bluuebunny/crossref_metadata_2025_split huggingface dataset:
In AML, Cbl Serves as Nexus for Signaling from Flt3, and Is Required for Coupling JNK1 in a Pathway of Survival and Proliferation Involving c-jun/AP-1.
Authors: Hartman Amy D. , Wilson-Weekes Annique , Suvannasankha Attaya , Burgess Gem S. , Hincher Kathryn J. , Cripe Larry D. , Boswell H. Scott 
Published: April 2020

Abstract
Here we demonstrate by immunoprecipitation and immunoblot, cbl is among the most heavily tyrosine phosphorylated adaptor proteins in primary AML blasts with Flt3 signaling, in the context of either mutation or overexpression/autocrine mechanisms. The human leukemic cell lines MV-4-11 and THP-1 model primary AML blasts in terms of Flt3 signaling by these respective criteria and demonstrate identical coupling between Flt3 and p85, the PI-3-kinase adaptor, by coimmunoprecipitation/blot experiments. Although cbl has no direct binding site on Flt3, it binds tightly to p85 SH2 by virtue of its tyrosine phosphorylation, also demonstrated by co-IP in cell lines and primary cells. Tyrosine phosphorylated cbl is a docking site for CrkII/L SH2’s and this provides a branch point for signals from Flt3 to PI-3-kinase or JNK, respectively, because CrkII(L) is known to bind JNK1 through SH3: polyproline interaction to serve as scaffolding; and interaction of JNK1 and CrkII/L was also observed by co-IP. In a survey of primary AML cases (n=33) there was a strict relationship between expression levels of (active) Flt3 and phospho-c-jun as readout for JNK activity level (p=0.001, r=0.54). To demonstrate the functional relevance of these interactions, siRNA knockdown of components was pursued in the cell lines and in primary AML blasts. JNK1 knockdown, and, to a much lesser degree, JNK2 knockdown, led to loss of phospho-c-jun expression in MV-4-11 and THP-1. Indeed, Flt3 signaling is required for JNK signaling because knockdown of Flt3 led to total loss of p-jun and c-jun expression in MV-4-11 and patient blast. By contrast, cbl knockdown led to selective loss of JNK signaling to p-jun without significantly affecting Flt3 or its downstream activating phosphorylation of AKT. Thus, despite binding by cbl to p85, cbl is not required for PI-3-kinase signaling because of redundancy supplied by the p85-Flt3 interaction. Further, by use of LY294002 to inhibit PI-3-kinase, PI-3-kinase is also not required for JNK signaling. However, selective inhibition of JNK signaling by the small molecule approach in these cell lines and in primary AML blasts leads to loss of proliferation, induction of apoptosis, and synergistic killing with daunorubicin. These observations form the platform for a phase I trial of JNK inhibition in refractory, multidrug-resistant, and Flt3-driven AML.
Leukemic Transformation Induces the Constitutive Expression of Indoleamine 2,3-Dyoxigenase Which Directly Increases CD4+CD25+ T Regulatory Cells.
Authors: Curti Antonio , Pandolfi Simona , Aluigi Michela , Valzasina Barbara , Isidori Alessandro , Ferri Elisa , Rutella Sergio , Durelli Ilaria , Horenstein Alberto L. , Fiore Francesca , Massaia Massimo , Piccioli Milena , Palmerini Emanuela , Pileri Stefano , Colombo Mario P. , Baccarani Michele , Lemoli Roberto M. 
