interacts_with DB00181?
Effective dasatinib uptake may occur without human organic cation transporter 1 ( O15245 ) : implications for the treatment of imatinib-resistant chronic myeloid leukemia . We have previously shown that imatinib uptake into chronic myeloid leukemia ( CML ) cells is dependent on human organic cation transporter 1 ( O15245 ; O15245 ) , and that low O15245 expression is an important determinant of clinical outcome to imatinib treatment . We hypothesized that dasatinib might be transported differently than imatinib , possibly accounting for its favorable effects in imatinib-resistant patients . (14)C-dasatinib uptake was greater in KCL22-transfected cells with pcDNA3- O15245 plasmid ( high O15245 -expressing cells ) than in control cells ( P = .02 ) . However , hOCT inhibitors did not decrease dasatinib uptake into either control or primary cells , in contrast to their block on imatinib uptake . Dasa-tinib decreased the level of phosphorylated CrkL to 49.9 % in control and 40.3 % in high O15245 -expressing cells . Dasa-tinib efflux was investigated in confluent P08183 -transfected MDCKII cell monolayers . Both dasatinib and imatinib were transported from the basal to the apical layer , indicating that they were transported by P08183 , which was confirmed using the P08183 inhibitor PSC833 ( P = .001 and P < .001 , respectively ) . Compared with imatinib , dasatinib achieved superior intracellular levels and P11274 - P00519 suppression even in cells with low or blocked O15245 . Efflux of dasatinib and imatinib appear similar via P08183 . Dasatinib may therefore offer an advantage over imatinib in patients with low O15245 expression . Enhancement of cytotoxicity of natural product drugs against multidrug resistant variant cell lines of human head and neck squamous cell carcinoma and breast carcinoma by tesmilifene . N,N-diethyl-2-[4-(phenylmethyl)phenoxyl]ethanamine ( tesmilifene ) , a tamoxifen derivative with antihistamine activity , greatly enhanced the survival of doxorubicin-treated , advanced stage breast cancer patients in a phase III trial . However , the molecular basis of tesmilifene action is not firmly established . The effects of tesmilifene on activity of several anticancer drugs was investigated using human head and neck squamous cell carcinoma ( HNSCC ) and breast carcinoma cell lines as a model system . Multidrug resistant ( MDR ) variants of an HNSCC cell line , HN-5a/V15e , and a breast carcinoma cell line , MCF-7/V25a , both highly overexpressed mdr1 ( P08183 ) mRNA and the proteins P-glycoprotein and glutathione transferase-pi . Drug sensitivities were measured by a vital stain after 4 days of continuous exposure to anticancer drug in the absence and presence of tesmilifene at a concentration that alone had no antiproliferative effect . DB04905 had minimal effect on drug cytotoxicity against the parental cell lines . However , the same tesmilifene treatment enhanced cytotoxicity of docetaxel , paclitaxel , epirubicin , doxorubicin , and vinorelbine against both MDR cell lines by up to 50 % . Flow cytometric measurement of annexin V/propidium iodide staining demonstrated that tesmilifene increased the killing of HN-5a/V15e cells caused by docetaxel after 24 and 48h exposure . DB04905 increased accumulation of radiolabelled vincristine in HN-5a/V15e cells , over 4h , by up to 100 % . The results suggest that tesmilifene might be effective in the treatment of tumors that are resistant to natural product drugs . The mechanism of enhancement appears to be related to expression of an ABC pump-dependent , MDR phenotype . Alteration of intracellular histamine H2 receptor cycling precedes antagonist-induced upregulation . Long-term administration of a histamine H2 receptor ( P25021 ) antagonist ( inverse agonist ) induces upregulation of P25021 in parietal cells , which may be relevant to the rebound hypersecretion of gastric acid that occurs after withdrawal of treatment . The mechanisms underlying this effect are unknown . We hypothesized that the P25021 upregulation could be related to receptor trafficking and used P25021 -green fluorescent protein ( P25021 -GFP ) to test the hypothesis . Human P25021 -GFP was generated and functionally expressed in P29320 -293 cells . Binding of the P25021 antagonist [3H]tiotidine was performed to quantify P25021 expression , and P25021 -GFP was imaged in living cells by confocal and evanescent wave microscopy . The binding affinity of [3H]tiotidine was not significantly different between P25021 -GFP- and wild-type P25021 -expressing P29320 -293 cells , both of which had constitutive activity of adenylate cyclase . Visualization of P25021 -GFP revealed that the agonist-induced P25021 internalization and the antagonist-induced recycling of the internalized P25021 from the recycling endosome within 2 h . Long exposure to the antagonist increased GFP fluorescence in the plasma