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71.
Hyun‐Joo Lee MinSeok Chang Jong‐Mook Kim HyeJin Hong KiEun Maeng Jane Koo ShinJae Chang Myung‐Sam Cho 《Biotechnology progress》2013,29(2):432-440
Host cell lines developed by genetic engineering sometimes show instabilities in maintaining their genetically acquired phenotypes. Previously, a hybrid host cell line, designated as hybrid of kidney and B cells (HKB), capable of retaining selected phenotypes originally existing in the parental cells was developed via fusion of 293 cells and HH514‐16 cells. Although HKB did indeed successfully preserve several favorable phenotypes, the expression of Epstein‐Barr virus (EBV) specific nuclear antigen 1 (EBNA1), which should be constitutively expressed for host cells to utilize oriP expression vector in transient production of therapeutic proteins, was observed to be unstable. Here, in an attempt to obtain stable expression of EBNA1, a cell type that contains an integrated EBV genome, rather than HH514‐16 cells, which harbor an episomal EBV genome, was applied for fusion with 293 cells. Fusion of 293 cells with Namalwa cells led to the creation of a new type of hybrid, F2N, which was able to stably express EBNA1 while not producing EBV particles. One of the F2N clones, F2N78, was observed to maintain EBNA1 expression for more than 1 year under serum‐free suspension culture conditions along with human specific glycosyl phenotypes observed previously in HKB. In addition, F2N78 was demonstrated to be an appropriate host cell line for both the transient and stable production of recombinant therapeutics with the features of safety expected of production cell lines for human use. © 2013 American Institute of Chemical Engineers Biotechnol. Prog., 29: 432–440, 2013 相似文献
72.
Roxanne Dutia Andrea J. Kim Matthew Modes Robert Rothlein Jane M. Shen Ye Edward Tian Jumana Ihbais Sam F. Victory Carmen Valcarce Sharon L. Wardlaw 《PloS one》2013,8(6)
Activation of brain melanocortin-4 receptors (MC4-R) by α-melanocyte-stimulating hormone (MSH) or inhibition by agouti-related protein (AgRP) regulates food intake and energy expenditure and can modulate neuroendocrine responses to changes in energy balance. To examine the effects of AgRP inhibition on energy balance, a small molecule, non-peptide compound, TTP2515, developed by TransTech Pharma, Inc., was studied in vitro and in rodent models in vivo. TTP2515 prevented AgRP from antagonizing α-MSH-induced increases in cAMP in HEK 293 cells overexpressing the human MC4-R. When administered to rats by oral gavage TTP2515 blocked icv AgRP-induced increases in food intake, weight gain and adiposity and suppression of T4 levels. In both diet-induced obese (DIO) and leptin-deficient mice, TTP2515 decreased food intake, weight gain, adiposity and respiratory quotient. TTP2515 potently suppressed food intake and weight gain in lean mice immediately after initiation of a high fat diet (HFD) but had no effect on these parameters in lean chow-fed mice. However, when tested in AgRP KO mice, TTP2515 also suppressed food intake and weight gain during HFD feeding. In several studies TTP2515 increased T4 but not T3 levels, however this was also observed in AgRP KO mice. TTP2515 also attenuated refeeding and weight gain after fasting, an effect not evident in AgRP KO mice when administered at moderate doses. This study shows that TTP2515 exerts many effects consistent with AgRP inhibition however experiments in AgRP KO mice indicate some off-target effects of this drug. TTP2515 was particularly effective during fasting and in mice with leptin deficiency, conditions in which AgRP is elevated, as well as during acute and chronic HFD feeding. Thus the usefulness of this drug in treating obesity deserves further exploration, to define the AgRP dependent and independent mechanisms by which TTP2515 exerts its effects on energy balance. 相似文献
73.