Published: April 2020

Abstract
The expression of the catalytic enzyme of tryptophan, indoleamine 2,3-dyoxigenase (IDO) in different cellular subsets, including solid tumors, has been recently identified as a T-cell inhibitory effector pathway. Here, we show that unlikely normal hematopoietic CD34+ cells expressing IDO only upon stimulation with IFN-γ, acute myeloid leukemia (AML) cells constitutively express IDO and inhibit allogeneic T-cell proliferation. IDO expression correlates with increased CD4+CD25+Foxp3+ T cells in AML patients at diagnosis. In vitro, IDO+ AML cells increase the percentage and the absolute number of CD4+CD25bright T cells, also expressing surface CTLA-4 and Foxp3 mRNA. The addition of the IDO-inhibitor, 1-methyl tryptophan (1-MT), completely abrogates such increase. Purified CD4+CD25+ T cells obtained from coculture with IDO+ AML cells are not proliferating and unable to produce IL-2. They also inhibit naive T-cell proliferation and Th1 skewing. Co-culture with IDO+ AML cells results in the conversion of CD4+CD25− into CD4+CD25+ T cells and the addition of 1-MT completely abrogates this effect. In mice, intrasplenic injection of IDO-expressing leukemia/lymphoma A20 cells induces the increase of CD4+CD25+ T cells, which can be blocked by 1-MT treatment. These data suggest that AML cells have the capacity to directly increase a population of T regulatory cells through the constitutive expression of the functionally active form of IDO. IDO expression can be regarded as a novel mechanism of leukemia escape from immune control and its inhibition may represent a novel anti-leukemia therapeutic strategy.
Gene Expression Profiling of Bulk CD138+ Plasma Cells in Preneoplastic Gammopathy/Asymptomatic Myeloma Identifies Distinct Subsets of Patients.
Authors: Dhodapkar Madhav V. , Zhan Fenghuang , Rasmussen Erik , Burington Bart , Durie Brian , Crowley John , Barlogie Bart , Shaughnessy John 
Published: April 2020

Abstract
Gene expression profiling of plasma cells (GEP-PC) has provided major insights into myeloma pathobiology. However the data about GEP-PC in preneoplastic gammopathy (MGUS) or asymptomatic myeloma (AM) are limited, and gene expression patterns that might predict outcome in these patients have not been defined. We analyzed GEP (using U133Plus Affymetrix microarrays), of plasma cells isolated by immuno-magnetic bead selection with CD138 microbeads, from the bone marrow of patients with MGUS (n=16) and asymptomatic myeloma (AM; n=18) enrolled in a prospective South West Oncology Group (SWOG) observational study. Data from normal plasma cells (PCs) and from 105 myeloma PCs were included as controls. Myeloma PCs were randomly selected to include at least 15 patients from each of the 7 subgroups previously identified based on GEP of myeloma tumor cells (Zhan and Shaughnessy, ASH 2004). After the suppression of immunoglobulin (Ig) genes, there were 1297 genes that significantly differed in expression between MGUS-PCs and MM-PCs, and 1099 genes that differed between MGUS-PCs and normal PCs with a 1% false discovery rate. Hierarchical cluster analysis of all samples was performed using 1000 plasma cell signature genes that were most differentially expressed between normal and myeloma PCs. These data demonstrated that both MGUS and AM samples were distributed between normal and MM samples. A prediction analysis of microarrays (PAM) model (PNAS99:6567, 2002) utilizing 134 genes was then developed to determine if the signature from these genes in MGUS/AM was more similar to normal or to myeloma plasma cells. In this analysis, 11/16 (69%) of the MGUS samples were more similar to normal PC, compared to 6/18 (33%) of the AM samples (p=0.04). At present, there are no reliable phenotypic markers to distinguish between normal and malignant PCs within the bulk CD138+ population. Gene expression spikes for cyclin D1 and MAF/MAF-B were seen in both MGUS and AM cohorts, including in some patients with normal PC signature. These data provide the largest comparison to date, of GEP of PCs in preneoplastic versus malignant gammopathies and suggest that GEP may be a useful tool to prospectively identify subsets of patients within the MGUS/AM population with dominant normal PC or MM PC signatures, and potentially differing prognosis. Further analysis of differentially expressed genes between MGUS/MM PCs identified in this dataset may allow insights into the genomic changes in tumor cells underlying the malignant progression of myeloma.
