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cytotoxicity evaluation of anatase and rutile tio₂ thin films on cho-k1 cells in vitro.cytotoxicity of titanium dioxide (tio₂) thin films on chinese hamster ovary (cho-k1) cells was evaluated after 24, 48 and 72 h of culture. the tio₂ thin films were deposited using direct current magnetron sputtering. these films were post-deposition annealed at different temperatures (300, 500 and 800 °c) toward the anatase to rutile phase transformation. the root-mean-square (rms) surface roughness of tio₂ films went from 2.8 to 8.08 nm when the annealing temperature was increased from 300 to 8 ...201628773740
[effect of low molecular weight polyacrylic acid on normal and gamma-irradiated chinese hamster cells]. 20167192423
potent inhibition of herg channels by the over-the-counter antidiarrheal agent loperamide.the aim of this study was to determine the in vitro electrophysiological properties of loperamide. the authors' hypothesis was that loperamide is a potent blocker of the current carried by the human ether-à-go-go-related gene (herg) potassium channel.201629759761
experimentally-induced animal models of prediabetes and insulin resistance: a review.animal models are very common in diabetes research when rodents are mostly used in this regard. although a number of animal models of type 1 and type 2 diabetes (t2d) are developed in last few decades, the numbers of animal models of prediabetes and insulin resistance are very scanty. due to the rapidly changing pathogenesis of the disease and preventive study, the popularity of the prediabetic and insulin resistance models are largely increased in the recent years. some genetically or spontaneo ...201629648707
impedance-based analysis of mu opioid receptor signaling and underlying mechanisms.the mu opioid receptor is a g-protein coupled receptor able to signal through the gαi/o class of g-protein and β-arrestin pathways, stimulating down-stream effector pathways. signaling bias occurs when different receptor agonists lead to different signaling outcomes. traditionally these have been studied using end-point assays. real-time cellular analysis platforms allow for the analysis of the holistic effects of receptor activation as an integrated output. while this allows for different ligan ...201630338303
[subunit specific modulation of glycine receptors by ginkgolic acid.]ginkgo biloba extract is a multicomponent pharmacological agent widely used in neurological disorders therapy. it was shown that ginkgolic acid, a constituent of lipophylic ginkgo biloba extract, has numerous biological activities. in the present study we have focused on the features of ginkgolic acid action on αl and α2 glycine receptors that make part of the inhibitory system of the brain. using whole-cell configuration of patch-clamp recording we analysed effects of ginkgolic acid on differen ...201629975473
[effects of low radiation doses on chinese hamster cells].the induction of cytogenetic damages after irradiation with single dose of gamma-rays (0.1-2 gy) have been studied. it is shown non-linear curve for the induction of chromosome aberrations, detected by anaphase method. after irradiation in s-stage of the cell cycle at dose below 0.2 gy the cells were more radiosensitive than after irradiation with doses 0.3-2 gy. between the phases of high radiosensitivity and radioresistance the reversal dose-effect relation was observed. this phenomenon was no ...20169889779
[characterization of the adaptive response to the action of gamma-rays, induced by low doses of 14c in chinese hamster fibroblasts].it has been studied the correlation of the mitotic activity of the chromosome aberrations and apoptosis, in the v-79 cells pre-exposure to an adapting dose of ionizing radiation from 14c-thymidine prior to an acute challenge dose of gamma-rays. in spite of that the incubation of the cells with isotope increased of the yield of the chromosome aberrations, but the cells became more resistant to following gamma-irradiation. increasing the adaptive dose of the 14c on degree didn't influence on the p ...20169876490
microsatellite instability is co-selectable with gene amplification in a mammalian mutator phenotype.the effect of an unbalanced nucleotide pool on the stability of dinucleotide (ca)n microsatellite sequences was investigated in the mutator phenotype csa7 clone isolated from chinese hamster chef18 cell line. a series of clones isolated from chef18 and csa7 cells and resistant to 6-thioguanine (tg) were shown to be stable at the three examined microsatellite loci. furthermore, the clones isolated from chef18 cells and resistant to n-phosphonacetyl-l-aspartate (pala) were stable, whereas those is ...201610368687
targeting hypoxia in tumors using 2-nitroimidazoles with peptidic chelators for technetium-99m: effect of lipophilicity.tumor hypoxia is an important prognostic factor for response to therapy. radiolabeled 2-nitroimidazoles have been used for imaging hypoxia, and the octanol/water partition coefficient (p) of these compounds appears to play a crucial role in their suitability for imaging. a series of 11 2-nitroimidazoles coupled to peptidic chelators for (99m)tc with divergent p was developed and evaluated in an in vitro system. two classes of n(3)s chelators were used: dialkyl-gly-ser-cys-linker-2-nitroimidazole ...201610821657
selection of n-(phosphonacetyl)-l-aspartate resistant chinese hamster mutants in the presence of the uridine uptake inhibitor dipyridamole.in mammalian cells selected in culture for resistance to pala the cad gene is amplified and these cells are a widely used model system to study gene amplification. selection of resistant mutants is routinely performed in medium supplemented with dialyzed serum, because the cytotoxic effect of pala is reversed by uridine, which is contained in serum. we have shown that in chinese hamster cells dipyridamole reduced uridine uptake to less than 5% with limited effect on cell survival. moreover, in m ...20167733632
inhibition of methotrexate-induced chromosomal damage by vanillin and chlorophyllin in v79 cells.methotrexate (mtx), a chemotherapeutic agent used to treat cancer, produces cytogenetic damage and has a cytostatic effect in a variety of test systems. several antigenotoxic agents have been studied in various in vitro and in vivo systems. however, data are limited regarding their ability to modulate mtx-induced genotoxicity. in the present study, vanillin (va) and chlorophyllin (chl) were used as antigenotoxic agents to study their ability to minimize the dna damage caused by mtx. exponentiall ...20169485540
[the mutagenic action of gamma radiation on chinese hamster cells. the cytogenetic characteristics of the mutants at the hprt locus].the peculiarities of mutation induction and cytogenetic characteristics of spontaneous and radiation-induced hprt mutant clones have been studied. the linear-quadratic dependence of the mutation induction on radiation dose has been found. high heterogeneity of cytogenetic parameters (aneuploidy and chromosome aberration frequency) has been shown in the mutants.20169026291
[the lethal and mutagenic actions of radiations with different lets on mammalian cells].inactivation and induction of mutations in the hgprt locus in chinese hamster cells after irradiation with accelerated heavy ions in the let range of 20 to 367 kev/micron were studied. in both cases, inactivation and induction of mutations, the let dependence of rbe is described by a curve with a local maximum in the range of 80 to 100 kev/micron. the maximum rbe value for the mutagenic action is almost twice as high as that for inactivation. however, the rbe coefficients of the mutation inducti ...20169181964
