bluuebunny/crossref_metadata_2025_split huggingface dataset:
Co-Delivery of Sleeping Beauty Transposons Encoding FVIII and Indoleamine 2,3-Dioxygenase Attenuate Inhibitor Formation Resulting in Long-Term FVIII Expression.
Authors: Liu Li , Fletcher Bradley S. 
Published: April 2020

Abstract
Gene therapy in adult hemophilia A mice is known to generate a robust immune response against the newly produced FVIII protein. This phenomenon has been extensively reported for both viral and non-viral vectors. T cell activation promotes the proliferation of B cells that produce antibodies against FVIII (inhibitors) leading to a loss of circulating FVIII protein and activity. Approaches to attenuate this immune response have included the use of cytotoxic chemotherapeutic agents (such as cyclophosphamide) or the use of antibodies or soluble fusion proteins that prevent T cell activation (such as the fusion protein between the cytotoxic T lymphocyte-associated antigen 4 and the immunoglobulin heavy chain, CTLA-4-Ig). Recently, the mechanism by which CTLA-4-Ig exerts its effect has been elucidated and is thought to involve tryptophan catabolism by upregulation of the enzyme indoleamine 2,3-dioxygenase (IDO). Catabolites of tryptophan degradation, such as kynurenine, have been shown to block T cell proliferation and promote T cell apoptosis. Here we report that hydrodynamic co-administration of Sleeping Beauty transposons encoding both FVIII and IDO are able to attenuate, but not fully block, inhibitor formation. With this approach, long term expression of FVIII at therapeutic levels (∼10% of normal) can be achieved in adult animals without the need for additional immune suppression. Only in animals receiving FVIII and IDO together can we detect FVIII protein by western blot at 24 weeks. Serum kynurenine levels are slightly elevated after gene delivery in animals receiving the IDO gene, but fall to within normal levels by 24 weeks. These results suggest that modulation of the levels of tryptophan catabolites in vivo can influence the formation of FVIII inhibitors.
Mesenchymal Stromal Cells Genetically Engineered To Overexpress Insulin-Like Growth Factor-1 Provide Paracrine Support for Cell-Based Erythropoietin Therapy of Chronic Renal Failure-Induced Anemia.
Authors: Kucic Terrence , Copland Ian B. , Cuerquis Jessica , Coutu Daniel L. , Chalifour Lorraine E. , Gagnon Raymonde F. , Galipeau Jacques 
Published: April 2020

Abstract
Mesenchymal stromal cells (MSC) are a population of non-hematopoietic progenitors native to the bone marrow that are amenable to genetic engineering, making them attractive delivery vehicles for the in vivo production of therapeutic proteins, such as erythropoietin (Epo). We have previously demonstrated that MSC engineered to secrete Epo can be used for the long-term correction of renal failure-induced anemia [Eliopoulos et al., J Am Soc Nephrol. June 2006]. However, limited long-term transplanted cell survival compromises the efficacy of MSC-based gene therapy approaches. The current study provides evidence that co-implantation of MSC overexpressing Insulin-like growth factor-1 (IGF-I) improves MSC-based gene therapy of anemia by providing paracrine support to Epo-secreting MSC within a synthetic subcutaneous organoid. The IGF-I receptor was found to be expressed in murine MSC by RT-PCR, and protein expression was confirmed by immunoblot. We also demonstrated MSC MAPK pathway responsiveness to IGF-I stimulation in vitro and subsequent improvement of MSC survival following staurosporin-induced apoptosis. Murine MSC were transduced to overexpress either Epo or IGF-I (hereafter MSC-Epo and MSC-IGF) using retroviral vectors. MSC-Epo were subsequently admixed in a collagen matrix and implanted by subcutaneous injection in both naïve mice and a murine model of chronic renal failure, in combination with either MSC-IGF or null MSC. Mice receiving MSC-Epo in conjunction with MSC-IGF experienced a greater and significantly sustained elevation in hematocrit compared to control mice. In addition, mice co-implanted with MSC-IGF and MSC-Epo demonstrated a significant improvement in cardiac function compared to controls. In conclusion, cell-based gene therapy using co-implanted MSC-IGF represents a promising new strategy for the treatment of renal failure-induced anemia, as well as for the improvement of gene-enhanced MSC survival within implanted matrices.
