interacts_with DB06813?
Managing the underlying cause of cystic fibrosis : a future role for potentiators and correctors . Cystic fibrosis ( CF ) , a severe genetic disease , is caused by mutations that alter the structure and function of P13569 , a plasma membrane channel permeable to chloride and bicarbonate . Defective anion transport in CF irreversibly damages the lungs , pancreas , liver , and other organs . CF mutations cause loss of P13569 function in multiple ways . In particular , class 3 mutations such as p.Gly551Asp strongly decrease the time spent by P13569 in the open state ( gating defect ) . Instead , class 2 mutations impair the maturation of P13569 protein and its transport from the endoplasmic reticulum to the plasma membrane ( trafficking defect ) . The deletion of phenylalanine 508 ( p.Phe508del ) , the most frequent mutation among CF patients ( 70-90 % ) , destabilizes the P13569 protein , thus causing both a trafficking and a gating defect . These two defects can be overcome with drug-like molecules generically called correctors and potentiators , respectively . The potentiator Kalydeco™ ( also known as DB08820 or VX-770 ) , developed by Vertex Pharmaceuticals , has been recently approved by the US FDA and the European Medicines Agency ( P15941 ) for the treatment of CF patients carrying at least one P13569 allele with the p.Gly551Asp mutation ( 2-5 % of all patients ) . In contrast , the corrector VX-809 , which significantly improves p.Phe508del- P13569 trafficking in vitro , is still under study in clinical trials . Because of multiple defects caused by the p.Phe508del mutation , it is probable that rescue of the mutant protein will require combined treatment with correctors having different mechanisms of action . This review evaluates the status of experimental and clinical research in pharmacotherapy for the CF basic defect . A phase II study of pralatrexate with vitamin B12 and folic acid supplementation for previously treated recurrent and/or metastatic head and neck squamous cell cancer . BACKGROUND : DB06813 ( Fotolyn(TM) ; Allos Therapeutics Inc. ) is an antifolate dihydrofolate reductase ( P00374 ) inhibitor . We conducted a phase II study of pralatrexate with folic acid and B12 supplementation in patients with recurrent and/or metastatic head and neck squamous cell cancer ( R/M HNSCC ) . PATIENTS AND METHODS : This was a single-arm , Simon optimal two stage phase II study . Patients with R/M HNSCC previously treated with chemotherapy were eligible . The study was initiated with a dosing schedule of pralatrexate 190 mg/m(2) biweekly on a 4-week cycle with vitamin supplementation . Due to toxicity concerns , the dosing was modified to 30 mg/m(2) weekly for 3 weeks in a 4-week cycle with vitamin supplementation . Radiologic imaging was to be obtained about every 2 cycles . RESULTS : Thirteen subjects were enrolled ; 12 were treated . Seven of the twelve patients had previously received ≥2 lines of chemotherapy . The most common grade 3 toxicity was mucositis ( 3 patients ) . Seven patients did not complete two cycles of therapy due to progression of disease ( 4 ) , toxicity ( 1 ) , death ( 1 ) , and withdrawal of consent ( 1 ) . Two deaths occurred : one due to disease progression and the other was an unwitnessed event that was possibly related to pralatrexate . No clinical activity was observed . The median overall survival was 3.1 months . The study was closed early due to lack of efficacy . CONCLUSIONS : DB06813 does not possess clinical activity against previously treated R/M HNSCC . Evaluation of pralatrexate in other clinical settings of HNSCC management with special considerations for drug toxicity may be warranted . Endogenous concentrations of ouabain act as a cofactor to stimulate fluid secretion and cyst growth of in vitro ADPKD models via DB02527 and P00533 -Src-MEK pathways . In autosomal-dominant polycystic kidney disease ( ADPKD ) , renal cysts develop by aberrant epithelial cell proliferation and transepithelial fluid secretion . We previously showed that ouabain increases proliferation of cultured human ADPKD cells via stimulation of the P01133 receptor ( P00533 ) -Src-MEK/ P29323 signaling pathway . We examined whether ouabain affects fluid secretion and in vitro cyst growth of human ADPKD cell monolayers , ADPKD cell microcysts cultured in a three-dimensional collagen matrix , and metanephric organ cultures from Pkd1(m1Bei) mice . Physiological concentrations of ouabain alone did not affect net transepithelial basal-to-apical fluid transport