[the antimutagenic activity of compounds of modified polyglucin].the effect of modified polygluquin compounds on gamma-irradiated nuclei of v-79 chinese hamster cells has been studied. antimutagenic properties of the compounds are determined by aldehyde groups and oxidized chains.20161703176
[antimutagenic activity of dextran gammaphos derivatives].in experiments with v-79 chinese hamster cell culture the influence of dextran gammaphos derivatives on the mutagenic effects of gamma-radiation was studied by the number of cells with micronuclei and fragmented nuclei. products of interaction between gammaphos and dialdehyde dextran were shown to have a higher antimutagenic activity than gammaphos.20161719580
hyperthermic sensitization by the radical initiator 2,2'-azobis (2-amidinopropane) dihydrochloride (aaph). i. in vitro studies.aaph (2,2'-azobis-(2-amidinopropane dihydrochloride)) is a water-soluble, heat-labile azo compound which undergoes thermal decomposition to produce carbon-centred free radicals. these carbon-centred radicals might be directly cytotoxic or may react with oxygen to produce potentially cytotoxic alkoxyl and peroxyl radicals. the rate of free radical production as a result of aaph thermal decomposition increases with increasing temperature. we have evaluated the efficacy of aaph as a heat sensitizer ...20168064185
[effect of chronic neutron irradiation on fibroblast cell culture in chinese hamster].the effect of monoenergy (30 gev) neutron radiation on chinese hamster fibroblasts has been studied by micronuclear test. it has been shown that in the range of doses up to 60 cgy rbe is equal to unity.20168293117
[stimulation of fibroblasts' growth in culture by low-dose ionizing radiation in acute irradiation].the effect of low irradiation doses (10-100 cgy) on the growth of chinese hamster fibroblasts was examined. it was shown that the number and the mean size of colonies increased after irradiation with doses of 10-40 cgy. dependence of the effect on dose is extremal with maximum at 10-20 cgy. irradiation by dose of 10 cgy did not affect on the cell cycle duration but significantly shortened the las-phase of growth.20168293114
embryotoxicity and mutagenicity of mycotoxins.embryotoxicity: aflatoxin b1 (afb1), g1 (afg1), and patulin (pa) were investigated in nmri mice for embryotoxic and teratogenic activity. these three mycotoxins were injected intraperitoneally or given orally on day 12 and 13 of pregnancy. afb1 (15, 45, and 90 mg/kg ip or 45 mg/kg po) produced moderate retardation in fetal development and a dose-related increase of cleft palates, wavy ribs, and diaphragm changes. the effects after injection of afg1 (45 to 90 mg/kg ip) were reduction of fetal wei ...20162121956
the pathobiology of ozone-induced damage.ozone remains one of the three most important air pollutants worldwide, yet little direct documented evidence of its genotoxicity exists. the interest in the pathology of ozone exposure and the molecular events that underlie its course stems from dna damage caused by oxygen stress including hydroxyl radicals, superoxide, singlet oxygen, and hydrogen peroxide. although the tissue damage associated with ozone inhalation occurs at both the conducting airway and the alveolus, the cellular and mechan ...20162185706
deoxyribonucleotide pools as targets for mutagenesis by n-methyl-n-nitrosourea.this paper describes a biological test of the hypothesis that one or more components of the intracellular nucleotide pool represent a significant target for the mutagenic effects of alkylating agents. in other words, we ask whether mutagenesis can occur either through alkylation of susceptible nucleotide residues in dna, or through alkylation of a free nucleotide, followed by its incorporation into dna. our approach is based upon the premise that if a nucleotide pool is a mutagenic target, then ...20163260658
induction of cad gene amplification by restriction endonucleases in v79,b7 chinese hamster cells.the restriction endonucleases pvuii, bamhi and ecori were tested for their ability to induce gene amplification in v79,b7 chinese hamster cells. the results indicate that treatment with these enzymes efficiently increases the frequency of clones resistant to n-phosphonacetyl-l-aspartate, indicating induction of cad gene amplification.20162563307
the effect of l-2-oxothiazolidine on glutathione levels in cultured mammalian cells.chinese hamster ovary cells were cultured with increasing concentrations (7 x 10(-8) m-7 x 10(-2) m) of l-2-oxothiazolidine in an attempt to increase intracellular glutathione levels in these multiply-passaged mammalian cells. in a series of 7 expts., intracellular glutathione levels increased significantly at the highest concentrations tested and were protective against oxygen radical-induced genetic toxicity.20163627156
effect of heavy metals on cellular growth, metabolism and integrity of cultured chinese hamster kidney cells.the effects of cdcl2, na2cro4, naaso2 and niso4 on cultured chinese hamster kidney cells were studied over a concentration range for 48 h. release of lactate dehydrogenase, a parameter of cell viability, was not closely related to cell proliferation except for na2cro4. a better correlation was obtained between glucose consumption and lactate production, and cellular growth. when studying toxic effects of metals in cell-culture systems, metabolic parameters should be determined in addition to cel ...20163840935
misonidazole increases the toxicity of bcnu for hypoxic cells.misonidazole (miso) increased the toxicity of bcnu for hypoxic chinese hamster ovary (cho) cells in vitro, but had no effect on aerobic toxicity of bcnu. survival was reduced by up to 10(-3) with an exposure to miso (imm x 4 hr) that had no direct toxicity for hypoxic cells. preincubation of cells with miso under hypoxic conditions followed by exposure to bcnu in air also resulted in increased toxicity. related effects were observed when cho cells were exposed to serum from mice that had receive ...20167107395
the use of drugs which deplete intracellular glutathione in hypoxic cell radiosensitization.diethylmaleate (dem) is a thiol-binding reagent with specificity toward glutathione. treatment of chinese hamster ovary (cho) cells in vitro with 2 x 10(-4) m dem for one hour results in a decrease in glutathione content to less than 5% of control, without cytotoxicity. this treatment results in dose-modifying sensitization to radiation under hypoxic conditions, with no effect on the shoulder of the radiation survival curve. no effect on the radiation sensitivity of oxygenated cells was seen. de ...20167107365
[changes in the cell membranes of chinese hamster fibroblasts during laser and x-ray irradiation studied with the use of a fluorescent probe]. 20167089203
host cell protein dynamics in recombinant cho cells: impacts from harvest to purification and beyond.during the production of recombinant protein products, such as monoclonal antibodies, manufacturers must demonstrate clearance of host cell impurities and contaminants to appropriate levels prior to use in the clinic. these include host cell dna and rna, product related contaminants such as aggregates, and importantly host cell proteins (hcps). despite the importance of hcp removal, the identity and dynamics of these proteins during cell culture and downstream processing (dsp) are largely unknow ...201723328085