Intravenous Administration of Manganese Superoxide Dismutase-Plasmid Liposomes (MnSOD-PL) in a Mouse Model Protects Against Whole Body Irradiation.
Authors: Greenberger Joel S. , Smith Tracy , Schlesselman James J. , Epperly Michael W. 
Published: April 2020

Abstract
Organ specific localized administration of manganese superoxide dismutase plasmid/liposome complexes (MnSOD-PL) confers protection of the rodent lung, esophagus, oral cavity, intestine and bladder from single fraction and fractionated ionizing irradiation by a mechanism dependent upon mitochondrial localized antioxidant effects. Intravenous injection of MnSOD-PL significantly increased survival of both male and female C57BL/6NHsd mice from the LD 50/30 whole body irradiation dose of 9.5 Gy (p = 0.0001 or 0.0059, respectively). To determine if systemic MnSOD-PL mediated improved survival at the expense of a deleterious delayed increase in development of cancer, life shortening, or neurodegenerative disease we followed all survivors. Fifty male and female C57BL/6NHsd mice per group were injected intravenously with MnSOD-PL (100 μg plasmid DNA in 0.1 ml) and irradiated with control mice to 0, 1 or 9.5 Gy whole body irradiation 24 hrs later. Moribund or dying mice were sacrificed, examined for tumors and bone marrow isolated stained with Wright Geimsa, and examined for abnormal hemogram. Small intestine was fixed in formalin, sectioned and examined for intestinal changes. Female mice pretreated with MnSOD-PL prior to 9.5 Gy (LD 50/30) had an increased survival over the first 30 days which is the time frame for death due to hematopoietic or intestinal damage with 90% survival compared to 58% for the control irradiated mice (p = 0.0059). Between 30 and 530 days there was no significant change in survival rate between MnSOD-PL treated and control irradiated 9.5 Gy or 1.0 Gy groups. Beginning at day 400, all groups have a similar death rate which can be attributed of the age of the mice. Male mice pretreated with MnSOD-PL and irradiated to 9.5 Gy had 75% survival after 30 days compared to 25% for the control mice (p &lt; 0.0001). Between 30 and 375 days after irradiation, there was no significant change in survival rate between MnSOD-PL treated compared to control irradiated 9.5 or 1.0 Gy groups. There was no detectable difference in the incidence of deaths associated with tumor (thymoma), marrow, or intestinal damage. All mice are being followed until they become moribund as a result of irradiation or old age. The data indicate, at this time, that systemic MnSOD-PL treatment is not detectably harmful to surviving total body irradiated mice.
High Frequency of Loss of Heterozygosity Due to Uniparental Disomy or Allele Deletion of Ocular Adnexal MALT-Type Lymphoma.
