bluuebunny/crossref_metadata_2025_split huggingface dataset:
Targeting of Heat Shock Protein 32 (Hsp32) in Neoplastic Cells by Styrene Maleic Acid Zinc Protoporphyrin (SMA-ZnPP) Is Associated with Reduced Growth and Induction of Apoptosis.
Authors: Gleixner Karoline V. , Matthias Mayerhofer , Vales Anja , Gruze Alexander , Kneidinger Michael , Hoermann Gregor , Krauth Maria T. , Kondo Rudin , Sonneck Karoline , Samorapoompichit Puchit , Lackner Eva , Greish Khaled , Pickl Winfried F. , Sillaber Christian , Zielinski Christoph C. , Maeda Hiroshi , Valent Peter 
Published: April 2020

Abstract
Heat shock protein 32 (Hsp32), also known as heme oxygenase-1 (HO-1), is a stress-related survival factor that has recently been implicated in enhanced survival of neoplastic cells. We here show that Hsp32/HO-1 is expressed abundantly in primary neoplastic cells in various solid tumors and hematopoietic neoplasms such as acute myeloid leukemia (AML) or chronic myeloid leukemia (CML), and in respective cell lines including the AML cell lines HL60, KG1, KG1a, and U937, CML cell lines K562 and KU812, eosinophilic leukemia cell line EOL-1, mast cell leukemia cell line HMC-1, myeloma cell lines RPMI8226 and U266, breast cancer cell line MDA MB 231, lung cancer cell line A549, pancreatic carcinoma cell line BxPC-3, colon carcinoma cell lines COLO201, COLO205, COLO320DM, and DLD-1, and the ovarian carcinoma cell line OVCAR-3. Expression of Hsp32 mRNA was demonstrable by RT-PCR and Northern blotting, and expression of the Hsp32 protein by Western blotting and immunocytochemistry. The CML-specific oncoprotein BCR/ABL and the transforming oncoprotein KIT D816V that is expressed in neoplastic mast cells, were found to promote expression of Hsp32 in Ba/F3 cells. In order to examine the functional role of Hsp32 in neoplastic cells, a specific siRNA was employed. Expression of Hsp32 siRNA resulted in reduced viability and induction of apoptosis in all cell lines tested. To further explore the value of Hsp32 as a target in neoplastic cells, a novel specific Hsp32-targeting compound, styrene maleic acid copolymer zinc protoporphyrin micelles (SMA-ZnPP) was applied. Exposure to SMA-ZnPP resulted in a significant decrease in proliferation determined by 3H-thymidine uptake, in all cell lines as well as in all primary neoplastic cells tested (AML, n=5; CML, n=5; mastocytosis, n=3; breast cancer, n=2; lung cancer, n=1). As assessed by AnnexinV-staining, Tunel assay and electron microscopy, the growth-inhibitory effects of SMA-ZnPP were found to be associated with induction of apoptosis. In each cell type, the effect of SMA-ZnPP on growth was dose-dependent and found to occur at pharmacologic concentrations (IC50 1–30 μM). Moreover, SMA-ZnPP was found to synergize with various anti-neoplastic drugs (tumor cell lines: cisplatin, leukemias: cytarabine, myeloma: bortezomib) in producing growth inhibition. In summary, these data show that Hsp32/HO-1 is an important survival factor and novel interesting target in various hematopoietic and non-hematopoietic neoplasms. Based on these data, it seems desirable to explore the value of the Hsp32-targeting drug SMA-ZnPP in clinical trials in patients with leukemias and solid tumors.
Mutation in Genes Impacting on the RAS-RAF-MEK-ERK Pathway Are Found at High Incidence in Childhood Acute Lymphoblastic Leukemia at Both Diagnosis and at Relapse.