identification of a single base-pair mutation of taa (stop codon) → gaa (glu) that causes light chain extension in a cho cell derived igg1.we describe here the identification of a stop codon taa (stop) → gaa (glu) = stop221e mutation on the light chain of a recombinant igg1 antibody expressed in a chinese hamster ovary (cho) cell line. the extended light chain variants, which were caused by translation beyond the mutated stop codon to the next alternative in-frame stop codon, were observed by mass spectra analysis. the abnormal peptide peaks present in tryptic and chymotryptic lc-ms peptide mapping were confirmed by n-terminal sequ ...201723018810
characterization of monoclonal antibody size variants containing extra light chains.size exclusion chromatography (sec) is the most commonly used method to separate and quantify monoclonal antibody (mab) size variants. mab-a is an igg1 subtype humanized monoclonal antibody recombinantly produced in chinese hamster ovary (cho) cells. sec analysis of mab-a resolved a peak, named peak 1, which elutes between monomer and dimer peaks. mab-a lots produced from different clones and production scales all have 0.2-0.3% of sec peak 1. electron spray ionization--time of flight mass spectr ...201723255003
very high density of chinese hamster ovary cells in perfusion by alternating tangential flow or tangential flow filtration in wave bioreactor™-part ii: applications for antibody production and cryopreservation.a high cell density perfusion process of monoclonal antibody (mab) producing chinese hamster ovary (cho) cells was developed in disposable wave bioreactor™ using external hollow fiber (hf) filter as cell separation device. tangential flow filtration (tff) and alternating tangential flow (atf) systems were compared and process applications of high cell density perfusion were studied here: mab production and cryopreservation. operations by perfusion using microfiltration (mf) or ultrafiltration (u ...201723436783
a high-throughput media design approach for high performance mammalian fed-batch cultures.an innovative high-throughput medium development method based on media blending was successfully used to improve the performance of a chinese hamster ovary fed-batch medium in shaking 96-deepwell plates. starting from a proprietary chemically-defined medium, 16 formulations testing 43 of 47 components at 3 different levels were designed. media blending was performed following a custom-made mixture design of experiments considering binary blends, resulting in 376 different blends that were tested ...201723563583
evidence for a mammalian late-g1 phase inhibitor of replication licensing distinct from geminin or cdk activity.pre-replication complexes (pre-rcs) are assembled onto dna during late mitosis and g1 to license replication origins for use in s phase. in order to prevent re-replication of dna, licensing must be completely shutdown prior to entry into s phase. while mechanisms preventing re-replication during s phase and mitosis have been elucidated, the means by which cells first prevent licensing during late g1 phase are poorly understood. we have employed a hybrid mammalian / xenopus egg extract replicatio ...201721983086
genomics in mammalian cell culture bioprocessing.explicitly identifying the genome of a host organism including sequencing, mapping, and annotating its genetic code has become a priority in the field of biotechnology with aims at improving the efficiency and understanding of cell culture bioprocessing. recombinant protein therapeutics, primarily produced in mammalian cells, constitute a $108 billion global market. the most common mammalian cell line used in biologic production processes is the chinese hamster ovary (cho) cell line, and althoug ...201722079893
toward a bioengineered heparin: challenges and strategies for metabolic engineering of mammalian cells.heparin is the most widely used pharmaceutical to control blood coagulation in modern medicine. a health crisis that took place in 2008 led to a demand for production of heparin from non-animal sources. since chinese hamster ovary (cho) cells are capable of producing heparan sulfate (hs), a related polysaccharide naturally, and heparin and hs share the same biosynthetic pathway, we hypothesized that heparin could be produced in cho cells by metabolic engineering. we developed stable human n-deac ...201722714556
expression and biological characterization of an anti-cd20 biosimilar candidate antibody: a case study.the cd20 molecule is a non-glycosylated protein expressed mainly on the surface of b lymphocytes. in some pathogenic b cells, it shows an increased expression, thus becoming an attractive target for diagnosis and therapy. rituximab is a chimeric antibody that specifically recognizes the human cd20 molecule. this antibody is indicated for the treatment of non-hodgkin lymphomas and autoimmune diseases, such as rheumatoid arthritis and systemic lupus erythematosus. in this work, we describe the sta ...201722647435
wavelength-dependent backscattering measurements for quantitative real-time monitoring of apoptosis in living cells.apoptosis--programmed cell death--is a cellular process exhibiting distinct biochemical and morphological changes. an understanding of the early morphological changes that a cell undergoes during apoptosis can provide the opportunity to monitor apoptosis in tissue, yielding diagnostic and prognostic information. there is avid interest regarding the involvement of apoptosis in cancer. the initial response of a tumor to successful cancer treatment is often massive apoptosis. current apoptosis dete ...201720059251
xgfr*, a novel affinity-matured bispecific antibody targeting igf-1r and egfr with combined signaling inhibition and enhanced immune activation for the treatment of pancreatic cancer.the epidermal growth factor receptor (egfr) and the insulin-like growth factor-1 receptor (igf-1r) play critical roles in tumor growth, providing a strong rationale for the combined inhibition of igf-1r and egfr signaling in cancer therapy. we describe the design, affinity maturation, in vitro and in vivo characterization of the bispecific anti-igf-1r/egfr antibody xgfr*. xgfr* is based on the bispecific igg antibody xgfr, which enabled heterodimerization of an igf-1r binding scfab heavy chain w ...201726984378
droplet size influences division of mammalian cell factories in droplet microfluidic cultivation.the potential of using droplet microfluidics for screening mammalian cell factories has been limited by the difficulty in achieving continuous cell division during cultivation in droplets. here, we report the influence of droplet size on mammalian cell division and viability during cultivation in droplets. chinese hamster ovary (cho) cells, the most widely used mammalian host cells for biopharmaceuticals production were encapsulated and cultivated in 33, 180 and 320 pl droplets for 3 days. perio ...201727535608
cell-free systems based on cho cell lysates: optimization strategies, synthesis of "difficult-to-express" proteins and future perspectives.nowadays, biotechnological processes play a pivotal role in target protein production. in this context, chinese hamster ovary (cho) cells are one of the most prominent cell lines for the expression of recombinant proteins and revealed as a safe host for nearly 40 years. nevertheless, the major bottleneck of common in vivo protein expression platforms becomes obvious when looking at the production of so called "difficult-to-express" proteins. this class of proteins comprises in particular several ...201727684475
