation-induced premature senescence may be a novel mechanism underlying hematopoietic stem cell depletion and BM failure in FA. <B style=%
Investigations on the Mechanisms of Resistance to Pixantrone in Tumor Cells.
Authors: Hazlehurst Lori A. , Alsina Melissa , Hacker Miles P. 
Published: October 2019

Abstract
Pixantrone is an aza-anthracenedione currently undergoing Phase 3 clinical trials and has shown significant activity in non-Hodgkin’s lymphoma. Preclinical studies with the aza-anthracenediones demonstrated a remarkable structure activity requirement, i.e., the ring nitrogen must be in the 2 position for anti-tumor activity. More importantly, Pixantrone which lacks hydroxyl groups at either the 1 or 4 positions of the chromophore, lacked any cardiotoxicity in pre-clinical trials. These pre-clinical findings have been substantiated during clinical development of Pixantrone. Given that there is a very real possibility that Pixantrone will progress to broader clinical use, we reasoned that understanding potential mechanisms by which tumor cells may become resistant to the drug were essential. By continuous in vitro exposure of MCF-7 cells to increasing concentrations of Pixantrone, we have developed a cell line (MCF7/aza) 20 fold resistant to the drug. These cells are cross resistant with mitoxantrone, surprisingly to a much great degree of resistance to mitoxantrone than to Pixantrone. Two compounds BBR 3438 and BBR 3576, represent a new class of anticancer drugs- the aza-anthrapyrazoles- have entered clinical trials and both were cross resistant in the MCF 7/aza cells. Using western blotting techniques we have demonstrated that the MCF 7/aza cells express elevated levels of BCRP but not Pgp. Further, resistance to Pixantrone was reversed with fumitremorgin C, a classic BCRP inhibitor that reverses mitoxantrone resistance in BCRP expressing cells. Resistance to Pixantrone was not reversed by verapamil, the classic Pgp inhibitor that reverses mitoxantrone resistance in MDR cells. Microscope evaluation of cells treated with Pixantrone demonstrated a unique intracellular distribution of the drug in MCF 7/aza cells in that the drug was sequestered in cytoplasmic vesicles rather than in the nucleus. More recently wehave completed a microarray analysis of MCF 7/Aza compared to MCF 7 cells and not surprisingly found a large number of genetic alterations in the resistant phenotype. One of the most striking alterations was a greater than 30 fold increase in BCRP. The results of our studies indicate that Pixantrone, a non-cardiotoxic mitoxantrone analogue, induced a resistant phenotype more closely related to that seen with mitoxantrone than the multi-drug resistance seen with many other anti-cancer drugs. It is of interest that an aza-anthracenedione, BBR 2378, was not cross resistant in MCF 7/aza cells. The unique structural feature of BBR 2378 is the presence of tertiary amines at the terminus of both sides arms of the compound in contrast to Pixantrone, BBR 3438, BBR 3576 or mitoxantrone all of which lack tertiary amine side arms. BBR 2378 has been previously been shown to be active in MDR cell linesas well. These results suggest that side arms play a pivotal role in determining activity or resistance in a variety of drug resistance tumor phenotypes.