Authors: Asakura Yoshitaka , Ogawa Seishi , Kato Motohiro , Yamamoto Go , Maeshima Akiko , Matsuno Yoshihiro , Suzuki Shigenobu , Kaneko Akihiro , Nomoto Junko , Tanimoto Kazuki , Sekiguchi Naohiro , Maruyama Dai , Kim Sung-Won , Watanabe Takashi , Tobinai Kensei , Kobayashi Yukio 
Published: April 2020

Abstract
Majority of malignant lymphoma arising from the ocular adnexae are ocular adnexal MALT lymphomas (OAL). Several genetic abnormalities, including t(14;18)(q32;q21), trisomy 18 and trisomy 3 have been reported in OAL. However, none of them are found in more than half of cases with OAL by conventional methods. High density Single Nucleotide Polymorphism (SNP) array analysis with CNAG/AsCNAR algorithm allows high-resolution and genome-wide detection of both loss of heterozygosity (LOH) and copy number abnormality, especially uniparental disomy (UPD), without depending on the availability of paired normal DNA (Yamamoto et al, Am J Hum Genet. 2007; 81:114–26). UPD is acquired by somatic recombination and therefore not detected by conventional cytogenetic analysis or array CGH. In this study we analyzed DNA from OAL for the presence of LOH with or without copy number changes. Tissue samples from patients with OAL at our institute between 1995 and 2003 (N=32) were subject to SNP-array (250K NspI) analysis with CNAG/AsCNAR algorithm. The patients included 21 males and 11 females, and the median age of 56.5 years at the time of diagnosis (range, 15–90 years). Clinical stage was I (29 cases), II (1 case), and IV (2 cases). Trisomy of 3, 18, and 21 were found in 9, 7 and 1 cases, respectively. Overall, LOH due to UPD more than one chromosomal band were found in 16 cases (50%). Recurrent one allele deletion was detected at 6q23-24, 12p13 and 17q21 (N=15, 15 and 21, respectively). In total, 28 (82%) among 32 cases showed one or more of the above changes. Characterization of these recurrent genetic abnormalities might reveal the pathogenesis of OAL.
Cytokine-Dependent Imatinib Resistance in Mouse Bcr-Abl(+), Arf-Null Lymphoblastic Leukemia.
Authors: Williams Richard T. , Besten Willem den , Boulos Nidal , Sherr Charles J. 
Published: April 2020

Abstract
Retroviral vector-mediated transduction of the BCR-ABL tyrosine kinase into Arf-deficient mouse bone marrow progenitors allows rapid ex vivo outgrowth of pure pre-B cell populations that induce a highly aggressive form of ALL when inoculated intravenously into healthy syngeneic mice. These leukemias resist therapy with the BCR-ABL kinase inhibitor imatinib, and surprisingly, drug resistance is non-tumor-cell autonomous. We now demonstrate that expression of the p185 BCR-ABL isoform (hereafter p185) in Arf-null pre-B cells is sufficient to generate polyclonal populations of leukemia-initiating cells (LICs), only 20 of which induce fatal, imatinib-resistant disease within one month after infusion. These p185+ Arf-null LICs are pre-B cells by immunophenotypic criteria, lack evidence of myeloid and stem cells markers, exhibit immunoglobulin heavy chain gene rearrangements and random proviral insertions, and are almost universally capable of initiating lethal, transplantable leukemias in healthy syngeneic recipient mice. Therefore, these LICs do not represent rare ‘cancer stem cells’. Resistance to targeted therapy with imatinib in this model system is not dependent upon mutations within the BCR-ABL kinase domain, but, rather, is mediated through interactions between the LICs and the host environment. While cytokines like IL-7 are dispensable for the efficient generation of p185+, Arf-null cells, these LICs remain IL-7-responsive and are significantly protected from imatinib-induced cytostasis by IL-7 in vitro. Introduction of p185 into Arf-null hematopoietic progenitors that also lack the common gamma chain (γc) for cytokine receptors bypasses their cytokine requirements and permits pre-B cell development. These p185+, Arf-null, γc-null LICs also initiate ALL but exhibit much greater sensitivity to imatinib in vivo, allowing a significant fraction of treated mice to sustain long term remissions even when therapy is discontinued. Therefore, salutary cytokines expressed in the hematopoietic microenvironment facilitate leukemic proliferation and confer resistance to targeted therapy. Not surprisingly, single agent therapy with dasatinib, a second generation BCR-ABL kinase inhibitor, is dramatically more efficacious in this pre-clinical Ph+ ALL model, although some recipient mice still succumb to disease despite continuous drug treatment. Using vectors co-expressing p185 and luciferase, we are now initiating dasatinib therapy when recipient animals have detectable disease burdens, thereby permitting a more dynamic assessment of therapeutic response and subsequent failure. We speculate that in human Ph+ ALL, the frequent deletion of the INK4A-ARF locus (∼50% of cases at diagnosis) might help to maintain LIC survival in the face of targeted therapy and so facilitate the emergence of drug-resistant BCR-ABL variants. As such, INK4A-ARF deletion might prove to be a poor prognostic indicator.