a novel fusion protein, achr-fc, ameliorates myasthenia gravis by neutralizing antiacetylcholine receptor antibodies and suppressing acetylcholine receptor-reactive b cells.most patients with myasthenia gravis (mg) have elevated levels of autoantibodies against the nicotinic acetylcholine receptor (achr) at the neuromuscular junction, which leads to muscle weakness. we developed a fusion protein, achr-fc, as a novel therapeutic biomolecule for patients with mg and examined its efficacy. achr-fc was expressed by chinese hamster ovary cells and purified. we examined the neutralizing activity and cellular cytotoxicity of achr-fc using anti-achr antibody-producing hybr ...201727677608
cationic lipids bearing succinic-based, acyclic and macrocyclic hydrophobic domains: synthetic studies and in vitro gene transfer.in this communication we describe the construction of four succinic-based cationic lipids, their formulation with plasmid dna (pdna), and an evaluation of their in vitro gene delivery into chinese hamster ovarian (cho-k1) cells. the cationic lipids employed in this work possess either a dimethylamine or trimethylamine headgroup, and a macrocyclic or an acyclic hydrophobic domain composed of, or derived from two 16-atom, succinic-based acyl chains. the synthesized lipids and a co-lipid of neutral ...201727662033
fish-based analysis of clonally derived cho cell populations reveals high probability for transgene integration in a terminal region of chromosome 1 (1q13).a basic goal in the development of recombinant proteins is the generation of cell lines that express the desired protein stably over many generations. here, we constructed engineered chinese hamster ovary cell lines (cho-s) with a pcho-hvr1 vector that carried an extracellular domain of a vegf receptor (vr) fusion gene. forty-five clones with high hvr1 expression were selected for karyotype analysis. using fluorescence in situ hybridization (fish) and g-banding, we found that pcho-hvr1 was integ ...201727684722
reduction of ammonia and lactate through the coupling of glutamine synthetase selection and downregulation of lactate dehydrogenase-a in cho cells.chinese hamster ovary (cho) cell cultivation for production of therapeutic proteins is accompanied by production of metabolic wastes, mostly ammonia and lactate. to reduce ammonia production, the glutamine synthetase (gs) system was used to develop therapeutic monoclonal antibody (mab)-producing cho cells (sm-0.025). additionally, the lactate dehydrogenase-a (ldh-a) was downregulated with shrna to reduce lactate production in sm-0.025. the resulting mab-producing cell lines (#2, #46, and #52) pr ...201727704181
fam60a plays a role for production stabilities of recombinant cho cell lines.recombinant cho (chinese hamster ovary) cell lines producing therapeutic proteins often lose their production capability during long-term cultivation. to ensure that cho production cell lines can be up-scaled to high-volume bioreactors, labor intensive stability studies of several months have to be performed to deselect clones that are losing productivity over time. the ability to predict whether clones will produce recombinant proteins at constant high levels, for example, through determination ...201727617904
mar-mediated transgene integration into permissive chromatin and increased expression by recombination pathway engineering.untargeted plasmid integration into mammalian cell genomes remains a poorly understood and inefficient process. the formation of plasmid concatemers and their genomic integration has been ascribed either to non-homologous end-joining (nhej) or homologous recombination (hr) dna repair pathways. however, a direct involvement of these pathways has remained unclear. here, we show that the silencing of many hr factors enhanced plasmid concatemer formation and stable expression of the gene of interest ...201727575535
fx knockout cho hosts can express desired ratios of fucosylated or afucosylated antibodies with high titers and comparable product quality.during antibody dependent cell cytotoxicity (adcc) the target cells are killed by monocytes and natural killer cells. adcc is enhanced when the antibody heavy chain's core n-linked glycan lacks the fucose molecule(s). several strategies have been utilized to generate fully afucosylated antibodies. a commonly used and efficient approach has been knocking out the fut8 gene of the chinese hamster ovary (cho) host cells, which results in expression of antibody molecules with fully afucosylated glyca ...201727666939
specific immune response to phospholipase b-like 2 protein, a host cell impurity in lebrikizumab clinical material.host cell proteins are manufacturing process-related impurities that may co-purify with the product despite extensive efforts to optimize the purification process. the risks associated with these impurities can vary and may be patient and/or therapeutic dependent. therefore, it is critical to monitor and control the levels of these impurities in products and their potential impact on safety and efficacy. lebrikizumab is a humanized immunoglobulin g4 monoclonal antibody (mab) that binds specifica ...201727739010
'click'-xylosides as initiators of the biosynthesis of glycosaminoglycans: comparison of mono-xylosides with xylobiosides.different mono-xylosides and their corresponding xylobiosides obtained by a chemo-enzymatic approach featuring various substituents attached to a triazole ring were probed as priming agents for glycosaminoglycan (gag) biosynthesis in the xylosyltransferase-deficient pgsa-745 chinese hamster ovary cell line. xylosides containing a hydrophobic aglycone moiety were the most efficient priming agents. mono-xylosides induced higher gag biosynthesis in comparison with their corresponding xylobiosides. ...201727618481
lengthening of high-yield production levels of monoclonal antibody-producing chinese hamster ovary cells by downregulation of breast cancer 1.the establishment process of high-producing chinese hamster ovary (cho) cells for therapeutic protein production is usually laborious and time consuming because of the low probability of obtaining stable, high-producing clones over a long term. thus, development of an efficient approach is required to establish stable, high-producing cells. this study presents a novel method that can efficiently establish sustainably high-producing cell lines by acceleration of transgene amplification and suppre ...201727742176
the effect of particle size on the genotoxicity of gold nanoparticles.despite the increasing biomedical applications of gold nanoparticles (aunps), their toxicological effects need to be thoroughly understood. in the present study, the genotoxic potential of commercially available aunps with varying size (5, 20, and 50 nm) were assessed using a battery of in vitro and in vivo genotoxicity assays. in the comet assay, 20 and 50 nm aunps did not induce obvious dna damage in hepg2 cells at the tested concentrations, whereas 5 nm nps induced a dose-dependent increment ...201727770565