in ADPKD monolayers or growth of cultured ADPKD microcysts . In contrast , in the presence of forskolin or 8-bromo- DB02527 , ouabain significantly enhanced ADPKD fluid secretion and microcyst expansion . Ouabain exerted this effect by enhancing DB02527 -dependent Cl(-) secretion via the P13569 . Similarly , ouabain accelerated DB02527 -dependent cyst enlargement in Pkd1(m1Bei) mice metanephroi , with a more prominent response in homozygous than heterozygous mice . Ouabain had no effect on fluid secretion and cystogenesis of normal human kidney cells and caused only slight cystic dilations in wild-type mouse kidneys . The effects of ouabain in ADPKD cells and Pkd1(m1Bei) metanephroi were prevented by inhibitors of P00533 ( AG1478 ) , Src ( Q99463 ) , and MEK ( U0126 ) . Together , our results show that ouabain , used in physiological concentrations , has synergistic effects on DB02527 -mediated fluid secretion and cyst growth , via activation of the P00533 -Src-MEK pathway . These data provide important evidence for the role of ouabain as an endogenous hormone that exacerbates ADPKD cyst progression . LC-MS based assay to measure intracellular compound levels in Mycobacterium smegmatis : linking compound levels to cellular potency . P00374 ( P00374 ) plays a central role in maintaining cellular pool of tetrahydrofolic acid , a cofactor necessary for DNA , RNA and protein synthesis . The clinical validation of P00374 as antibacterial target was established by the success of trimethoprim ( P54849 ) . P00374 is also an attractive target for identifying anti-tuberculosis molecules however , due to observed weak cellular potency , no P00374 inhibitors have been developed as drugs so far . P54849 and its analogs have poor cellular potency on Mycobacterium tuberculosis and Mycobacterium smegmatis cells . We found a mutant strain of M. smegmatis , mc²155 to be sensitive to P54849 whereas wild type strain was not inhibited by P54849 . We utilized this system to probe if poor or lack of activity of P54849 is a consequence of poor intracellular compound levels . An LC-MS based method was developed for measuring P54849 and rifampicin ( Q9HBH0 ) in M. smegmatis . Using the assay , equivalent Q9HBH0 levels were observed in both strains however , P54849 was detected only in mc²155 cells , hence proving a positive correlation between potency and compound levels . To the best of our knowledge this is the first time LC-MS method has been used to measure compound levels in mycobacterial cells . We propose it to be a valuable tool to understand the lack of potency or resistance mechanisms in antimycobacterial drug development . Augmentation by citalopram of risperidone-induced monoamine release in rat prefrontal cortex . RATIONALE : A typical antipsychotics ( APDs ) , e.g. olanzapine and risperidone , have been reported to be effective adjunctive treatment for depression if selective serotonin ( 5-HT ) reuptake inhibitors ( SSRIs ) alone are ineffective . OBJECTIVES AND METHODS : We utilized microdialysis in awake , freely moving rats to study the effect of risperidone in combination with citalopram , an SSRI , on extracellular 5-HT , dopamine ( DA ) , and norepinephrine ( NE ) efflux in rat medial prefrontal cortex ( mPFC ) . RESULTS : DB00734 ( 1.0 mg/kg , s.c. ) , given alone , significantly increased 5-HT , DA , and NE concentrations in the mPFC . DB00215 ( 10 mg/kg , s.c. ) , by itself , produced a significant increase in 5-HT levels only . The combination of risperidone and citalopram produced significantly greater increases in efflux of both DA and NE than risperidone alone . However , the effect of this combination on extracellular 5-HT concentrations was not significantly different than that of citalopram alone . The augmentation of DA and NE efflux induced by risperidone plus citalopram could be partially blocked by the selective P08908 antagonist , WAY 100635 ( 0.2 mg/kg , s.c. ) . CONCLUSIONS : The results suggest that the ability of atypical APDs to augment the therapeutic efficacy of SSRIs in major depression and treatment-resistant depression may be due , at least in part , to potentiation of SSRI-induced increases in cortical DA and NE . The contributions of P08908 receptor stimulation and 5- Q13049 and alpha2 adrenergic receptor antagonism to this augmentation are discussed . Gating properties of Q14524 mutations and the response to mexiletine in long-QT syndrome type 3 patients . BACKGROUND : DB00379 ( Mex ) has been proposed as a gene-specific therapy for patients with long-QT syndrome type 3 ( LQT3 ) caused by mutations in the cardiac sodium channel gene ( Q14524 ) . The degree of QT shortening and the protection from arrhythmias vary among patients harboring different mutations . We tested whether the clinical response to Mex in LQT3 could be predicted by the biophysical properties of the different mutations . METHODS AND RESULTS : We identified 4 Q14524 mutations in 5 symptomatic LQT3 patients with different responses to Mex ( 6 to 8 mg . kg(-1) . d(-1) ) . We classified the mutations as sensitive to Mex ( P1332L , R1626P ; >/= 10 % of QTc shortening and QTc < 500 ms or no arrhythmias ) or insensitive to Mex ( S941N , M1652R ; negligible or no QTc shortening and sudden death ) . We measured Na(+) current from P29320 293 cells transfected with wild-type ( WT ) or mutant Nav1.5 . All mutations showed impaired inactivation of Na(+) current , but the mutations identified in patient responders to Mex ( P1332L , R1626P ) showed a hyperpolarizing shift of V(1/2) of steady-state inactivation . Furthermore , Mex produced use-dependent block with the order R1626P=P1332L > S941N=WT > M1652R , suggesting that Mex-sensitive mutants present prolonged recovery from Mex block . CONCLUSIONS : We propose that voltage dependence of channel availability and shifts of V(1/2) of steady-state inactivation correlate with the clinical response observed in LQT3 patients . This supports the view that the response to Mex is mutation specific and that in vitro testing may help to predict the response to therapy in LQT3 . Celecoxib with chemotherapy in colorectal cancer . P35354 ( P35354 ) is the enzyme that normally synthesizes prostaglandins during an inflammatory response . Many primary and metastatic cancers express P35354 , and its presence is correlated with tumor angiogenesis , more invasive tumor phenotype , resistance to apoptosis , and systemic immunosuppression . The expression of P35354 is associated with a worse prognosis . Inhibition of prostaglandin synthesis may be beneficial in human malignancy . Regular consumption of nonsteroidal anti-inflammatory drugs ( NSAIDs ) decreases the incidence of , and mortality rate resulting from , a number of types of gastrointestinal cancers . Premalignant colonic lesions regress following the administration of nonspecific P36551 inhibitors , such as sulindac ( DB00605 ) . Advanced solid tumor patients treated with indomethacin ( DB00328 ) survive twice as long as do such patients who receive supportive care alone . The U.S . Food and Drug Administration has approved specific P35354 inhibitors for the treatment of arthritis , pain , and familial adenomatous polyposis . Preclinical studies show that these drugs block angiogenesis , suppress solid tumor metastases , and slow the growth of implanted gastrointestinal cancer cell lines . The P35354 inhibitors have safely and effectively been combined with chemotherapeutic agents in experimental studies . Ongoing clinical trials are currently assessing the potential therapeutic role of P35354 inhibitors in both prevention and treatment of a diverse range of human cancers . Characterization of the aggregation responses of camel platelets . BACKGROUND : Despite evidence of active hemostasis , camel platelets barely respond to common aggregating agents at standard doses used for human platelet aggregation . OBJECTIVES : The purpose of the study was to find out whether camel platelets can be activated by high doses or combinations of aggregation agonists , and to characterize the receptor that mediates the aggregation response to adenosine diphosphate ( ADP ) , the most potent agonist for camel platelets known so far . METHODS : Aggregation studies were performed with platelet-rich plasma ( PRP ) in response to multiple doses or combinations of ADP , epinephrine ( P08473 ) , collagen , and arachidonic acid ( AA ) . Aggregation responses to ADP were performed before and after the addition of the ADP receptor ( Q9H244 ) antagonist DB00758 . RESULTS : Camel platelets responded to ADP at doses higher than the standard dose for human platelets , and to combinations of P08473 and other agonists , while no aggregation was elicited with P08473 or AA alone . DB00758 blocked the ADP-induced aggregation responses in a dose-dependent fashion in vitro . CONCLUSIONS : Camel platelet aggregation can be activated by increasing the dose of some agonists such as ADP , but not AA or P08473 . Irreversible aggregation of camel platelets could also be triggered by a combination of P08473 and ADP , and collagen and AA . Inhibition with