The MMSET Gene Contributes to the Growth, Adhesion, and Tumorigencitiy of t(4;14) Multiple Myeloma Cells.
Authors: Lauring Josh , Abukhdeir Abde M. , Konishi Hiroyuki , Garay Joseph P. , Gustin John P. , Wang Qiuju , Arceci Robert J. , Matsui William , Park Ben H. 
Published: April 2020

Abstract
Multiple myeloma (MM) is an incurable hematological malignancy characterized by recurrent chromosomal translocations. The t(4;14)(p16;q32) is associated with the worst prognosis of any patient subgroup in MM, although the basis for this poor prognosis is unknown. The t(4;14) is unusual in that it involves two potential target genes on chromosome 4: fibroblast growth factor receptor 3 (FGFR3) and multiple myeloma SET domain (MMSET). MMSET is universally over-expressed in t(4;14) MM, whereas FGFR3 expression is lost in one third of cases, suggesting a role for MMSET in myeloma pathogenesis. Nonetheless, the role of MMSET in t(4;14) MM has remained unclear. Here we demonstrate a role for MMSET in t(4;14) MM cells. Using homologous recombination-mediated gene targeting, we disrupted the N-terminal and full-length isoforms of MMSET in t(4;14)+ KMS-11 MM cells. Disruption of the translocated MMSET allele revealed that this allele accounts for most of the MMSET transcription in t(4;14) MM cells. Accordingly, selective targeting of the translocated allele, but not the non-translocated allele, led to reduced colony formation in methylcellulose and reduced tumor formation in nude mouse xenografts. Down-regulation of MMSET expression in t(4;14) MM cell lines by stable RNA interference (RNAi) led to a slower growth in liquid culture, a significant reduction in colony formation in methylcellulose, and decreased tumorigenicity in vivo. Additionally, MMSET knockdown led to partial cell cycle arrest of adherent MM cells and reduced the ability of MM cells to adhere to extracellular matrix. Cells with targeted disruption or knockdown of MMSET exhibited changes in expression levels of potential target genes, including several adhesion molecules. These results provide the first direct evidence that translocation-mediated overexpression of MMSET plays a critical role in t(4;14) MM and suggest that therapies targeting this gene could impact this particular subset of poor-prognosis patients.
Constitutive Expression of Lef-1 in Murine Hematopoietic Progenitors Causes Myeloid and Lymphoid Leukemias Propagated by a Leukemic Stem Cell Population with Lymphoid Characteristics.