Mindy I. Davis Atsuo T. Sasaki Min Shen Brooke M. Emerling Natasha Thorne Sam Michael Rajan Pragani Matthew Boxer Kazutaka Sumita Koh Takeuchi Douglas S. Auld Zhuyin Li Lewis C. Cantley Anton Simeonov 《PloS one》2013,8(1)
Phosphoinositide kinases regulate diverse cellular functions and are important targets for therapeutic development for diseases, such as diabetes and cancer. Preparation of the lipid substrate is crucial for the development of a robust and miniaturizable lipid kinase assay. Enzymatic assays for phosphoinositide kinases often use lipid substrates prepared from lyophilized lipid preparations by sonication, which result in variability in the liposome size from preparation to preparation. Herein, we report a homogeneous 1536-well luciferase-coupled bioluminescence assay for PI5P4Kα. The substrate preparation is novel and allows the rapid production of a DMSO-containing substrate solution without the need for lengthy liposome preparation protocols, thus enabling the scale-up of this traditionally difficult type of assay. The Z’-factor value was greater than 0.7 for the PI5P4Kα assay, indicating its suitability for high-throughput screening applications. Tyrphostin AG-82 had been identified as an inhibitor of PI5P4Kα by assessing the degree of phospho transfer of γ-32P-ATP to PI5P; its inhibitory activity against PI5P4Kα was confirmed in the present miniaturized assay. From a pilot screen of a library of bioactive compounds, another tyrphostin, I-OMe tyrphostin AG-538 (I-OMe-AG-538), was identified as an ATP-competitive inhibitor of PI5P4Kα with an IC50 of 1 µM, affirming the suitability of the assay for inhibitor discovery campaigns. This homogeneous assay may apply to other lipid kinases and should help in the identification of leads for this class of enzymes by enabling high-throughput screening efforts. 相似文献
74.
Werner Scheuer Markus Thomas Petra Hanke Johannes Sam Franz Osl Diana Weininger 《MABS-AUSTIN》2016,8(3):562-573
Vascular endothelial growth factor (VEGF)-A blockade has been validated clinically as a treatment for human cancers. Angiopoietin-2 (Ang-2) is a key regulator of blood vessel remodeling and maturation. In tumors, Ang-2 is up-regulated and an unfavorable prognostic factor. Recent data demonstrated that Ang-2 inhibition mediates anti-tumoral effects. We generated a tetravalent bispecific antibody (Ang-2-VEGF-TAvi6) targeting VEGF-A with 2 arms based on bevacizumab (Avastin®), and targeting Ang-2 with 2 arms based on a novel anti-Ang-2 antibody (LC06). The two Ang-2-targeting single-chain variable fragments are disulfide-stabilized and fused to the C-terminus of the heavy chain of bevacizumab. Treatment with Ang-2-VEGF-A-TAvi6 led to a complete abrogation of angiogenesis in the cornea micropocket assay. Metastatic spread and tumor growth of subcutaneous, orthotopic and anti-VEGF-A resistant tumors were also efficiently inhibited. These data further establish Ang-2-VEGF bispecific antibodies as a promising anti-angiogenic, anti-metastatic and anti-tumor agent for the treatment of cancer. 相似文献
75.
Matthew J. Billard David J. Fitzhugh Joel S. Parker Jaime M. Brozowski Marcus W. McGinnis Roman G. Timoshchenko D. Stephen Serafin Ruth Lininger Nancy Klauber-Demore Gary Sahagian Young K. Truong Maria F. Sassano Jonathan S. Serody Teresa K. Tarrant 《PloS one》2016,11(4)
Triple negative breast cancer (TNBC) is a heterogeneous disease that has a poor prognosis and limited treatment options. Chemokine receptor interactions are important modulators of breast cancer metastasis; however, it is now recognized that quantitative surface expression of one important chemokine receptor, CXCR4, may not directly correlate with metastasis and that its functional activity in breast cancer may better inform tumor pathogenicity. G protein coupled receptor kinase 3 (GRK3) is a negative regulator of CXCR4 activity, and we show that GRK expression correlates with tumorigenicity, molecular subtype, and metastatic potential in human tumor microarray analysis. Using established human breast cancer cell lines and an immunocompetent in vivo mouse model, we further demonstrate that alterations in GRK3 expression levels in tumor cells directly affect migration and invasion in vitro and the establishment of distant metastasis in vivo. The effects of GRK3 modulation appear to be specific to chemokine-mediated migration behaviors without influencing tumor cell proliferation or survival. These data demonstrate that GRK3 dysregulation may play an important part in TNBC metastasis. 相似文献
76.
77.