membrane and also induced upregulation of P25021 -GFP estimated by the binding assay , whereas long exposure to the agonist enhanced degradative trafficking of P25021 -GFP . We examined whether the upregulation reflected an increase in receptor synthesis . Treatment with antagonist did not augment P25021 mRNA , and subsequent inhibition of protein synthesis by cycloheximide had no effect on P25021 upregulation . These findings suggested that upon exposure to an antagonist ( inverse agonist ) , the equilibrium between receptor endocytosis and recycling is altered before P25021 upregulation , probably via suppressing P25021 degradation . P17931 -independent Down-regulation of Q9UBS5 due to Treatment with Korean Herbal Extract HAD-B Reduces Proliferation of Human Colon Cancer Cells . OBJECTIVES : Many efforts have shown multi-oncologic roles of galectin-3 for cell proliferation , angiogenesis , and apoptosis . However , the mechanisms by which galectin-3 is involved in cell proliferation are not yet fully understood , especially in human colon cancer cells . METHODS : To cluster genes showing positively or negatively correlated expression with galectin-3 , we employed human colon cancer cell lines , SNU-61 , SNU-81 , SNU-769B , SNU-C4 and SNU- P01031 in high-throughput gene expression profiling . Gene and protein expression levels were determined by using real-time quantitative polymerase chain reaction ( PCR ) and western blot analysis , respectively . The proliferation rate of human colon cancer cells was measured by using a 3- ( 4 , 5- dimethylthiazol-2-yl ) -2 , 5-diphenyltetrazolium bromide ( MTT ) assay . RESULTS : Expression of γ-aminobutyric acid B receptor 1 ( Q9UBS5 ) showed a positive correlation with galectin-3 at both the transcriptional and the translational levels . Downregulation of galectin-3 decreased not only Q9UBS5 expression but also the proliferation rate of human colon cancer cells . However , Korean herbal extract , HangAmDan-B ( HAD-B ) , decreased expression of Q9UBS5 without any expressional change of galectin-3 , and offset γ-aminobutyric acid ( GABA ) -enhanced human colon cancer cell proliferation . CONCLUSIONS : Our present study confirmed that Q9UBS5 expression was regulated by galectin-3 . HAD-B induced galectin-3-independent down-regulation of Q9UBS5 , which resulted in a decreased proliferation of human colon cancer cells . The therapeutic effect of HAD-B for the treatment of human colon cancer needs to be further validated . The P08908 -receptor agonist flibanserin reduces drug-induced dyskinesia in O75916 -deficient mice . Drug-induced dyskinesia is a major complication of dopamine replacement therapy in advanced Parkinson 's disease consisting of dystonia , chorea and athetosis . Agonists at P08908 -receptors attenuate levodopa-induced motor complications in non-human primates . Mice with increased dopamine D2 receptor ( P14416 ) signalling due to the lack of expression of the regulator of G-protein signalling 9 ( O75916 ) also develop dyskinesia following levodopa treatment . We investigated whether the P08908 -receptor agonist flibanserin compared with buspirone reduces motor abnormalities induced by levodopa or quinelorane , a selective dopamine D2-receptor agonist . Following dopamine depletion via reserpine , 40 mice ( 20 wild-type and 20 O75916 knock-out ) were treated with flibanserin or buspirone in combination with levodopa or quinelorane . Motor behaviour was analysed using open field analysis . O75916 knock-out mice displayed significantly more drug-induced dystonia ( p < 0.04 ; t test ) than wild type . In quinelorane-treated wild-type mice flibanserin as well as buspirone significantly reduced dystonia ( p < 0.05 ) . In O75916 knock-out animals again both reduced quinelorane-induced dystonia . However , flibanserin was significantly more effective ( p = 0.003 ) . Following reserpine pretreatment and administration of levodopa wild-type and Q99697 9 knock-out mice showed mild to moderate dystonia . Surprisingly , 10 mg/kg buspirone increased dystonia in both animal groups , whereas it was decreased by 10 mg/kg flibanserin . However , compared with levodopa alone only the increase of dystonia by buspirone was significant ( p < 0.04 ) . DB04908 showed promising antidyskinetic effects in a model of drug-induced dyskinesia . Our data underline the possible benefit of P08908 agonists in drug-induced dyskinesia . Novel MEK inhibitor trametinib and other retinoblastoma gene ( RB ) -reactivating agents enhance efficacy of 5-fluorouracil on human colon cancer cells . Chemotherapy for colorectal cancer has become more complicated and diversified with the appearance of molecular-targeting agents . DB00544 ( DB00544 ) has been a mainstay of chemotherapy for colorectal cancer , but it is still unknown whether the combining of DB00544 with novel molecular-targeting agents is effective . P04818 ( TS ) is a direct