The Paternally Expressed Gene 10 (PEG10) on Chromosome 7q21 Is Overexpressed and Imprinted in High-Risk B-CLL.
Authors: Kainz Birgit , Bilban Martin , Shehata Medhat , Heintel Daniel , Kienle Dirk , Kroemer Elisabeth , Le Trang , Kroeber Alexander , Schwarzinger Ilse , Gleiss Andreas , Chott Andreas , Doehner Hartmut , Zoechbauer9 Sabine , Heller9 Gerwin , Fonatsch Christa , Gaiger Alexander , Stilgenbauer Stephan , Zielinski Christoph , Wagner Oswald , Jaeger Ulrich ,   German CLL Study Group
Published: April 2020

Abstract
Introduction: To establish surrogate markers for IgVH mutation status in B-CLL, we performed microarray analysis with mutated and unmutated patient samples. Among the most differentially expressed genes was PEG10, a maternally imprinted gene located on chromosome 7q21. PEG10 is known to be expressed during placental development and in hepatocellular carcinoma. Here we have investigated the association of PEG10 with B-CLL subtypes and risk factors as well as its transcriptional regulation. In addition, we studied its expression in other hematologic malignancies.
Methods and Results: In our microarray experiments, PEG10 was highly expressed (2.1-fold) in unmutated samples. We confirmed these results by real time PCR in 42 B-CLL patients and found a significant correlation between PEG10 expression and unmutated IgVH status (P=0.007). High PEG10 expression was associated with chromosomal del(11q) (P=0.008), a shorter time to treatment (P=0.009) as well as a positive Coombs test (P=0.04). High expression was found in B-CLL and sorted CD19+ B-CLL cells, plasmoblastic DLBCL as well as in the Burkitt’s lymphoma cell line Ramos. In normal tissues, relatively high levels were seen in brain, liver, lung and kidney, but these expression levels were lower than in CD19+ B-CLL cells. Expression in sorted B cells from healthy donors was lower than two orders of magnitude. Intracellular expression of the PEG10 protein in B-CLL cells was confirmed by immunofluorescence studies using antibodies that recognize specifically PEG10-RF1. To address the question whether PEG10 overexpression results from loss of imprinting, we performed allele-specific as well as methylation-specific PCR. Imprinting was maintained in 6 patients with B-CLL. Analysis of a mother and son, both having B-CLL, confirmed that at least in this familial case the maternal PEG10 allele was silenced as expected. Interestingly, similar expression patterns were observed for the neighbouring gene sarcoglycan epsilon indicating a common mechanism of deregulation of the 7q21 locus in high risk B-CLL.
Conclusions: We show that PEG10 is differentially expressed in B-CLL and is associated with del(11q), a shorter time to treatment and unmutated IgVH mutation status, suggesting its role as a marker for high risk B-CLL. PEG10 overexpression is unlikely to result from loss of imprinting. Further studies of the gene locus at 7q21 are in progress.
Aberrant Expression of Ikaros Induces T-Cell Leukemia/Lymphoma in Mice without Wild-Type Allelic Loss.