production and purification of recombinant glucocerebrosidase in transgenic rice cell suspension cultures.gaucher disease, which is caused by deficiency of glucocerebrosidase (gcd), is currently treated by enzyme replacement therapy. plant-based systems produce glycoproteins and can be combined with targeting strategies to generate proteins with terminal mannose structures for macrophage uptake. however, the gliding step for the purification is essential since the produced protein still exists inside cells. in the case of rice-amylase 1a (ramy1a) secretion signal peptide, gcd protein is secreted out ...201727817046
charge heterogeneity: basic antibody charge variants with increased binding to fc receptors.we identified active isoforms of the chimeric anti-gd2 antibody, ch14.18, a recombinant antibody produced in chinese hamster ovary cells, which is already used in clinical trials. (1,2,3) we separated the antibody by high resolution ion-exchange chromatography with linear ph gradient elution into acidic, main and basic charge variants on a preparative scale yielding enough material for an in-depth study of the sources and the effects of microheterogeneity. the binding affinity of the charge vari ...201727559765
reduced cytotoxicity in pcb-exposed chinese hamster ovary (cho) cells pretreated with vitamin e.the aim of this study was to evaluate protective effects of vitamin e (50 -150 μm) in ovary cells upon cytotoxic effects induced by two structurally distinct pcb congeners - planar "dioxin-like" pcb 77 and non-planar di-ortho-substituted pcb 153 with an emphasis on identifying differences in the mechanism of vitamin e action depending on the structure of congeners. application of three bioassays confirmed that pcbs decrease ovarian cell proliferation with slightly profound effects of pcb 77. pcb ...201727865896
detection of a phosphorylated glycine-serine linker in an igg-based fusion protein.molecular mass determination by electrospray ionization mass spectrometry of a recombinant igg-based fusion protein (mab1-f) produced in human embryonic kidney (hek) cells demonstrated the presence of a dominant +79 da product variant. using lc-ms tryptic peptide mapping analysis and collision-induced dissociation (cid) and electron-transfer/higher-energy collision dissociation fragmentations, the modification was localized to the c-terminal serine residue of a glycine-serine linker [(g4s)2] of ...201727661266
identification of an igg cdr sequence contributing to co-purification of the host cell protease cathepsin d.recombinant therapeutic monoclonal antibodies (mabs) must be purified from host cell proteins (hcps), dna, and other impurities present in chinese hamster ovary (cho) cell culture media. hcps can potentially result in adverse clinical responses in patients and, in specific cases, have caused degradation of the final mab product. as reported previously, residual traces of cathepsin d caused particle formation in the final product of mab-1. the current work was focused on identification of a prima ...201727798957
differences in trpc3 and trpc6 channels assembly in mesenteric vascular smooth muscle cells in essential hypertension.canonical transient receptor potential (trpc)3 and trpc6 channels of vascular smooth muscle cells (vsmcs) mediate stretch- or agonist-induced cationic fluxes, contributing to membrane potential and vascular tone. native trpc3/c6 channels can form homo- or heterotetrameric complexes, which can hinder individual trpc channel properties. the possibility that the differences in their association pattern may change their contribution to vascular tone in hypertension is unexplored. functional characte ...201727861908
establishment of monoclonal antibodies against cell surface domains of asct2/slc1a5 and their inhibition of glutamine-dependent tumor cell growth.human alanine-serine-cysteine transporter 2 (asct2; slc1a5) is a major transporter of the amino acid glutamine that is known to be overexpressed in certain malignant tumors. in this study, we generated specific monoclonal antibodies (mabs) against asct2 by establishing an asct2-expressing chinese hamster ovary cell line that was used to immunize mice and rats. the mabs km4008, km4012, and km4018 against asct2 were isolated through a cell-based screen; these specifically bound to asct2-positive c ...201727865832
lysosomal alpha-mannosidase and alpha-mannosidosis.lysosomal alpha-mannosidase with acidic ph optimum is ubiquitous in human tissues where is expressed in two major forms, a and b that are the product of a single gene located on chromosome 19. mutations in the gene encoding for alpha-mannosidase cause alpha- mannosidosis, an autosomal recessive disease, resulting in the accumulation of unprocessed mannose containing oligosaccharide material. this rare disease has an estimated incidence of 1/500.0.00 live births and clinically is divided into thr ...201727814608
selection of chemically defined media for cho cell fed-batch culture processes.two cho cell clones derived from the same parental cho(bc®) cell line and producing the same monoclonal antibody (bc-g, a low producing clone; bc-p, a high producing clone) were tested in four basal media in all possible combinations with three feeds (=12 conditions) in fed-batch cultures. higher amino acid feeding did not always lead to higher mab production. the two clones showed differences in cell physiology, metabolism and optimal medium-feed combinations. during the phase transitions of al ...201727900626
a framework to quantify karyotype variation associated with cho cell line instability at a single-cell level.chinese hamster ovary (cho) cells, the major mammalian host cells for biomanufacturing of therapeutic proteins, have been extensively investigated to enhance productivity and product quality. however, cell line instability resulting in unexpected changes in productivity or product quality continues to be a challenge. based on previous reports about causes and characteristics of production instability, we hypothesized that chromosomal rearrangements due to genomic instability are associated with ...201727922175
segmented linear modeling of cho fed-batch culture and its application to large scale production.we describe a systematic approach to model cho metabolism during biopharmaceutical production across a wide range of cell culture conditions. to this end, we applied the metabolic steady state concept. we analyzed and modeled the production rates of metabolites as a function of the specific growth rate. first, the total number of metabolic steady state phases and the location of the breakpoints were determined by recursive partitioning. for this, the smoothed derivative of the metabolic rates wi ...201727869296
improved recombinant antibody production by cho cells using a production enhancer dna element with repeated transgene integration at a predetermined chromosomal site.chinese hamster ovary (cho) cells are one of the most useful host cell lines for the production of biopharmaceutical proteins. although a series of production processes have been refined to improve protein productivity and cost performance, establishing producer cells is still time-consuming and labor-intensive. recombinase-mediated site-specific gene integration into a predetermined chromosomal locus may enable predictable protein expression, reducing the laborious process of cell screening. we ...201727856232