target of DB00544 , and the low TS level has been generally supposed to sensitize DB00544 's efficacy . We therefore hypothesized that RB-reactivating agents could enhance the efficacy of DB00544 , because the RB-reactivating agents could suppress the function of transcription factor E2F of TS gene promoter . We used three RB-reactivating agents , trametinib/GSK1120212 ( MEK inhibitor ) , fenofibrate ( Q07869 α agonist ) , and LY294002 ( PI3K inhibitor ) , with DB00544 against human colon cancer HT-29 and HCT15 cells . DB08911 induced p15 and p27 expression and reduced cyclin D1 levels in HT-29 cells . Fenofibrate also dephosphorlated P27361 /2 and reduced cyclin D1 levels in HT-29 cells . LY294002 induced p27 expression in HCT15 cells . All three agents caused dephosphorylation of RB protein and P55008 -phase arrest with a reduction of TS expression . As a consequence , the combination of DB00544 with each of the agents resulted in a significant decrease of colony numbers in HT-29 or HCT15 cells . These results suggest " RB-reactivation therapy " using molecular-targeting agents to be a new strategy for DB00544 -based chemotherapy . In particular , we strongly expect trametinib , which was discovered in Japan and was recently submitted to FDA for approval , to be used together with established regimens for colorectal cancer . Complex formation with the Type B gamma-aminobutyric acid receptor affects the expression and signal transduction of the extracellular calcium-sensing receptor . Studies with P29320 -293 cells and neurons . We co-immunoprecipitated the Ca(2+)-sensing receptor ( CaR ) and type B gamma-aminobutyric acid receptor ( GABA-B-R ) from human embryonic kidney ( P29320 ) -293 cells expressing these receptors and from brain lysates where both receptors are present . CaRs extensively co-localized with the two subunits of the GABA-B-R ( Q96GN5 and R2 ) in P29320 -293 cell membranes and intracellular organelles . Coexpressing CaRs and GABA-B-R1s in P29320 -293 cells suppressed the total cellular and cell surface expression of CaRs and inhibited phospholipase C activation in response to high extracellular [ Ca(2+) ] ( [Ca(2+)](e) ) . In contrast , coexpressing CaRs and GABA-B-R2s enhanced CaR expression and signaling responses to raising [Ca(2+)](e) . The latter effects of the O75899 on the CaR were blunted by coexpressing the Q9UBS5 . Coexpressing the CaR with Q9UBS5 or R2 enhanced the total cellular and cell surface expression of the Q9UBS5 or R2 , respectively . Studies with truncated CaRs indicated that the N-terminal extracellular domain of the CaR participated in the interaction of the CaR with the Q9UBS5 and R2 . In cultured mouse hippocampal neurons , CaRs co-localized with the Q9UBS5 and R2 . CaRs and GABA-B-R1s also co-immunoprecipitated from brain lysates . The expression of the CaR was increased in lysates from Q9UBS5 knock-out mouse brains and in cultured hippocampal neurons with their Q9UBS5 genes deleted in vitro . Thus , CaRs and GABA-B-R subunits can form heteromeric complexes in cells , and their interactions affect cell surface expression and signaling of CaR , which may contribute to extracellular Ca(2+)-dependent receptor activation in target tissues . Puerarin alleviates noise-induced hearing loss via affecting PKCγ and GABAB receptor expression . Noise-induced hearing loss ( NIHL ) often results from prolonged exposure to high levels of noise . Our previous study revealed that during the development of NIHL , the expression of protein kinase C γ subunit ( PKCγ ) and GABAB receptor ( GABABR ) was changed within the cochlear nuclear complex ( CNC ) , suggesting that these molecules might be the potential targets for the treatment of NIHL . As an extending study , here we focused on puerarin , a major isoflavonoid extracted from Pueraria lobota , which has been used in the treatment of cardiovascular and cerebrovascular diseases , and investigated whether it could protect against NIHL by acting on PKCγ and GABABR . Transgenic Q99259 -GFP knock-in mice were subjected to the NIHL model and their auditory functions were evaluated by the auditory brainstem response thresholds and distortion product oto-acoustic emission signals . Our results showed that 200mg/kg puerarin treatment ameliorated the thresholds of auditory brainstem response of NIHL mice significantly . Triple immunofluorescence staining and electron microscopy results revealed that GFP-positive neurons in the superficial layers of CNC expressed both PKCγ and Q9UBS5 , and Q99259 -positive terminals contacted PKCγ- or Q9UBS5 -positive neurons . Immunoblotting and RT-PCR results showed that NIHL increased the expression of PKCγ but decreased that of Q9UBS5 and O75899 at both protein and mRNA levels in the CNC . Puerarin significantly attenuated the increased expression of PKCγ but elevated the reduced expression of Q9UBS5 and O75899 after noise exposure . Thus , we provided the first evidence that puerarin ameliorated the auditory functions of NIHL mice , and this effect may be due to its ability to regulate the expression of PKCγ and GABABR . Type B gamma-aminobutyric acid receptors modulate the function of the extracellular Ca2+-sensing receptor and cell differentiation in murine growth plate chondrocytes . Extracellular calcium-sensing receptors ( CaRs ) and metabotropic or type B gamma-aminobutyric acid receptors ( GABA-B-Rs ) , two closely related members of family C of the G protein-coupled receptor superfamily , dimerize in the formation of signaling and membrane-anchored receptor complexes . We tested whether CaRs and two GABA-B-R subunits ( Q96GN5 and R2 ) are expressed in mouse growth plate chondrocytes ( GPCs ) by PCR and immunocytochemistry and whether interactions between these receptors influence the expression and function of the CaR and extracellular Ca(2+)-mediated cell differentiation . Both CaRs and the Q9UBS5 and -R2 were expressed in the same zones of the growth plate and extensively colocalized in intracellular compartments and on the membranes of cultured GPCs . The Q9UBS5 co-immunoprecipitated with the CaR , confirming a physical interaction between the two receptors in GPCs . In vitro knockout of Q9UBS5 genes , using a Cre-lox recombination strategy , blunted the ability of high extracellular Ca(2+) concentration to activate phospholipase C and P27361 /2 , suppressed cell proliferation , and enhanced apoptosis in cultured GPCs . In GPCs , in which the Q9UBS5 was acutely knocked down , there was reduced expression of early chondrocyte markers , aggrecan and type II collagen , and increased expression of the late differentiation markers , type X collagen and osteopontin . These results support the idea that physical interactions between CaRs and GABA-B-R1s modulate the growth and differentiation of GPCs , potentially by altering the function of CaRs . Modulatory effects of heparin and short-length oligosaccharides of heparin on the metastasis and growth of LMD MDA-MB 231 breast cancer cells in vivo . Expression of the chemokine receptor P61073 allows breast cancer cells to migrate towards specific metastatic target sites which constitutively express P48061 . In this study , we determined whether this interaction could be disrupted using short-chain length heparin oligosaccharides . Radioligand competition binding assays were performed using a range of heparin oligosaccharides to compete with polymeric heparin or heparan sulphate binding to I(125) P48061 . DB01109 dodecasaccharides were found to be the minimal chain length required to efficiently bind P48061 ( 71 % inhibition ; P < 0.001 ) . These oligosaccharides also significantly inhibited P48061 -induced migration of P61073 -expressing LMD MDA-MB 231 breast cancer cells . In addition , heparin dodecasaccharides were found to have less anticoagulant activity than either a smaller quantity of polymeric heparin or a similar amount of the low molecular weight heparin pharmaceutical product , DB06822 . When given subcutaneously in a SCID mouse model of human breast cancer , heparin dodecasaccharides had no effect on the number of lung metastases , but did however inhibit ( P < 0.05 ) tumour growth ( lesion area ) compared to control groups . In contrast , polymeric heparin significantly inhibited both the number ( P < 0.001 ) and area of metastases , suggesting a differing mechanism for the action of polymeric and heparin-derived oligosaccharides in the inhibition of tumour growth and metastases . [ Differential gene expression in nasopharyngeal carcinoma cell with reduced and normal expression of 6A8 alpha-mannosidase ] . OBJECTIVE : To detect the differential display of mRNA expression between human nasopharyngeal carcinoma cell CNE-2L2 with reduced malignancy caused by transduction of a DNA antisense to 6A8 alpha-mannosidase cDNA ( AS cell ) and the wild type cell ( W cell ) . METHODS : Differential display of mRNA expression was analyzed using DNA microarray analysis . The datasets were confirmed by Northern blotting and RT-PCR . RESULTS : Out of the 1069 genes analyzed , 34 genes were up-regulated in AS cells relative to W cells . Conversely , 42 genes were down-regulated . The genes , up-regulation of which might have suppressive effect on tumor malignant behaviors , were P130 mRNA for 130K protein , TGF-betaIIR alpha , Q9UBS5 , P36897 , Q13829 , STANIN , E-CADHERIN , P35221 and 2 , P48378 , TMPO , etc . The genes , down-regulation of which might have suppressive effect on tumor malignant behaviors , were P16070 , Q92597 , P01137 , P46782 , LEGUMAIIN , P35520 , P13987 , P09661 , etc . The microarray datasets were confirmed by Northern blot and RT-PCR analysis . CONCLUSIONS : In comparison to the W cell , AS cell has up-regulation of 34 genes and down-regulation of 42 genes . Changes of the gene expression may play a role in