Authors: Kataoka Itaru , Ishimaru Fumihiko , Ichimura Koichi , Yoshino Tadashi , Tanimoto Mitsune 
Published: April 2020

Abstract
The Ikaros gene consisting of seven exons codes a family member of zinc finger transcription factors essential for establishment of immune systems. A number of isoform proteins are produced by alternative splicing. The activity of Ikaros is strictly regulated by total expression level or mutual dimerization with full-length and dominant-negative isoforms. Findings from Ikaros mutant mice revealed its characteristics as tumor suppressor in addition to functions of hemato-lymphoid development and differentiation. Mutant mice homozygous for lack of dimerization domains evolve T-cell leukemia and lymphoma. Heterozygous mutation of DNA binding domains can also lead to clonal expansion of T-cells through loss of heterozygosity. In both of murine models reported earlier, leukemias arise from the absence of full-length active isoforms. On the other hand, analyses for human diseases conducted by our group elucidated high incidence of over-expression of dominant-negative isoform in adult precursor B-lymphoblastic leukemia. Thus the mechanisms by which Ikaros contribute to leukemogenesis are critical issues to be clarified especially in human. Here, we demonstrate that over-expression of dominant-negative isoform causes leukemia and lymphoma without wild-type allelic loss by murine BMT assay. Retroviral gene transfer of dominant-negative isoform, Ik6, to murine hematopoietic cells developed lethal lympho-proliferative disorders in 3–6 months of latency with complete penetrantion. The diseases involved a variety of organs such as thymus, lymphnode, spleen, liver and bone marrow. Clonality of the infiltrating cells was confirmed by Southern blotting. The leukemic cells showed T-cell phenotype (B220-CD3e+CD4+CD8a+TCRb+Gr1-Mac1-) and express simultaneously full-length Ikaros proteins and transduced dominant-negative isoform. These data suggest that leukemogenesis mediated by decreased activity of Ikaros does not require deletion of full-length isoforms, and disproportional expression of Ikaros directed to dominant-negative isoform is enough to accumulate and immortalize lymphoid blasts in vivo. Silencing dominant-negative Ikaros proteins should be considered as a good candidate of molecular targeted therapy, and quantification of Ikaros isoforms might be useful for monitoring the minimal residual disease.
Potential Genetic Risk Factors for Chronic ITP of Childhood.
Authors: Williams Shirley A. , Imfeld Karen L. , Tai LeiQian , Nakagawa Peggy A. , Chang Mei , Nugent Diane J. 
Published: April 2020

Abstract
While well recognized as an autoantibody mediated disease, the pathophysiology of childhood ITP remains poorly understood. Approximately 20% of children diagnosed with acute ITP will have persistent thrombocytopenia for more than 6 months after the original diagnosis. One hypothesis is that the autoantibody coated platelets are bound by surface Fcγ receptors in the reticuloendothelium system, which then target these platelets for destruction. Although the FcγRIIIA receptor usually shows weak binding, the phenylalanine (F) to valine (V) mutation in the FcγRIIIA receptor results in a higher affinity for IgG1 and IgG3. To test the possibility that the FcγRIIIA receptor genotype is associated with increased autoantibody mediated platelet destruction in ITP, we screened more than 80 ITP patients for this single nucleotide polymorphism (SNP) using an amplicon-down format for a custom Nanochip molecular biology workstation microarray assay.
We observed a large shift in the allele frequencies of the FcγRIIIA receptor SNP (F158V) in ITP patients compared to controls (F =.36, V =.64 vs F =.70, V =.30; P&lt;.001). When we analyzed the ITP patients with regard to chronicity of ITP, we found that chronic ITP patients showed a larger V allele frequency shift (F =.45, V =.55) than acute ITP patients (F =.58, V =.42). The IL-1α-889 SNP genotypes of these ITP patients were also found to have allele frequencies significantly different from controls (C =.67, T =.33 vs C =.84, T =.16, P =.005).
FcγRIIIA SNP Genotype FF FV VV significance ITP patients 12.3 46.5 41 Controls 50.2 39.2 10.4 P&lt;0.001 IL-1 α SNP Genotype CC CT TT significance ITP patients 51.4 33.3 15.2 Controls 71 26.3 2.6 P=0.005
Additionally, we have shown that the VV genotype of FcγRIIIA receptor is associated with 100% response to IVIg treatment in childhood ITP (Tarantino et al J. Thromb. Haemost. 2005). Thus, we propose that the FcγRIIIA receptor and IL-1α-889 genotypes may be useful as potential risk factors for ITP and its response to therapy. Understanding the mechanism whereby these polymorphisms alter the incidence and course of ITP in children will provide critical information in the prevention and treatment of this syndrome.