Authors: Irving Julie , Minto Lynne , Matheson Elizabeth , Case Marian , Bailey Simon , Bown Nick , Hassan Rosline , Harrison Christine , Hall Andrew 
Published: April 2020

Abstract
Deregulation of the RAS-RAF-MEK-ERK signalling cascade, often caused by activating mutations in the RAS family of G-proteins, is a recurrent event in haematological malignancies including childhood acute lymphoblastic leukemia (ALL). However, deregulation of this pathway can also arise from mutational events in genes encoding other proteins which impact on this pathway including FLT3, PTPN11 and BRAF and such mutations have recently been identified in childhood ALL. The combined incidence of mutations in these genes, along with the RAS family is not known, but may be significant.
A comprehensive mutational screen of key exons of NRAS, KRAS2, FLT3, PTPN11 and BRAF was performed in 86 diagnostic and 49 relapse childhood ALL samples using denaturing high performance liquid chromatography (DHPLC) and direct sequencing and/or PCR product cloning. Using wild type cells spiked with cell lines bearing known mutations, the sensitivity of this screening assay was shown to be between 12–25%.
At diagnosis the combined mutation incidence was 35% (30 from 86) made up of NRAS (n=16), KRAS2 (n=15), FLT3 (n=3), PTPN11 (n=2), with 4 patients having more than one mutation. At relapse, 25 % (12 from 49) of cases showed mutations and included NRAS (n=4), KRAS2 (n=6), FLT3 (n=1) and PTPN11 (n=1). No mutations were identified in BRAF. While at relapse the mutated blasts were always the predominant population, in 7 of the 30 diagnostic cases, only a minor subpopulation of blasts housed the mutation and included all of the FLT3 insertion/deletion mutants. In 2 relapsed cases with mutations in KRAS2, mutations were not detectable by DHPLC in the matched diagnostic samples but were apparent at low level when a more sensitive allele-specific PCR was employed (assay sensitivity between 0.1–1%).
These data reveal a significant role of RAS-RAF-MEK-ERK pathway deregulation in leukemogenesis and implicate the pathway in some patients with progression to relapse. In addition, they highlight the genetic heterogeneity of leukemic blasts at diagnosis compared to those at relapse suggesting that clonal selection occurs in response to drug exposure. Inhibitors of the pathway, which are currently undergoing clinical trial, may be a novel therapeutic option for a significant proportion of children with relapsed ALL.
Dual Role of Pim-2 Depends on the Associated Proteome.
Authors: Levy Daphna , Cohen Amos M. , Don Jeremy 
Published: April 2020

Abstract
Pim-2, a Ser/Thr kinase, is a proto-oncogene originally identified as common proviral insertion sites in T and B cell lymphomas in mice. Deregulation of Pim-2 expression has been documented in several human malignancies, including leukemia, lymphoma, multiple myeloma and prostate cancer. In human non-Hodgkin’s lymphomas and in chronic lymphocytic leukemia Pim-2 is up-regulated and its expression correlates with disease activity. Pim-2 promotes cell survival in response to a wide variety of proliferative signals. Pim-2 promotes cell survival by phosphorylation of Bad and Cot. The goal of this study was to clarify the significance of our new unanticipated aspect of Pim-2′s function, namely, that over-expression of Pim-2 in HeLa cells led to cell cycle arrest at G1 and to increased apoptosis. We found that the G1 arrest was associated with increased (T14/Y15) phosphorylation of CDK2, increased proteosomic degradation of CDC25A, and increased levels of the CDK inhibitor p57. In addition, we found increased E2F-1 levels, which suggested the usage of the E2F-1 dependent apoptotic pathway. Using dominant negative forms of either E2F-1 or p73, which were co-expressed with Pim-2 in HeLa cells, revealed significant rescue of the G1 arrest and apoptotic phenomena. Silencing of Pim-2 in these cells, via siRNA, reversed the G1 arrest and pro-apoptotic effects, and verified the Pim-2 dependent specificity. We conclude that Pim-2 might play a dual role. Our data suggest that under certain environmental circumstances and in various cell types, Pim-2 appears to increase cell survival by abrogating some pro-apoptotic substrates, but under different proteomic associations Pim-2 might favor G1 arrest and apoptosis.
Computational Prediction of c-MYC Binding and Action by Integration of Multiple Data Sources.