the malignancy reduction of AS cell . [ CT analysis of developmental spinal canal stenosis ] . The cervical and the lumbar vertebrae were analyzed by computed tomography in two groups . One group consisted of the patients with developmental stenosis of the cervical spine ( the smallest AP diameter of the cervical spinal canal in the lateral view of X-ray was less than 12 mm ) and the other was the control group ( the smallest diameter was more than 14 mm ) . The canal to body ratio of the cross-sectional area ( P16152 ) , the modified Jones ' " canal to body ratio " ( JNS ) and the flat index of the vertebral canal ( FI ) were measured at C4, P01031 , P13671 , Q9NP50 and Q15004 . The shape of the vertebral canal of Q9NP50 and Q15004 was classified according to Baddeley . In the stenosis group , P16152 and JNS were smaller at every measured level and FI was smaller at C4 , P01031 and P13671 compared with the control group , Trefoil canals were found more frequently in the stenosis group . There was a clear relationship between the developmental stenosis of the cervical and the lumbar spinal canal . On treating patients with cervical spondylotic myelopathy , one should consider the possibility of pan-spinal canal stenosis . Molecular identification of the human O75899 : cell surface expression and coupling to adenylyl cyclase in the absence of Q9UBS5 . We have identified a gene encoding a GABAB receptor , the human O75899 , located on chromosome 9q22.1 , that is distinct from the recently reported rat Q9UBS5 . O75899 structurally resembles Q9UBS5 ( 35 % identity ) , having seven transmembrane domains and a large extracellular region , but differs in having a longer carboxy-terminal tail . O75899 is localized to the cell surface in transfected COS cells , and negatively couples to adenylyl cyclase in response to GABA , baclofen , and 3-aminopropyl(methyl)phosphinic acid in CHO cells lacking Q9UBS5 . DB00181 action is inhibited by the GABABR antagonist , 2-hydroxysaclofen . The human O75899 and Q9UBS5 genes are differentially expressed in the nervous system , with the greatest difference being detected in the striatum in which Q9UBS5 but not O75899 mRNA transcripts are detected . O75899 and Q9UBS5 mRNAs are also coexpressed in various brain regions such as the Purkinje cell layer of the cerebellum . Identification of a functional homomeric O75899 coupled to adenylyl cyclase suggests that the complexity of GABAB pharmacological data is at least in part due to the presence of more than one receptor and opens avenues for future research leading to an understanding of metabotropic GABA receptor signal transduction mechanisms . Immunocytochemical localization of Q9UBS5 receptor subunits in the basolateral amygdala . Gamma-aminobutyric acid B ( GABAB ) receptors ( GBRs ) are G-protein-coupled receptors that mediate a slow , prolonged form of inhibition in the basolateral amygdala ( P00519 ) and other brain areas . Recent studies indicate that this receptor is a heterodimer consisting of Q9UBS5 ( GBR1 ) and O75899 subunits . In the present investigation , antibodies to the Q9UBS5 subunit were used to study the neuronal localization of GBRs in the rat P00519 . GBR immunoreactivity was mainly found in spine-sparse interneurons and astrocytes at the light microscopic level . Very few pyramidal neurons exhibited perikaryal staining . Dual-labeling immunofluorescence analysis indicated that each of the four main subpopulations of interneurons exhibited GBR immunoreactivity . Virtually 100 % of large CCK+ neurons in the basolateral and lateral nuclei were GBR+ . In the basolateral nucleus 72 % of somatostatin ( Q8TE85 ) , 73 % of parvalbumin ( PV ) and 25 % of P01282 positive interneurons were GBR+ . In the lateral nucleus 50 % of somatostatin , 30 % of parvalbumin and 27 % of P01282 positive interneurons were GBR+ . Electron microscopic ( EM ) analysis revealed that most of the light neuropil staining seen at the light microscopic level was due to the staining of dendritic shafts and spines , most of which probably belonged to spiny pyramidal cells . Very few axon terminals ( Ats ) were GBR+ . In summary , this investigation demonstrates that the distal dendrites of pyramidal cells , and varying percentages of each of the four main subpopulations of interneurons in the P00519 , express GBRs . Because previous studies suggest that GBR-mediated inhibition modulates DB01221 -dependent EPSPs in the P00519 , these receptors may play an important role in neuronal plasticity related to emotional learning . Role of histamine receptors in the effects of histamine on the production of reactive oxygen species by whole blood phagocytes . AIMS : The diverse physiological functions of histamine are mediated through distinct histamine receptors . In this study we investigated the role of P25021 and Q9H3N8 in the effects of histamine on the production of reactive oxygen species