Cyclosporine Alone for the Treatment of Early Recurrences of TTP.
Authors: Cataland Spero R. , Jin Ming , Ferketich Amy K. , Zheng X. Long , George James N. , Wu Haifeng M. 
Published: April 2020

Abstract
Patients: The 3 patients detailed in this report presented to Ohio State University Hospitals and were enrolled on our prospective study of plasma exchange as initial therapy of TTP. Given the close outpatient follow-up during the first month after the tapering of plasma exchange, 2 patients were found to have an early recurrence of their TTP. The third patient had a recurrence 8 weeks after completing 6 months of CSA after being treated with concurrent CSA and PE for a previous acute episode of TTP. She completed the planned 6 months of cyclosporine therapy but relapsed 8 weeks after discontinuing the drug. Her recurrence was characterized by numbness and tingling in her right hand in addition to laboratory findings consistent with TTP. Complete demographic and clinical details of these 3 patients are shown in Table 1.
Methods: All 3 patients received oral cyclosporine (2–3 mg/kg) daily for six months in a divided dose. None of the patients underwent plasma exchange, infusion, or transfusion of blood products of any kind. Patients were seen daily to monitor for any clinical or laboratory findings that would warrant the re-initiation of plasma exchange therapy. Remissions were defined as a normal platelet count, normalization of LDH for at least 2 consecutive days, and improvement or stabilization of any renal and neurologic abnormalities. Serial measurements of ADAMTS13 activity and inhibitors were performed at regular intervals throughout therapy and follow-up.
Results: All three patients showed clear laboratory and clinical signs of improvement within the first 7 days of therapy and met criteria for clinical remission 14, 24, and 34 days respectively from the time they began treatment with cyclosporine. The third patient that presented with numbness and tingling in her right hand also had complete resolution of her neurologic symptoms within one week of restarting cyclosporine. Pretreatment ADAMTS13 activity was determined to be 33%, 25%, and 10% in all 3 patients respectively with all 3 patients having documented inhibitors of ADAMTS13.
Conclusions: Treatment with cyclosporine alone resulted in the suppression of ADAMTS13 inhibitors and normalization of ADAMTS13 activity.
Figure Figure
All three patients have completed the planned 6 months of cyclosporine and have maintained their remission 4, 3, and 1 month respectively after discontinuing cyclosporine.
Patient Demographic and Clinical Data Laboratory Studies at the TIme CSA Initiated Age Sex Race Therapy Preceeding Recurrence Time to Relapse After Most Recent Therapy Platelet Count (150-450 K/uL) LDH (100-190 U/L) PE=Plasma Exchange, CSA=Cyclosporine, S=Corticosteroids Patient 1 48 F C PE + S 17d 113 212 Patient 2 39 F C PE 6d 135 400 Patient 3 56 F AA CSA 56d 51 418
Mutation in the Zebrafish Homolog of the RNA Helicase DHX8 Result in Hematopoietic Defects.
Authors: English Milton A. , Lei Lin , Blake Trevor , Sood Raman , Liu Paul P. 