Authors: Chen Yili , Blackwell Thomas W. , Gao Jing , Hewagama Anura , Grifka Heather M. , Lee Angel W. , States David J. 
Published: April 2020

Abstract
c-MYC is an important proto-oncogene. Its actions are mediated by sequence specific binding of the c-MYC protein to genomic DNA. While many c-MYC recognition sites can be identified in c-MYC responsive genes, many others are associated with genes showing no c-MYC response. It is not yet known how the cell determines which of the many c-MYC recognition sites are biologically active and directly bind c-MYC protein to regulate gene expression. We have developed a computational model that predict c-MYC binding and functional activation as distinct processes. Our model integrates four types of evidence to predict functional c-MYC targets: genomic sequence, MYC binding, gene expression and gene function annotations. First, a Bayesian network classifier is used to predict c-MYC recognition sites likely to exhibit high occupancy binding in chromatin immunoprecipitation studies using several types of sequence information, including predicted DNA methylation using a computational model to estimate the likelihood of genomic DNA methylation. In the second step, the DNA binding probability of MYC is combined with the gene expression information from 9 independent microarray datasets in multiple tissues and the gene function annotations in Gene Ontology to predict the c-MYC targets. The prediction results were compared with the c-MYC targets in public MYC target database [www.myccancergene.org], which collected the c-MYC targets identified in biomedical literatures. In total, we predicted 599 likely c-MYC genes on human genome, of which 73 have been reported to be both bound and regulated by MYC, 83 are bound by MYC in vivo and another 93 are MYC regulated. The approach thus successfully identified many known c-MYC targets as well as suggesting many novel sites including many sites that are remote from the transcription start site. Our findings suggest that to identify c-MYC genomic targets, any study based on single high throughput dataset is likely to be insufficient. Using multiple gene expression datasets helps to improve the sensitivity and integration of different data sources helps to improve the specificity.
Summary of c-MYC Targets Prediction Microarray Dataset Data Source (Citation) Tissue Predicted Targets Binding&Regulation Reported Only Binding Reported Only Regulation Reported 1 PMID: 15778709 B Cell 421 61 60 56 2 PMID: 12086878 Prostate Cancer 428 56 65 76 3 PMID: 14722351 Prostate Cancer 50 4 7 13 4 PMID: 15254046 Prostate Cancer 66 19 8 14 5 PMID: 12747878 Breast Cancer 17 1 3 5 6 PMID: 11707567 Lung Cancer 295 51 42 59 7 PMID: 15820940 CML 8 1 1 2 8 PMID: 12704389 ALL 222 45 32 46 9 PMID: 11731795 ALL / MLL / AML 22 6 1 6 Total 599 73 83 93
Transcriptional Regulation of the SHP-1, a Candidate Tumor Suppressor Gene in Hematological Malignancies.
Authors: Nakase Koichi , Cheng Jihua , Zhu Quan , Marasco Wayne A. 
Published: April 2020

Abstract
The SH2-Containing Protein-Tyrosine Phosphatase-1 (SHP-1) is a hematopoietic cell specific protein-tyrosine phosphatase and a candidate tumor suppressor. It has been shown that expression of SHP-1 is repressed in HTLV-1 transformed lymphocyte cell lines, Adult T-cell Leukemia and other hematological malignancies. However, the mechanisms that regulate SHP-1 expression remain unclear. We previously identified that the −120 to +157 of the SHP-1 P2 promoter (core promoter) exhibits similar levels of activity as the wild type promoter. Here we report the involvement of NF-kB and Oct-1 in the transcriptional regulation of the SHP-1 P2 promoter. To elucidate the mechanisms by which SHP-1 promoter is regulated, in vitro DNase I footprinting experiments were performed. Incubation of nuclear extracts from Jurkat cells with the SHP-1 P2 promoter probes revealed two DNase I protected regions between −100 and −90 and between −80 and −55 in the SHP-1 P2 promoter. Elk-1, Ik-1, NF-kB and Oct-1 were identified through database search as likely candidate factors binding to these regions. Electrophoretic Mobility Shift Assay (EMSA) was carried out using a double strand DNA probe that covers the previous DNase I footprinting area (−120 to −19) and Jurkat nuclear extract. Transcription factors NF-kB and Oct-1 were confirmed as binding to the SHP-1 P2 promoter through antibody supershift, probe competition, and/or site directed mutagenesis. Results from in vivo Chromatin Immunoprecipitation (ChIP) assay with anti-NF-kB and anti-Oct-1 antibody also strongly suggested that Oct-1 and NF-kB bound the SHP-1 P2 promoter in Jurkat cells. In transient transfection experiments using luciferase as a reporter, mutations at Oct-1 binding sites and NF-kB binding sites showed significant reduction in SHP-1 promoter activity compared to wild type core promoter, further indicating the involvement of these factors in the SHP-1 P2 promoter transcription regulation. In summary, this is the first report demonstrating that Oct-1 and NF-kB are involved in the regulation of SHP-1 P2 promoter. Modulation of these transcription factors could provide new targets for treatment of hematological malignancies.