rational design of glycoengineered interferon-β analogs with improved aggregation state: experimental validation.recombinant human interferon-β (rhifn-β) used clinically has lower efficacy than expected due to protein instabilities such as aggregation. increasing molecular stability, glycoengineering has been used to improve clinical efficacy for a number of therapeutics; however, often labor-intensive trail-and-error approaches are used to identify additional glycosylation sites. in this study two rhifn-β analogs with one additional glycosylation site, l6t and s75n, identified by a rational in silico appr ...201727881683
flavonoids from heliotropium subulatum exudate and their evaluation for antioxidant, antineoplastic and cytotoxic activities ii.the flavonoids are the largest group of phenolic compounds isolated from a wide range of higher plants. these compounds work as antimicrobials, anti-insect agents and protect plants from other types of biotic and abiotic stresses. various researchers have suggested that flavonoids possessed antioxidant, antineoplastic and cytotoxic activities. the main objective of this study was to test dichloromethane fraction of resinous exudate of heliotropium subulatum for their antioxidant, antineoplastic ...201727905025
knockout of a difficult-to-remove cho host cell protein, lipoprotein lipase, for improved polysorbate stability in monoclonal antibody formulations.while the majority of host cell protein (hcp) impurities are effectively removed in typical downstream purification processes, a small population of hcps are particularly challenging. previous studies have identified hcps that are challenging for a variety of reasons. lipoprotein lipase (lpl)-a chinese hamster ovary (cho) hcp that functions to hydrolyze esters in triglycerides-was one of ten hcps identified in previous studies as being susceptible to retention in downstream processing. lpl may d ...201727943242
application of (13)c flux analysis to identify high-productivity cho metabolic phenotypes.industrial bioprocesses place high demands on the energy metabolism of host cells to meet biosynthetic requirements for maximal protein expression. identifying metabolic phenotypes that promote high expression is therefore a major goal of the biotech industry. we conducted a series of (13)c flux analysis studies to examine the metabolic response to igg expression during early stationary phase of cho cell cultures grown in 3l fed-batch bioreactors. we examined eight clones expressing four differe ...201728122259
a novel approach to characterize host cell proteins associated with therapeutic monoclonal antibodies.recombinant monoclonal antibody (mab) products are widely produced in the pharmaceutical industry using chinese hamster ovary (cho) cells. host cell proteins (hcps) are one of many process-related impurities generated during the production of mab products. the multi-analyte hcp enzyme linked immunosorbent assay (elisa) is the industry standard accepted assay to measure clearance of hcps from recombinant protein therapeutics. while similar platform processes are used for expression and purificati ...201728112396
expression and characterization of a codon-optimized blood coagulation factor viii.essentials recombinant factor viii (fviii) is known to be expressed at a low level in cell culture. to increase expression, we used codon-optimization of a b-domain deleted fviii (bdd-fviii). this resulted in 7-fold increase of the expression level in cell culture. the biochemical properties of codon-optimized bdd-fviii were similar to the wild-type protein.201728109042
from the cover: an investigation of the genotoxicity and interference of gold nanoparticles in commonly used in vitro mutagenicity and genotoxicity assays.the suitability of 4 in vitro assays, commonly used for mutagenicity and genotoxicity assessment, was investigated in relation to treatment with 14 nm citrate-stabilized gold nanoparticles (aunps). specifically, the ames test was conducted without metabolic activation, where no mutagenic effects were observed. high resolution transmission electron microscopy and cytoviva dark-field image analysis showed that aunps did not enter the bacterial cells, thus confirming the unreliability of the ames t ...201728108664
a highly efficient modified human serum albumin signal peptide to secrete proteins in cells derived from different mammalian species.signal peptides (sps) are key elements in the production of recombinant proteins; however, little information is available concerning different sp in mammalian cells other than cho. in order to study the efficiency of different sps to direct the traffic along the secretory pathway of the green fluorescence protein (gfp) and a scfv-fc fusion protein; cho-k1, hek293 and ns0 cell lines were transfected in a transient and stable way. sp of human azurocidin (az), modified human albumin (msa), modifie ...201728089882
production of human mutant biologically active hepatocyte growth factor in chinese hamster ovary cells.hepatocyte growth factor (hgf) is a potent multifunctional cytokine that affects proliferation, migration, and morphogenesis of various cells. hgf is secreted as an inactive single-chain precursor protein and activated by the cleavage of serine proteases to form heterodimers. in our current study, the cleavage site of hgf was blocked by replaced arg 494 of glu (r494e) that resulted in the single-chain hgf (r494e) unable to be cleaved by serine proteases. we established chinese hamster ovary (cho ...201728045643
asp73-dependent and -independent regulation of the affinity of ligands for human histamine h1 receptors by na().the affinity of ligands for g-protein-coupled receptors (gpcrs) is allosterically regulated by na(+) via a highly conserved aspartate residue (asp(2.50)) in the second transmembrane domain of gpcrs. in the present study, we examined the na(+)-mediated regulation of the affinity of ligands for gq/11-protein-coupled human histamine h1 receptors in chinese hamster ovary cells. the affinities of 3 agonists and 20 antihistamines were evaluated by their displacement curves against the binding of [(3)h ...201728040476
the fibrous form of intracellular inclusion bodies in recombinant variant fibrinogen-producing cells is specific to the hepatic fibrinogen storage disease-inducible variant fibrinogen.fibrinogen storage disease (fsd) is a rare disorder that is characterized by the accumulation of fibrinogen in hepatocytes and induces liver injury. six mutations in the γc domain (γg284r, γt314p, γd316n, the deletion of γg346-q350, γg366s, and γr375w) have been identified for fsd. our group previously established γ375w fibrinogen-producing chinese hamster ovary (cho) cells and observed aberrant large granular and fibrous forms of intracellular inclusion bodies. the aim of this study was to inve ...201728161763
a vector based on the chicken hypersensitive site 4 insulator element replicates episomally in mammalian cell.gene therapy in mammalian cells requires vectors exhibiting long-term stability and high expression. episomal gene expression vectors offer a safe and attractive alternative to those that integrate into the host cell genome. in the present study, we developed a new episomal vector based on the insulator, chicken hypersensitive site 4 (chs4). the chs4 element was artificially synthesized, cloned into the pegfp-c1 vector, and used to transfect chinese hamster ovary (cho) and human chang liver cell ...201728155604