Published: April 2020

Abstract
Hematopoiesis is a tightly regulated process that requires the coordinated efforts of many tissue-specific transcription factors. Lateral mesoderm tissue undergoes a series of restricted proliferation and differentiation that result in the production of the hematopoietic stem cells. These cells then differentiate to populate the different cell lineages. While some factors essential for this process have been defined, many remain unknown. Zebrafish offer a powerful model system in which it is possible to conduct forward phenotype-driven screening. We have undertaken a chemical mutagenesis screen using N-nitroso-N-ethylurea (ENU) in the zebrafish to uncover mutants in myelopoiesis and early hematopoiesis. Adult male fish were exposed to ENU in a protocol designed to mutagenize spermatagonia with a mutation frequency of approximately 1:1000. Mutagenized males were crossed to wild-type females to generate F1 females. Haploid embryos from F1 females were screened at 27 hours of development by RNA in situ hybridization using the myeloid-specific marker, leukocyte-specific plastin (l-plastin), and the stem cell marker, core binding factor b (cbfb). One mutant line identified from this screen, mummy, showed reduced cbfb expression and lack of l-plastin expression. Additional in situ staining has confirmed that mummy has a myeloid defect as other myeloid markers such as Lys.C, Mpo, and C/ebp1 were also significantly reduced. Mummy mutant also have a degreased expression of gata1 and bE1 globin (RBC markers) as well as the stem cell marker scl. These results suggested that the defect in mummy is at or up-stream of the HSC level. The mummy mutant embryos also suffer from widespread cell death and die around 36 hours post fertilization. Genetic mapping and positional cloning has identified the mutated gene as the zebrafish homolog of DHX8, which encodes an RNA helicase. The yeast homolog of dhx8, prp22, functions during mRNA splicing and RNA processing. Morpholino injection into wild-type embryos has confirmed that reduced expression of dhx8 can produce a similar phenotype. The wild-type zebrafish dhx8 cDNA but not the mutated dhx8 cDNA identified in mummy was able to partially rescue the mummy phenotype. Preliminary data from RT-PCR and microarray analysis suggest that mummy mutants have defects in the production of spliced transcripts of some genes but not others. Genes with decreased spliced transcripts in the mummy mutant include scl, cbfb, and gata1, which may explain the hematpoietic defects observed. Our results demonstrate that mutations in an RNA helicase involved in splicing could result in specific blockage of hematopoiesis.
Familial Erythrocytosis Associated with a Mutation in the HIF Prolyl Hydroxylase, PHD2.
Authors: Percy Melanie J. , Zhao Q. , Flores A. , Harrison C. N. , Lappin T. R.J. , Maxwell P. H. , McMullin M. F. , Lee F. S. 
Published: April 2020

Abstract
Normal erythropoiesis is precisely regulated by a negative feedback mechanism based on the oxygen status of the body. Whenever a deficit in oxygen supply is detected erythropoietin (Epo) is rapidly synthesised to increase circulating red cells. This process is under the control of the HIF transcription complex, which is composed of one alpha and one beta subunit. Although the alpha subunit is constitutively synthesized, it is maintained at a low level by continual targeting to the proteasome. Prolines 402 and 564 in the oxygen dependent degradation (ODD) domain of HIF-1α are hydroxylated in the presence of oxygen by members of the PHD family of prolyl hydroxylases. Once hydroxylated HIF-1α is captured by the von Hippel Lindau tumour suppressor gene product (VHL), ubiquitylation follows and HIF-1α is then targeted for proteasomal degradation. Defects in the hypoxia response pathway appear to be the most common cause of erythrocytosis associated with inappropriately normal or elevated serum Epo.
Several different mutations in the VHL gene have been detected in erythrocytosis individuals. Although these mutations explain a significant number of cases there remains a large cohort where the molecular defect is undefined. Thus we screened a group of such individuals for base changes in PHD 1–3. Two affected siblings and their deceased father were found to possess a heterozygous C to G change at base 950 in the coding sequence of PHD2, causing loss of proline at codon 317 and replacement with arginine. This base change was not present in their normal mother nor in 200 normal controls. Residue 317 is located in the enzyme’s active site and at a position two residues C-terminal to Asp315, which chelates the active site iron moiety. Functional analysis indicated that the Pro317Arg PHD2 variant bound to HIF-1α and HIF-2α more weakly and displayed substantially less HIF hydroxylase activity than the wild type protein. These results indicate that the Pro317Arg PHD2 mutant protein’s function will be impaired, less hydroxylation of HIF will occur, allowing more HIF to escape proteasomal degradation and to increase transcription of HIF target genes. Consequently, Epo gene expression will be elevated thereby allowing increased erythrocyte production and the development of erythrocytosis.