ERBB Signaling Promotes BCR-Abl Expression and Modulates Multiple Signaling Pathways as Assessed by High-Throughput, Reverse Phase Protein Array.
Authors: Nelson Laura D. , Siwak Doris R. , Lu Yiling , Miller Claudia , Chandra Joya , Hughes Dennis P.M. 
Published: April 2020

Abstract
ERBB family receptor tyrosine kinases, EGFR, Her-2, Her-3 and Her-4, are prognostic and therapeutic targets in multiple carcinomas, including breast, lung, and head and neck tumors. We previously reported aberrant expression and activation of Her-2 and Her-4 in a panel of Ph+ ALL cell lines: Z33, Z119, and Z181. Loss of cell viability and apoptotic DNA fragmentation were observed after treatment with the irreversible pan ERBB inhibitor, CI-1033, in vitro. In the present study, we define the mechanisms of cell death initiated by ERBB blockade. We examined the expression and Tyr1248 phosphorylation of Her-2 in Z119 and Z181 cell lines upon CI-1033 treatment and found a 4-fold decrease in Ph-Y1248 by Western blotting. We then assessed the impact upon downstream signaling pathways using reverse-phase protein microarray (RPPA) - a new high-throughput method to measure and profile the total expression and post-translational phosphorylation of signaling pathway components. Leukemia cells treated with CI-1033 assessed with RPPA using 34 different antibodies showed mTOR expression to be decreased by more than two-fold in Z119 and Z181. Concomitantly, we saw a more than three-fold decrease in phosphorylated S6-S240/244. This suggests that down-regulation of translational control caused by inhibition of activated ERBB-family receptors contributes to the reduced proliferation in these cells. We also observed a 3.5-fold decrease in phosphorylated glycogen synthase kinase (GSK)-3a/b 21/9 in Z119 and a 1.7-fold decrease in Z181. This suggests inactivation of GSK3, which is regulated by Akt/PKB signaling. We also observed a four-fold decrease in Bcr-Abl expression in Z181, identified as p190 by Western. This suggests that ERBB signaling helps maintain expression of Bcr-Abl protein, driving Ph+ lymphomagenesis, perhaps explaining why ERBB blockade induces apoptosis in Ph+ ALL. Elucidating the mechanism of this apoptosis, CI-1033 caused activation of caspase-3 in Z119 and Z181 as measured by DEVD-AMC cleavage, and caused PARP cleavage. Thus multiple, non-redundant signaling pathways downstream of ERBB can be downregulated with a small molecule inhibitor, CI-1033, causing reduced viability and increased apoptosis in ERBB+ Ph+ ALL cell lines. Our data suggest that inhibition of these signals may provide additional therapeutic efficacy to Ph+ ALL patients.
Figure Figure
Single Versus Double Umbilical Cord Blood Transplantation (UCBT): Higher Risk of Acute Graft-Versus-Host Disease (GVHD) but Lower Transplant Related Mortality (TRM) in Recipients of Double UCBT.