design, synthesis and biological activity of novel molecules designed to target parp and dna.in order to enhance the cytotoxic potential of poly(adp-ribose) polymerase (parp) inhibitors in brca1 or 2 deficient tumours, we designed a series of molecules containing a 1,2,3-triazene moiety tethered to a parp targeting scaffold. a cell-based selectivity assay involving a brca2-deficient chinese hamster cell line and its corresponding brca2 wild type transfectant, was used to predict the parp targeting potential of the latter agents. the results showed that adding a dna damaging function to ...201728003142
method comparison for analyzing wound healing rates.wound healing scratch assay is a frequently used method to characterize cell migration, which is an important biological process in the course of development, tissue repair, and immune response for example. the measurement of wound healing rate, however, varies among different studies. here we summarized these measurements into three types: i) direct rate average; ii) regression rate average; and iii) average distance regression rate. using chinese hamster ovary (cho) cells as a model, we compar ...201728177756
the vdac2-bak axis regulates peroxisomal membrane permeability.peroxisomal biogenesis disorders (pbds) are fatal genetic diseases consisting of 14 complementation groups (cgs). we previously isolated a peroxisome-deficient chinese hamster ovary cell mutant, zp114, which belongs to none of these cgs. using a functional screening strategy, vdac2 was identified as rescuing the peroxisomal deficiency of zp114 where vdac2 expression was not detected. interestingly, knockdown of bak or overexpression of the bak inhibitors bcl-xl and mcl-1 restored peroxisomal bio ...201728174205
small-molecule-biased formyl peptide receptor agonist compound 17b protects against myocardial ischaemia-reperfusion injury in mice.effective treatment for managing myocardial infarction (mi) remains an urgent, unmet clinical need. formyl peptide receptors (fpr) regulate inflammation, a major contributing mechanism to cardiac injury following mi. here we demonstrate that fpr1/fpr2-biased agonism may represent a novel therapeutic strategy for the treatment of mi. the small-molecule fpr1/fpr2 agonist, compound 17b (cmpd17b), exhibits a distinct signalling fingerprint to the conventional fpr1/fpr2 agonist, compound-43 (cmpd43). ...201728169296
hiv protease inhibitors alter amyloid precursor protein processing via β-site amyloid precursor protein cleaving enzyme-1 translational up-regulation.mounting evidence implicates antiretroviral (arv) drugs as potential contributors to the persistence and evolution of clinical and pathological presentation of hiv-associated neurocognitive disorders in the post-arv era. based on their ability to induce endoplasmic reticulum (er) stress in various cell types, we hypothesized that arv-mediated er stress in the central nervous system resulted in chronic dysregulation of the unfolded protein response and altered amyloid precursor protein (app) proc ...201727993242
high-throughput lipidomic and transcriptomic analysis to compare sp2/0, cho, and hek-293 mammalian cell lines.a combined lipidomics and transcriptomics analysis was performed on mouse myeloma sp2/0, chinese hamster ovary (cho), and human embryonic kidney (hek) cells in order to compare widely used mammalian expression systems. initial thin layer chromatography (tlc) analysis indicated that phosphatidylethanolamine (pe) and phosphatidylcholine (pc) were the major lipid components in all cell lines with lower amounts of sphingomyelin (sm) in sp2/0 compared to cho and hek, which was subsequently confirmed ...201727991764
assessing the genotoxicity of two commonly occurring byproducts of water disinfection: chloral hydrate and bromal hydrate.water disinfection treatments result in the formation of disinfection byproducts (dbps) that have been linked to adverse human health outcomes including higher incidence of bladder and colorectal cancer. however, data about the genotoxicity of dbps is limited to only a small fraction of compounds. chloral hydrate (ch) and bromal hydrate (bh) are two trihaloacetaldehydes commonly detected in disinfected waters, but little is known about their genotoxicity, especially bh. we investigated the genot ...201728010927
role of a tachykinin-related peptide and its receptor in modulating the olfactory sensitivity in the oriental fruit fly, bactrocera dorsalis (hendel).insect tachykinin-related peptide (trp), an ortholog of tachykinin in vertebrates, has been linked with regulation of diverse physiological processes, such as olfactory perception, locomotion, aggression, lipid metabolism and myotropic activity. in this study, we investigated the function of trp (bdtrp) and its receptor (bdtrpr) in an important agricultural pest, the oriental fruit fly bactrocera dorsalis. bdtrpr is a typical g-protein coupled-receptor (gpcr), and it could be activated by the pu ...201727923683
integrin αiibβ3-dependent erk signaling is regulated by src and rho kinases in both leu33 and pro33 polymorphic isoforms.platelet integrin αiibβ3 possesses a leu/pro polymorphism at residue 33 (leu33/hpa-1a or pro33/hpa-1b). the pro33 isoform has been suggested to exhibit prothrombotic features. αiibβ3-expressing cho (chinese hamster ovary) cells on immobilized fibrinogen show activation of the map kinase family member erk2, with an enhanced erk2 activity in pro33 cells compared to leu33 cells. in our present work, we examined how the leu/pro polymorphism modulates the erk2 activation stimulated by 2 differently t ...201727923225
improving the secretory capacity of chinese hamster ovary cells by ectopic expression of effector genes: lessons learned and future directions.chinese hamster ovary (cho) cells are the preferred cell factory for the production of therapeutic glycoproteins. although efforts primarily within bioprocess optimization have led to increased product titers of recombinant proteins (r-proteins) expressed in cho cells, post-transcriptional bottlenecks in the biosynthetic pathway of r-proteins remain to be solved. to this end, the ectopic expression of transgenes (effector genes) offers great engineering potential. however, studies on effector ge ...201727931938
integrated genome and protein editing swaps α-2,6 sialylation for α-2,3 sialic acid on recombinant antibodies from cho.immunoglobin g with α-2,6 sialylation has been reported to have an impact on antibody-dependent cellular cytotoxicity and anti-inflammatory efficacy. however, production of antibodies with α-2,6 sialylation from chinese hamster ovary cells is challenging due to the inaccessibility of sialyltransferases for the heavy chain n-glycan site and the presence of exclusively α-2,3 sialyltransferases. in this study, combining mutations on the fc regions to allow sialyltransferase accessibility with overe ...201727943633
mir-143 targets mapk7 in cho cells and induces a hyperproductive phenotype to enhance production of difficult-to-express proteins.in recent years, the number of complex but clinically effective biologicals such as multi-specific antibody formats and fusion proteins has increased dramatically. however, compared to classical monoclonal antibodies (mabs), these rather artificially designed therapeutic proteins have never undergone millions of years of evolution and thus often turn out to be difficult-to-express using mammalian expression systems such as chinese hamster ovary (cho) cells. in order to provide access to these so ...201728371547