Authors: Petropoulos Konstantin , Schessl Christina , Arseni Natalia , Rawat Vijay P.S. , Deshpande Aniruddha , Stadler Christiane , Hiddemann Wolfgang , Quintanilla-Martinez Leticia , Bohlander Stefan K. , Feuring-Buske Michaela , Buske Christian 
Published: April 2020

Abstract
Canonical Wnt signaling is critically involved in normal hematopoietic development and the self-renewal process of hematopoietic stem cells. Deregulation of this pathway has been linked to a large variety of cancers including leukemia. Lef-1, a key factor of the Wnt / beta-catenin signaling pathway, plays pivotal roles in lymphoid development, but little is known about the role of Lef-1 in myeloid hematopoiesis and leukemogenesis. We now show that Lef-1 is expressed in murine hematopoietic stem cells (‘LSK’, Sca+cKit+Lin−) and both myeloid and lymphoid subpopulations as well as in different human leukemias. Using a retroviral BM transplantation model, we demonstrate that ectopic expression of wild type Lef-1 (WT) and a constitutive active mutant of Lef-1 (CA) induces a severe disturbance of normal hematopoietic development: mice transplanted with bone marrow constitutively expressing Lef-1 had a significant increase in the number of circulating myeloid cells resulting in an inverted lymphoid-myeloid ratio in the peripheral blood (ratio: 0.28 (WT), 0.10 (CA) vs. 1.07 (GFP); p&lt;0.002). With a median latency of 12 month, transplanted mice succumbed to a lethal myeloproliferation (n=2) or acute myeloid or B-lymphoblastic leukemias (N=8). Both leukemia subtypes shared key biological characteristics such as positivity for IG DH-JH rearrangements. In addition, both subtypes were characterized by a biphenotypic cell population (B220+Mac1+Gr1+, BMG+++) as well as varying numbers of B220+Mac1−Gr1− (B+MG−) cells. In both leukemia subtypes single DJH–rearranged B+MG− cells had the highest seeding efficiency and were able to give rise to both BMG+++ and B−MG+ cells in vitro (mean seeding efficiency: B+MG−: 17.9%, vs. BMG+++: 4.7%, B−MG+: 4.0, p=0.005). Strikingly, the frequency of leukemia-initiating cells in AML was highest in the B+MG− population as determined by limit dilution transplantation assays (B+MG−: 1 in 433 vs. BMG+++/B−MG+: 1 in 8491 cells, p&lt;0.001). Expression analysis of malignant blast cells from both lymphoid and myeloid leukemias also revealed striking commonalities with regard to the transcription profile. Blast cells from both AML and ALL diseased mice uniformly showed expression of myelo-specific genes like C/ebpα and c-fms and lymphoid specific genes E2A and Ebf-1, but lacked expression of the B-cell differentiation regulator Pax-5. These findings demonstrate that balanced expression of Lef-1 is crucial for early normal hematopoietic differentiation and that deregulation of this factor induces the development of DJH-rearranged acute myeloid and lymphoid leukemias which are propagated by a leukemic stem cell with lymphoid characteristics.
PML Protects HIPK2 and p300 from SCF-Mediated Ubiquitin-Dependent Degradation To Activate Transcription.
Authors: Shima Yutaka , Shima Takito , Chiba Tomoki , Irimura Tatsuro , Kitabayashi Issay 
Published: April 2020

Abstract
The Pml gene is the target of t(15;17) chromosome translocation in acute promyelocytic leukemia. PML protein is known to localize in discrete nuclear speckles, named PML nuclear bodies (NBs). In NBs, PML interacts with several transcription factors, such as p53 and AML1, and their co-activators, such as HIPK2 and p300. PML activates transcription of their target genes. PML is thought to stabilize transcription factor complex and function as a mediator in transcription activation, but little is known about the molecular mechanism by which PML activates transcription. To clarify the role of PML in transcription regulation, we purified the PML complex and identified a novel F-box protein (FBP), Skp1, and Cullin1 (Cul1) in the PML complex by LC/MS/MS analysis. FBPs form SCF ubiquitin ligase complexes with Skp1, Cul1 and ROC1 and mediate recognition of specific substrates for ubiquitination. We found that the FBP that we identified here also forms a SCF complex with Skp1, Cul1 and ROC1. To identify substrates for the SCF complex, we tested several proteins that could bind to PML, and found that the FBP promotes degradation of HIPK2 and p300. These degradations were inhibited in the presence of a proteasome inhibitor, MG132. The FBP stimulated ubiquitination of HIPK2. These results suggest that the SCF promotes degradation of these proteins by the ubiquitin-proteasome pathway. The fact that the SCF is a part of the PML complex suggests that PML plays a role in the SCF-mediated degradation of HIPK2 and p300 by the ubiquitin-proteasome pathway. In order to clarify the role of PML in degradation of HIPK2 and p300, we tested effects of PML on the degradation and found that PML inhibited the SCF-mediated degradation of HIPK2 and p300 without inhibition of ubiquitination. To clarify roles of HIPK2, PML IV and the FBP in p53-dependent transcription, we performed reporter analysis using the MDM2 promoter in H1299 cells. Since the FBP promotes degradation of HIPK2, we initially thought that the FBP might inhibit activation of p53-dependent transcription by HIPK2 and PML IV. However, the FBP, HIPK2 and PML synergistically stimulated the p53-dependent transcriptional activation. Taken together our data suggest that the SCF-induced ubiquitination of transcription co-activators HIPK2 and p300 plays a critical role in transcriptional regulation, and that PML stimulates transcription by protecting HIPK2 and p300 from ubiquitin-dependent degradation.