Authors: MacMillan Margaret L. , Brunstein Claudio , DeFor Todd E. , Blazar Bruce R. , Weisdorf Daniel J. , Wagner John E. 
Published: April 2020

Abstract
Transplantation of 2 partially HLA matched UCB units has been shown to be a safe and effective means to overcome the cell-dose barrier in adolescents and adults. In addition, preliminary data suggest that double UCBT is associated with a greater graft-versus-leukemia effect (Blood2005:106:93a). To date, there has been no comparative analysis on the risk of acute GVHD in this population of patients. Therefore, we compared rates of acute GVHD in recipients of single (n=210) versus double (n=169) UCBT. Double UCB recipients were more likely to be older, transplanted in recent years, treated with a non myeloablative therapy, received mycophenolate mofetil as GVHD prophylaxis, and less likely to receive antithymocyte globulin (ATG) as part of the conditioning. Incidences of grades II–IV and III–IV acute GVHD were higher in recipients of double than single UCBT recipients (60% vs 33%, p.01) and (21% vs 11%, p=.01), respectively. Adjusting for differences between groups, two factors were associated with the development of grade II–IV acute GVHD in Cox regression: use of 2 UCB units (RR 2.0 vs 1.0 [95% CI, 1.3–3.2, p.01]) and absence of ATG in the preparative regimen (RR 1.0 vs 0.5 [95% CI, 0.3–0.8, p.01]). Other factors that were tested in the model but not determined to be significantly associated with acute GVHD were recipient age, gender, weight, diagnosis, time from diagnosis to UCBT, HLA disparity, total and CD3 cell dose, CMV serostatus, conditioning regimen, and GVHD prophylaxis. Despite increased risk of acute GVHD, TRM 1 year was significantly lower in recipients of double UCBT (17%, 95% CI, 5–29%) as compared to recipients of a single UCBT (47%, 95% CI, 26–68%; p =.02) (Figure 1) among those that developed grade III–IV acute GVHD. Survival at 1 year among those with grade III–IV acute GVHD was significantly higher after double UCBT (67%, 95% CI, 51–83%) than after single UCBT (41%, 95% CI 21–61%; p=.04) (Figure 2).
Figure 1 Figure 1. Figure 2 Figure 2.
In conclusion, risk of acute GVHD is significantly higher in recipients of two partially HLA matched UCB units, yet it does not appear to adversely effect TRM or survival. ATG in the preparative therapy, however, does not appear to reduce TRM despite its favorable association with reduced GVHD. Impact of GVHD on immune recovery and infection risk in recipients of single and double UCBT is under investigation.
Inhibition of UBE2Q2 Sensitizes Cells to Mitotic Arrest and Cytotoxicity Caused by Vincristine.
Authors: Crawford David F. , Banerjee Sami , Brooks William S. 
Published: April 2020

Abstract
A microarray study was previously performed to identify G2/M-specific genes and genes with altered expression in response to G2 DNA damage checkpoint activation. We predicted that many of these proteins would be mitotic regulatory proteins which might serve as potential targets in the treatment of malignancies. Included in this set were several proteins involved in ubiquitin-mediated protein degradation including a putative ubiquitin conjugating enzyme known as UBE2Q2. We have demonstrated that this gene is expressed in a cell cycle dependent manner and that the encoded protein can function as an ubiquitin conjugating enzyme (or E2) in vitro. To assess the role of this protein in mitosis and the cellular response to treatment with chemotherapy agents, we inhibited UBE2Q2 function by expressing a dominant-negative version of the protein. With dominant-negative UBE2Q2 (DN-UBE2Q2) expression, no alteration in normal cell cycle kinetics is observed in unperturbed cells. However, DN-UBE2Q2 expression causes a dramatic change in the cellular response to vincristine and other microtubule inhibiting agents (MIAs). While control cells transiently arrest in mitosis after low dose vincristine treatment, DN-UBE2Q2-expressing cells have an impaired ability to escape this arrest which leads to a dramatic potentiation of the mitotic arrest. Cells expressing DN-UBE2Q2 are also much more sensitive to the cytotoxic effects of both vincristine and paclitaxel, MIAs with distinct mechanisms of action. Although DN-UBE2Q2 causes sensitization to all MIAs tested, it does not confer increased sensitivity to etoposide or doxorubicin, two DNA damaging agents, indicating that the effect is specific for microtubule disruption. The increased sensitivity to MIAs conferred by DN-UBE2Q2 expression is a result of caspase-mediated apoptosis and can be blocked using a caspase inhibitor. In order to independently confirm these findings, we also inhibited UBE2Q2 expression using siRNA. As with dominant-negative expression, we found that RNAi directed against UBE2Q2 enhanced the vincristine-induced mitotic arrest. These findings suggest that inhibition of UBE2Q2 might be used to augment the effectiveness of MIAs in the treatment of hematologic and other malignancies.