a semi-empirical glycosylation model of a camelid monoclonal antibody under hypothermia cell culture conditions.the impact of cell culture environment on the glycan distribution of a monoclonal antibody (mab) has been investigated through a combination of experiments and modeling. a newly developed cho duxb cell line was cultivated at two levels of initial glutamine (gln) concentrations (0, 4 mm) and incubation temperatures of (33 and 37 °c) in batch operation mode. hypothermia was applied either through the entire culture duration or only during the post-exponential phase. beyond reducing cell growth and ...201728285402
tbx20 controls the expression of the kcnh2 gene and of herg channels.long qt syndrome (lqts) exhibits great phenotype variability among family members carrying the same mutation, which can be partially attributed to genetic factors. we functionally analyzed the kcnh2 (encoding for kv11.1 or herg channels) and tbx20 (encoding for the transcription factor tbx20) variants found by next-generation sequencing in two siblings with lqts in a spanish family of african ancestry. affected relatives harbor a heterozygous mutation in kcnh2 that encodes for p.t152hfsx180 kv11 ...201728049825
high-resolution imaging and quantification of plasma membrane cholesterol by nanosims.cholesterol is a crucial lipid within the plasma membrane of mammalian cells. recent biochemical studies showed that one pool of cholesterol in the plasma membrane is "accessible" to binding by a modified version of the cytolysin perfringolysin o (pfo*), whereas another pool is sequestered by sphingomyelin and cannot be bound by pfo* unless the sphingomyelin is destroyed with sphingomyelinase (smase). thus far, it has been unclear whether pfo* and related cholesterol-binding proteins bind unifor ...201728167768
atf6β-based fine-tuning of the unfolded protein response enhances therapeutic antibody productivity of chinese hamster ovary cells.the dynamics of protein folding and secretion are key issues in improving the productivity and robustness of chinese hamster ovary (cho) producer cells. high recombinant protein secretion in cho producer clones triggers the activation of the unfolded protein response (upr), an intracellular response to the accumulation of unfolded and misfolded proteins in the endoplasmic reticulum (er). we previously reported that the human microrna (mirna) mir-1287 enhances productivity in igg expressing cho c ...201728165157
dedicated high dose rate (192)ir brachytherapy radiation fields for in vitro cell exposures at variable source-target cell distances: killing of mammalian cells depends on temporal dose rate fluctuation.afterloading brachytherapy is conducted by the stepwise movement of a radioactive source through surgically implanted applicator tubes where at predefined dwell positions calculated dwell times optimize spatial dose delivery with respect to a planned dose level. the temporal exposure pattern exhibits drastic fluctuations in dose rate at a given coordinate and within a single treatment session because of the discontinuous and repeated source movement into the target volume. this could potentially ...201728145285
engineering the expression of an anti-interleukin-13 antibody through rational design and mutagenesis.high levels of protein expression are key to the successful development and manufacture of a therapeutic antibody. here, we describe two related antibodies, ab001 and ab008, where ab001 shows a markedly lower level of expression relative to ab008 when stably expressed in chinese hamster ovary cells. we use single-gene expression vectors and structural analysis to show that the reduced titer is associated with the vl cdr2 of ab001. we adopted two approaches to improve the expression of ab001. fir ...201728130326
preferential hydrolysis of truncated oxidized glycerophospholipids by lysosomal phospholipase a2.truncated oxidized glycerophospholipids (ox-pls) are bioactive lipids resulting from oxidative stress. the catabolic pathways for truncated ox-pls are not fully understood. lysosomal phospholipase a2 (lpla2) with phospholipase a and transacylase activities is a key enzyme in phospholipid homeostasis. the present study assessed whether lpla2 could hydrolyze truncated ox-pls. incubation of lpla2 with liposomes consisting of 1,2-o-octadecenyl-sn-glycero-3-phosphocholine (dodpc)/1,2-dioleoyl-sn-glyc ...201727993948
development of a cho-based cell-free platform for synthesis of active monoclonal antibodies.chinese hamster ovary (cho) cells are routinely optimized to stably express monoclonal antibodies (mabs) at high titers. at the early stages of lead isolation and optimization, hundreds of sequences for the target protein of interest are screened. typically, cell-based transient expression technology platforms are used for expression screening, but these can be time- and resource-intensive. here, we have developed a cell-free protein synthesis (cfps) platform utilizing a commercially available c ...201728350472
production of recombinant human acid α-glucosidase with high-mannose glycans in gnt1 rice for the treatment of pompe disease.lysosomal storage diseases are a group of inherited metabolic disorders. patients are treated with enzyme replacement therapy (ert), in which the replacement enzymes are required to carry terminal mannose or mannose 6-phosphate residues to allow efficient uptake into target cells and tissues. n-acetylglucosaminyltransferase-i (gnti) mediates n-glycosylation in the cis cisternae of the golgi apparatus by adding n-acetylglucosamine to the exposed terminal mannose residue of core n-glycan structure ...201728363873
inhibition of glycosylation on a camelid antibody uniquely affects its fcγri binding activity.glycoengineering of mabs has become common practice in attempts to generate the ideal mab candidate for a wide range of therapeutic applications. the effects of these glycan modifications on the binding affinity of igg mabs for fcγriiia and their cytotoxicity are well known. however, little is understood about the effect that these modifications have on binding to the high affinity fcγri receptor. this study analyzed the effect of variable n-glycosylation on a human-llama hybrid mab (eg2-hfc, 80 ...201727721040
a strategy to accelerate protein production from a pool of clones in chinese hamster ovary cells for toxicology studies.in the biopharmaceutical industry, a clonally derived cell line is typically used to generate material for investigational new drug (ind)-enabling toxicology studies. the same cell line is then used to generate material for clinical studies. if a pool of clones can be used to produce material for ind-enabling toxicology studies ("pool for tox strategy") during the time a lead clone is being selected for clinical material production, the toxicology studies can be accelerated significantly (approx ...201728371489
an aromatic amino acid within intracellular loop 2 of the prostaglandin ep2 receptor is a prerequisite for selective association and activation of gαs.we previously demonstrated that the aromatic moiety of tyr(143) within the intracellular loop 2 (icl2) region of the prostaglandin ep2 receptor plays a crucial role in gs coupling. here we investigated whether the icl2 of the ep2 receptor directly binds to gαs and whether an aromatic moiety affects this interaction. in chinese hamster ovary cells, mutations of tyr(143) reduced the ability of the ep2 receptor to interact with g proteins as demonstrated by gtpγs sensitivity, as well as the ability ...201728336329
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