by phagocytes in whole blood . MAIN METHODS : Changes in reactive oxygen species ( ROS ) production by whole blood phagocytes after treatment with histamine , Q9H3N8 agonists ( 4-methylhistamine , VUF8430 ) , P25021 agonist ( dimaprit ) and their combinations with Q9H3N8 antagonist ( JNJ10191584 ) and P25021 antagonist ( ranitidine ) were determined using the chemiluminescence ( CL ) assay . To exclude the direct scavenging effects of the studied compounds on the CL response , the antioxidant properties of all compounds were measured using several methods ( TRAP , ORAC , and luminol-HRP-H2O2 based CL ) . KEY FINDINGS : DB11320 , 4-methylhistamine , VUF8430 and dimaprit inhibited the spontaneous and OZP-activated whole blood CL in a dose-dependent manner . On the other hand , only VUF8430 was able to inhibit PMA-activated whole blood CL . DB00863 , but not JNJ10191584 , completely reduced the effects of histamine , 4-methylhistamine and dimaprit . The direct scavenging ability of tested compounds was negligible . SIGNIFICANCE : Our results demonstrate that the inhibitory effects of histamine on ROS production in whole blood phagocytes were caused by P25021 . Our results also suggest that Q9H3N8 agonists in concentrations higher than 10(-6)M may also influence ROS production via binding to P25021 . DB09280 - DB08820 in Patients with Cystic Fibrosis Homozygous for Phe508del P13569 . Influence of immunomodulatory drugs on the cytotoxicity induced by monoclonal antibody 17-1A and interleukin-2 . Patients treated with monoclonal antibodies and cytokines for cancer receive often co-medication , which may influence treatment efficacy . Therefore , we investigated with a flowcytometric cytotoxicity assay the effect of several immunomodulatory drugs on antibody dependent cellular cytotoxicity ( ADCC ) , interleukin-2 ( P60568 ) induced cytotoxicity and P60568 -induced-ADCC . We found that dexamethasone markedly inhibited the P60568 induced cytotoxicity and the P60568 -induced-ADCC . DB00904 , a P46098 serotonin receptor antagonist augmented significantly ADCC . Clemastine , a histamine type-2 receptor antagonist augmented the P60568 -induced-ADCC . The P01375 antagonist thalidomide suppressed ADCC whereas pentoxifylline proved to be ineffective . Other tested drugs namely ibuprofen and indomethacin , both prostaglandin E2 antagonists , cimetidine a histamine type-2 receptor antagonist , the opioid pethidine , prostaglandin E2 and histamine exerted minor effects or had no influence on the tested parameters . We conclude that glucocorticosteroids should be avoided with monoclonal antibody and cytokine treatment . According to our in vitro data the other drugs tested did not have a negative impact on cellular cytotoxicity and ADCC . Synthesis , biological activity and HPLC validation of 1,2,3,4-tetrahydroacridine derivatives as acetylcholinesterase inhibitors . The synthesis and biochemical evaluation of new hybrids of tacrine ( DB00382 ) and 4-fluorobenzoic acid ( 4-FBA ) possessing activity towards acetylcholinesterase ( P22303 ) and butyrylcholinesterase ( BuChE ) inhibition are presented . The compounds of interest were obtained from the reaction of activated 4-FBA and diamino derivatives of 1,2,3,4-tetrahydroacridine . The compounds P13671 -2KW/HCl , P13671 -4KW/HCl and P13671 -3KW/HCl have four-fold higher antiacetylcholinesterase activity than DB00382 . All of the acquired compounds present higher selectivity towards P22303 than DB00382 and lower selectivity towards BuChE . In addition , a rapid , selective and stability-indicating HPLC method was developed and validated for the determination of P13671 -2KW/HCl , P13671 -3KW/HCl and P13671 -4KW/HCl . DB00382 and 4-FBA were found to be the main impurities . Chromatographic separation was achieved isocratically on a Waters Symmetry C18 150 × 3.9 mm , 4 μm column with a mobile phase of acetonitrile/buffer ( 17 mM sodium dodecyl sulphate and 8.3 mM sodium dihydrogen phosphate , 50:50 v/v ) ( overall pH 4 ) . A 1.5 ml/min flow rate and a 247 nm wavelength were chosen for this method . P13671 -2KW/HCl , P13671 -3KW/HCl and P13671 -4KW/HCl were subjected to acidic and basic hydrolysis , chemical oxidation , thermal exposition at 60 °C and intense UV light . The limits of detection ( LOD ) and quantification ( LOQ ) were less than 2 μg/ml and 6 μg/ml for P13671 -2KW/HCl , P13671 -3KW/HCl and P13671 -4KW/HCl , 0.04 μg/ml and 0.12 μg/ml for DB00382 , 0.42 μg/ml and 1.41 μg/ml for 4-FBA , respectively . Molecular and biochemical analysis of calmodulin interactions with the calmodulin-binding domain of plant glutamate decarboxylase . We previously provided what to our knowledge is the first evidence that plant glutamate decarboxylase ( Q99259 ) is a calmodulin ( P62158 ) -binding protein . Here , we studied the Q99259 P62158 -binding domain in detail . A synthetic peptide of 26 amino acids corresponding to this