Midpoint attractors and species richness: Modelling the interaction between environmental drivers and geometric constraints
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Robert K. Colwell Nicholas J. Gotelli Louise A. Ashton Jan Beck Gunnar Brehm Tom M. Fayle Konrad Fiedler Matthew L. Forister Michael Kessler Roger L. Kitching Petr Klimes Jürgen Kluge John T. Longino Sarah C. Maunsell Christy M. McCain Jimmy Moses Sarah Noben Katerina Sam Legi Sam Arthur M. Shapiro Xiangping Wang Vojtech Novotny 《Ecology letters》2016,19(9):1009-1022
We introduce a novel framework for conceptualising, quantifying and unifying discordant patterns of species richness along geographical gradients. While not itself explicitly mechanistic, this approach offers a path towards understanding mechanisms. In this study, we focused on the diverse patterns of species richness on mountainsides. We conjectured that elevational range midpoints of species may be drawn towards a single midpoint attractor – a unimodal gradient of environmental favourability. The midpoint attractor interacts with geometric constraints imposed by sea level and the mountaintop to produce taxon‐specific patterns of species richness. We developed a Bayesian simulation model to estimate the location and strength of the midpoint attractor from species occurrence data sampled along mountainsides. We also constructed midpoint predictor models to test whether environmental variables could directly account for the observed patterns of species range midpoints. We challenged these models with 16 elevational data sets, comprising 4500 species of insects, vertebrates and plants. The midpoint predictor models generally failed to predict the pattern of species midpoints. In contrast, the midpoint attractor model closely reproduced empirical spatial patterns of species richness and range midpoints. Gradients of environmental favourability, subject to geometric constraints, may parsimoniously account for elevational and other patterns of species richness. 相似文献
78.
Kyu‐Won Kwak Hong‐Soo Choi Sung‐Hee Nam Myung‐Sae Han Eunsun Kim Kwanho Park Heui‐Sam Lee 《Entomological Research》2019,49(12):534-538
This report reveals the structure of a virus extracted from the Korean horn beetle Allomyrina dichotoma. The purified virus particle was 100% identical to Allomyrina virus lef‐8 sequence registered as KM_233709.1. The structure of this virus was confirmed to be closely related to that of the Nudiviridae family, and it was rod shaped and enveloped, and observed to be of approximately the mean length of a single viral nucleocapsid of 200–210 nm and mean diameter of 100–110 nm. These results provide an insight into the structural characteristics of the Nudiviridae family that can be used for nudiviral identification. 相似文献
79.
80.
Olusegun O. Osunkoya Jens G. Froese Sam Nicol Christine Perrett Kerri Moore Jason Callander Shane Campbell 《Austral ecology》2019,44(7):1123-1138
Invasive alien plant species threaten agriculture and biodiversity globally and require ongoing management to minimise impacts. However, the large number of invasive species means that a risk‐based approach to prioritisation is needed, taking into account the spatial scale of management decisions and myriad of available information. Here, we developed a risk‐based inventory of invasive plants in Queensland, Australia, using both current species distribution/abundance and the severity of their impacts. Our assessment followed a comprehensive data collection process including a scoping of local government pest management plans, herbarium records, the published literature and structured elicitation of expert knowledge during a series of regional stakeholder workshops. From ~300 plant species that were identified as established and/or emerging invaders in the State, only one‐third were considered by practitioners to pose significant risks across regions to be considered management priorities. We aggregated regional species lists into a statewide priority list and analysed the data set (107 species) for historical, geographical, floristic and ecological patterns. Regions on the mainland eastern seaboard of the State share similar invasive plant communities, suggesting that these regions may form a single management unit, unlike the western/inland and the extreme far north (Torres Strait Islands) regions, which share fewer invasive plant species. Positive correlations were detected between invasiveness and time since introduction for some but not all plant life forms. Stakeholders identified research and management priorities for the invasive plant list, including biological control options, public awareness/education, effective herbicide use, ecology/taxonomy and risk analysis. In the course of the exercise, a statewide invasive plant priority list of high‐, medium‐ and low‐impact scores for policy, research and management was compiled. Finally, our approach to invasive plant species prioritisation highlighted that planning and policy documents are not necessarily reflected at the grass‐root level in terms of species identity and management priorities. 相似文献