Pleiotrophin Promotes Myeloma Proliferation through the Activation of Known and Novel Receptors.
Authors: Gordon Melinda S. , Lee Hee Jin , Campbell Richard A. , Drucker Charles B. , Katz Matthew , Leuchter Richard K. , Bonavida Benjamin , Berenson James R. 
Published: April 2020

Abstract
Myeloma survival and proliferation in the bone marrow (BM) depends on the expression of a variety of autocrine and paracrine growth factors, including IL-6, insulin-like growth factor I (IGF-I), vascular endothelial growth factor (VEGF), and hepatocyte growth factor (HGF). We recently discovered an additional new autocrine myeloma growth factor, pleiotrophin (PTN). PTN is an 18kD heparin-binding protein normally expressed during early development and downregulated in adults, but aberrant PTN re-expression has been associated with a variety of aggressive solid tumors including neuroblastoma, glioma, melanoma and lung, breast and prostate cancers. We found that interference with PTN using a polyclonal anti-PTN antibody inhibited the proliferation of myeloma cells in vitro and in vivo by inducing cell cycle arrest but not apoptosis. To determine the mechanism by which PTN stimulates myeloma proliferation we analyzed the expression of the known PTN receptors, syndecans 1 (CD138) and 3, the anaplastic lymphoma kinase (ALK) and the receptor tyrosine phosphatase beta/zeta (RPTPβ/ζ) on myeloma cells. The expression of ALK and RPTPβ/ζ has not previously been investigated in any hematologic malignancy. In addition to syndecan 1 we found that a subset of myeloma cell lines and BM mononuclear cells (BMMCs) from myeloma patients are RPTPβ/ζ+ by RT-PCR, Western blot and flow cytometry. Myeloma cells, however, do not express ALK. RPTPβ/ζ inhibition by PTN binding leads to the accumulation of β-catenin, and downstream activation of Wnt, NF-κB, MAPK and Akt-mediated cell signals in cells from solid tumors, signaling pathways known to contribute to myeloma cell survival and proliferation. The myeloma cell line MM-1S and the SCID-hu myeloma model LAGλ-1 are both CD138+ but RPTPβ/ζ−. Nonetheless, the growth of these cells is inhibited by anti-PTN antibody. CD138 lacks cytoplasmic signaling motifs suggesting that additional novel receptors are required for PTN-stimulated cell growth in these cells. To continue to define PTN-mediated signaling in myeloma cells we compared the induction of PTN-regulated signaling pathways in RPTPβ/ζ+ RPMI 8226 cells compared to RPTPβ/ζ − MM-1s cells by phospho-protein Western blot. We found differences in PTN-stimulated tyrosine phosphorylation between RPTPβ/ζ+ and RPTPβ/ζ −cells. Specifically, we also discovered that RPTPβ/ζ+ RPMI 8226 cells activate the MAPK Erk1/2. In contrast, we found Akt, but not Erk1/2, to be activated by PTN in RPTPβ/ζ − MM-1S cells. We are now continuing our analysis of PTN signaling in these cells. We are using also identifying new PTN receptors on the surface of myeloma cells by affinity chromatography using biotinylated-PTN. These studies will provide potential new targets that should lead to the development of novel targeted anti-myeloma therapies.