Novel Tumor Suppressor Protein Programmed Cell Death 4 (PDCD4) Suppresses Activity of PI3K/Akt Pathway and Regulates Expression of p27 (Kip1) and c-myc, DAP5 and Willm’s Tumor (WT1) in Acute Myeloid Leukemia.
Authors: Ozpolat Bulent , Akar Ugur , Colburn Nancy H. , Lopez-Berestein Gabriel 
Published: April 2020

Abstract
Programmed cell death 4 (PDCD4) is a recently identied novel tumor suppressor protein that inhibits cap-dependent mRNA translation. PDCD4 inhibits tumor promoter incuced carcinogenesis and transformation by suppressing the helicase activity of eIF4A, leading to translational inhibition. Recently we found that PDCD4 is required for all-trans-retinoic acid (ATRA)-induced granulocytic differentiation of acute promyelocytic leukemia (APL) cells (Ozpolat&Akar et al, Mol Cancer Res, in press), type of acute myeloid leukemia characterized by a t(15;17) and a differentiation block. Here we investigated the downstream mediators or targets of PDCD4 in leukemia cell differentiation. ATRA is currently used as a first line standard treatment in APL. Recently, we reported that ATRA induces translational suppression through multiple posttranscriptional mechanisms that involve suppression of translation initiation, a rate limiting step of protein synthesis (Harris&Ozpolat et al, Blood, 104 (5) 2004). We found that ATRA treatment induced PDCD4 expression in NB4 APL, HL60 AML, primary APL patient leukemia cells and normal human CD34+ bone marrow progenitors cells during granulocytic differentiation. However, ATRA/maturation resistant NB4.R1 and HL60R cells failed to express and translocate PDCD4 into nucleus after ATRA treatment. To identify downstream targets of PDCD4 we knock downed PDCD4 expression by siRNA and examined changes in the expression of target proteins that are known to be regulated by ATRA by Western blot analysis in NB4 cells. We found that PDCD4 represses c-myc, and WT1 expression however, it is required for the expression of DAP5 (death associated protein 5), and cyclin dependent kinase inhibitor p27KIP1, and but not c-jun, p21Cip1, and tissue transglutaminase (TG2). Inhibition of PDCD4 by siRNA resulted in upregulation of phospho-P70S6K, suggesting that PDCD4 inhibits activity of PI3K/Akt pathway. RT-PCR analysis revealed that mRNA of these proteins did not change suggesting that PDCD4 tumor suppressor protein regulates expression of these important cellular proteins at translational level and suppresses PI3K/Akt pathway. Furthermore we rapamycin, a specific mTOR inhibitor currently in clinical trials in AML, induced a marked expression of PDCD4, which regulates c-myc and p27 Kip1, revealing a novel mechanism of action of rapamycin, providing new rationale for targeting translational pathways as a therapeutic intervention in the treatment of AML. Overall, data suggest that PDCD4 regulates expression of critical cellular proteins involved in differentiation of leukemia cells and PDCD4 mediated translational control may be an important regulatory mechanism for regulation of gene expression.