domain forms a stable complex with Ca2+/ P62158 with a 1:1 stoichiometry , and amino acid substitutions suggest that tryptophan-485 has an indispensable role in P62158 binding . Chemical cross-linking revealed specific P62158 / Q99259 interactions even in the absence of Ca2+ . However , increasing KCI concentrations or deletion of two carboxy-terminal lysines abolished these interactions but had a mild effect on P62158 / Q99259 interactions in the presence of Ca2+ . We conclude that in the presence of Ca(2+)-hydrophobic interactions involving tryptophan-485 and electrostatic interactions involving the carboxy-terminal lysines mediate P62158 / Q99259 complex formation . By contrast , in the absence of Ca2+ , P62158 / Q99259 interactions are essentially electrostatic and involve the carboxy-terminal lysines . In addition , a tryptophan residue and carboxy-terminal lysines are present in the P62158 -binding domain of an Arabidopsis Q99259 . Finally , we demonstrate that petunia Q99259 activity is stimulated in vitro by Ca2+/ P62158 . Our study provides a molecular basis for Ca(2+)-dependent P62158 / Q99259 interactions and suggests the possible occurrence of Ca(2+)-independent P62158 / Q99259 interactions . Interplay between inhibitory ferric and stimulatory curcumin regulates phosphorylation-dependent human cystic fibrosis transmembrane conductance regulator and ΔF508 activity . Curcumin potentiates cystic fibrosis transmembrane conductance regulator ( P13569 ) activation in an DB00171 -independent but phosphorylation-dependent manner . The underlying molecular mechanisms are unclear . Here , P29320 -293T cells cultured in an Fe(3+)-containing medium were transiently transfected with P13569 constructs , and the role of the inhibitory Fe(3+) bridge between intracellular loop 3 and the regulatory domain of P13569 in this pathway was investigated . The results showed that ethylenediaminetetraacetic acid ( DB00974 ) stimulated phosphorylation-dependent P13569 activation and the stimulation was suppressed by the deletion of the regulatory domain or the insertion of a C832A mutation that removes the Fe(3+)-binding interface . Furthermore , curcumin potentiation of P13569 was significantly weakened not only by Fe(3+)-insensitive mutations at the interface between the regulatory domain and intracellular loop 3 but also by N-ethylmaleimide or DB00974 pretreatment that removes Fe(3+) . More importantly , potentiation of P13569 was completely suppressed by sufficient Fe(3+) . Finally , the insertion of Fe(3+)-insensitive H950R/S768R increased the curcumin-independent activity of ΔF508 but weakened its curcumin potentiation . Thus , Fe(3+) homeostasis in epithelia may play a critical role in regulating P13569 activity , and targeting Fe(3+)-chelating potentiators may direct new therapies for cystic fibrosis . Identification of cell surface targets for HIV-1 therapeutics using genetic screens . Human immunodeficiency virus ( HIV ) drugs designed to interfere with obligatory utilization of certain host cell factors by virus are less likely to encounter development of resistant strains than drugs directed against viral components . Several cellular genes required for productive infection by HIV were identified by the use of genetic suppressor element ( GSE ) technology as potential targets for anti-HIV drug development . Fragmented cDNA libraries from various pools of human peripheral blood mononuclear cells ( PBMC ) were expressed in vitro in human immunodeficiency virus type 1 ( HIV-1 ) -susceptible cell lines and subjected to genetic screens to identify GSEs that interfered with viral replication . After three rounds of selection , more than 15000 GSEs were sequenced , and the cognate genes were identified . The GSEs that inhibited the virus were derived from a diverse set of genes including cell surface receptors , cytokines , signaling proteins , transcription factors , as well as genes with unknown function . Approximately 2.5 % of the identified genes were previously shown to play a role in the HIV-1 life cycle ; this finding supports the biological relevance of the assay . GSEs were derived from the following 12 cell surface proteins : P61073 , CCR4 , P32248 , P20702 , P16070 , Q08722 , P34810 , Q07108 , P04233 , Q99062 , Q9UBS5 , and P20333 . Requirement of some of these genes for viral infection was also investigated by using RNA interference ( RNAi ) technology ; accordingly , 10 genes were implicated in early events of the viral life cycle , before viral DNA synthesis . Thus , these cell surface proteins represent novel targets for the development of therapeutics against HIV-1 infection and AIDS . Stage-dependent inhibition of Plasmodium falciparum by potent Ca2+ and calmodulin modulators . The effects of Ca2+ channel blockers , verapamil , nicardipine and diltiazem , and of potent calmodulin ( P62158 ) inhibitors , trifluoperazine ( Q9HCM9 ) , calmidazolium , W-7 and W-5 , on Plasmodium falciparum in culture were examined . Among Ca2+ blockers , nicardipine was the most potent with the 50 % inhibitory concentration ( IC50 ) of 4.3 microM at 72 h after culture . Parasites were more sensitive to calmidazolium and W-7 with IC50 of 3.4 and 4.5 microM , respectively , than to Q9HCM9 and W-5 . All Ca2+ blockers and P62158 inhibitors suppressed parasite development at later stages . DB00622 , diltiazem , calmidazolium and W-5 also retarded parasite development at earlier stages and/or subsequent growth following pretreatment . Verapamil , nicardipine , Q9HCM9 and calmidazolium reduced erythrocyte invasion by merozoites . Fluorescence microscopy with the cationic fluorescent dye rhodamine 123 revealed that nicardipine , Q9HCM9 and calmidazolium depolarized both the plasma membrane and mitochondrial membrane potentials of the parasite . It is therefore considered that although all Ca2+ and P62158 antagonists tested here influence parasite development at later stages , they are multifunctional , having effects not directly associated with Ca2+ channels or P62158 . Alloreactive memory T cells are responsible for the persistence of graft-versus-host disease . Graft-vs-host disease ( GVHD ) is caused by a donor T cell anti-host reaction that evolves over several weeks to months , suggesting a requirement for persistent alloreactive T cells . Using the C3H.SW anti-C57BL/6 ( B6 ) mouse model of human GVHD directed against minor histocompatibility Ags , we found that donor CD8(+) T cells secreting high levels of P01579 in GVHD B6 mice receiving C3H.SW naive CD8(+) T cells peaked by day 14 , declined by day 28 after transplantation , and persisted thereafter , corresponding to the kinetics of a memory T cell response . Donor CD8(+) T cells recovered on day 42 after allogeneic bone marrow transplantation expressed the phenotype of P16070 (high)CD122(high)CD25(low) , were able to homeostatically survive in response to P60568 , P13232 , and P40933 and rapidly proliferated upon restimulation with host dendritic cells . Both allogeneic effector memory ( P16070 (high)CD62L(low) ) and central memory ( P16070 (high)CD62L(high) ) CD8(+) T cells were identified in B6 mice with ongoing GVHD , with effector memory CD8(+) T cells as the dominant ( > 80 % ) population . Administration of these allogeneic memory CD8(+) T cells into secondary B6 recipients caused virulent GVHD . A similar allogeneic memory P01730 (+) T cell population with the ability to mediate persistent GVHD was also identified in BALB/b mice receiving minor histocompatibility Ag-mismatched B6 T cell-replete bone marrow transplantation . These results indicate that allogeneic memory T cells are generated in vivo during GVH reactions and are able to cause GVHD , resulting in persistent host tissue injury . Thus , in vivo blockade of both alloreactive effector and memory T cell-mediated host tissue injury may prove to be valuable for GVHD prevention and treatment . Association study of 45 candidate genes in nicotine dependence in Han Chinese . Numerous genetic linkages , association studies have been performed in different ethnic groups and revealed many susceptibility loci and genes for nicotine dependence . However , limited similar researches were performed in Han Chinese . This study was designed to investigate the association of candidate genes with nicotine dependence in Han Chinese . We genotyped 384 SNPs within 45 candidate genes with nicotine dependence in a Han Chinese population consisting 223 high nicotine dependent subjects and 257 low nicotine dependent subjects by employing GoldenGate genotyping assay ( Illumina ) . Following association analysis was performed using PLINK software . Individual SNP-based association analysis revealed that nine SNPs located in P35462 ( rs2630351 ) , P21918 ( rs1967550 ) , Q9Y6R4 ( rs2314378 ) , DDC ( rs11575461 ) , Q05901 ( rs4954 ) , O75899 ( rs2779562 ) , P14416 ( rs11214613 and rs6589377 ) and P43681 ( rs2236196 ) were significantly associated with FTND after correction for multiple testing with the p values from 2.59×10(-7) to 9.99×10(-5) . Haplotype-based association analysis revealed haplotype G-A-A formed by rs2630351 , rs167771 and rs324032 and haplotype G-G-G-A formed by rs3773678 , rs2630349 , rs2630351 and rs167771 in P35462 ; haplotype of G-A formed by rs2779562 and rs2808566 in O75899 and haplotype of T-T-A-G-A formed by rs6832644 , rs4057797 , rs9764 , rs4552421 and rs10033119 in P25929 are associated with FTND ( p=3.61×10(-7)-8.78×10(-6) ) . Our results provided confirmation of the previous findings that P14416 , P35462 , DDC , Q05901 , O75899 and P43681 are associated with nicotine dependence . Furthermore , we for the first time report a significant association between nicotine dependence and P21918 , Q9Y6R4 and P25929 . These findings need independent replication in the future studies .