clopidogrel suggests that camel platelets express the ADP receptor , Q9H244 . Understanding platelet function in camels will add to the understanding of platelet function in health and disease . Extracellular signal-regulated kinase/mitogen-activated protein kinases block internalization of delta-opioid receptors . Translocation of G protein-coupled receptors ( GPCRs ) from the cell membrane to cytosol depends on the kind of ligand activating the receptor . This principle is clearly demonstrated for opioid receptors , because diverse opiate agonists rapidly induce receptor internalization , whereas morphine almost fails . We report here the impact of mitogen-activated protein ( Q96HU1 ) kinase isoforms extracellular signal-regulated kinase ( P29323 )1/2 on the internalization of delta-opioid receptors ( DORs ) expressed in human embryonic kidney ( P29320 )293 cells . Receptor activation by etorphine turned out to transiently phosphorylate P29323 / Q96HU1 kinases and bring about Q8IXH6 internalization within 20 min . In contrast , prolonged exposure of HEK293 cells to morphine excited persistent phosphorylation of P29323 / Q96HU1 kinases , and those cells failed to internalize the opioid receptor . When P29323 / Q96HU1 kinase phosphorylation was blocked by 2'-Amino-3'-methoxyflavone ( PD98059 ) , morphine gained the ability to strongly induce Q8IXH6 endocytosis . The importance of activated Q96HU1 kinases for Q8IXH6 internalization is further demonstrated by glutamate and paclitaxel because these substances induce phosphorylation of P27361 /2 and concomitantly prevent Q8IXH6 sequestration by etorphine . In addition , receptor internalization by morphine was facilitated by inhibition of protein kinase C and opioid-mediated transactivation of epidermal growth factor receptor ( P00533 ) , both activating P29323 / Q96HU1 kinases by opioids . The mechanism affording Q8IXH6 internalization by PD98059 may relate to arrestin , which uncouples GPCRs and thus triggers receptor internalization . Arrestin considerably translocates toward the cell membrane upon Q8IXH6 activation by morphine in presence of the Q96HU1 kinase blocker , but it fails in the absence of PD98059 . We conclude that P29323 / Q96HU1 kinase activity prevents opioid receptor desensitization and sequestration by blocking arrestin 2 interaction with activated DORs . Flow cytometric analysis of mammalian glial cultures treated with methotrexate . DB00563 ( MTX ) is an antineoplastic drug that acts by competitive inhibition of the enzyme dihydrofolate reductase ( P00374 ) . MTX treatment of cultured cell lines leads to the emergence of resistant cell populations . Studies using stepwise selection procedures have demonstrated that MTX resistance conferred by overproduction of P00374 can be caused by P00374 gene amplification . We examined the effect of MTX on cells whose origin more closely approximates the in vivo condition by developing a culture system using dissociated brain tissue from 17-19 day old mouse embryos . At the first passage , cultures were divided into control and MTX groups . Cells were treated with the same or successively higher concentrations of MTX at each passage over a 3-4 month period . The first passage eliminated neurons and left a glial culture comprised of approximately 90 % astrocytes . We used the Fluorescence Activated Cell Sorter in conjunction with fluorescent dyes to measure P00374 content , DNA content , size , and viability of glial cells following MTX treatment . MTX-treated cells divided but grew more slowly and were larger than untreated cells . Stepwise selection in 30/60/90 nM or 60/120 nM MTX resulted in significant two- to threefold increases in fluorescence , and hence P00374 levels . Slot hybridizations assays demonstrated a threefold increase in P00374 gene copy number in the DNA from the 30/60/90 cultures . Thus , our findings were consistent with the results obtained from somatic cell lines , and lend support to the hypothesis that gene amplification may be a common mechanism for the acquisition of resistance in many types of cells . They also indicate that glial cells may be a specific target for cytotoxic effects of MTX on the central nervous system . Evaluation of the pharmacokinetics , preclinical and clinical efficacy of pralatrexate for the treatment of T-cell lymphoma . INTRODUCTION : Peripheral T-cell lymphomas ( PTCLs ) are a heterogeneous group of T-cell neoplasms . Most patients with PTCL have a poor outcome with conventional therapies and are not cured without stem-cell transplantation . DB06813 , a novel antifolate chemotherapeutic agent , was rationally designed to impede folate metabolism by inhibiting dihydrofolate reductase ( P00374 ) and to be more efficiently internalized into tumor cells . DB06813 is the first drug that is FDA approved for patients with relapsed and refractory PTCL . AREAS COVERED : DB06813 has been used as a single agent and in combination with other agents in clinical trials for non-Hodgkin 's lymphoma and Hodgkin 's disease as well as in solid tumors . This review will cover the development of pralatrexate , the pharmacokinetics of pralatrexate , preclinical findings with pralatrexate and clinical studies of pralatrexate in hematologic malignancies . EXPERT OPINION : DB06813 has significant activity in vitro , and in early Phase I/II trials , responses were noted in patients with aggressive T-cell lymphomas . The DB06813 in Patients with Relapsed or Refractory Peripheral T-Cell Lymphoma trial demonstrated the activity of pralatrexate across a spectrum of heavily pretreated patients with different aggressive T-cell lymphoma subtypes , and studies in cutaneous T-cell lymphoma have shown efficacy at different doses and schedules . The most frequent adverse events in these trials were mucositis , reversible thrombocytopenia and fatigue . DB09280 - DB08820 in Patients with Cystic Fibrosis Homozygous for Phe508del P13569 . Distinct mechanistic activity profile of pralatrexate in comparison to other antifolates in in vitro and in vivo models of human cancers . PURPOSE : This study evaluated mechanistic differences of pralatrexate , methotrexate , and pemetrexed . METHODS : Inhibition of dihydrofolate reductase ( P00374 ) was quantified using recombinant human P00374 . Cellular uptake and folylpolyglutamate synthetase ( Q05932 ) activity were determined using radiolabeled pralatrexate , methotrexate , and pemetrexed in NCI-H460 non-small cell lung cancer ( NSCLC ) cells . The tumor growth inhibition ( TGI ) was assessed using MV522 and NCI-H460 human NSCLC xenografts . RESULTS : Apparent K ( i ) values for P00374 inhibition were 45 , 26 , and > 200 nM for pralatrexate , methotrexate , and pemetrexed , respectively . A significantly greater percentage of radiolabeled pralatrexate entered the cells and was polyglutamylatated relative to methotrexate or pemetrexed . In vivo , pralatrexate showed superior anti-tumor activity in both NSCLC models , with more effective dose-dependent TGI in the more rapidly growing NCI-H460 xenografts . CONCLUSIONS : DB06813 demonstrated a distinct mechanistic and anti-tumor activity profile relative to methotrexate and pemetrexed . DB06813 exhibited enhanced cellular uptake and increased polyglutamylation , which correlated with increased TGI in NSCLC xenograft models . Ca2+-calmodulin and janus kinase 2 are required for activation of sodium-proton exchange by the Gi-coupled 5-hydroxytryptamine 1a receptor . The type 1 sodium-proton exchanger ( P19634 ) is expressed ubiquitously and regulates key cellular functions , including mitogenesis , cell volume , and intracellular pH . Despite its importance , the signaling pathways that regulate P19634 remain incompletely defined . In this work , we present evidence that stimulation of the 5-hydroxytryptamine 1A ( P08908 ) receptor results in the formation of a signaling complex that includes activated O60674 ( Jak2 ) , Ca2+/calmodulin ( P62158 ) , and P19634 , and which involves tyrosine phosphorylation of P62158 . The signaling pathway also involves rapid agonist-induced association of P62158 and P19634 as assessed by coimmunoprecipitation studies and by bioluminescence resonance energy transfer studies in living cells . We propose that P19634 is activated through this pathway : P08908 receptor --> G(i2)alpha and/or G(i3)alpha --> Jak2 activation --> tyrosine phosphorylation of P62158 --> increased binding of P62158 to P19634 --> induction of a conformational change in P19634 that unmasks an obscured proton-sensing and/or proton-transporting region of P19634 --> activation of P19634 . The G(i/o)-coupled P08908 receptor now joins a handful of Gq-coupled receptors and hypertonic shock as upstream activators of this emerging pathway . In the course of this work , we have presented clear evidence that P62158 can be activated through tyrosine phosphorylation in the absence of a significant role for elevated intracellular Ca2+ . We have also shown for the first time that the association of P62158 with P19634 in living cells is a dynamic process . DB00472 induces preventive and complex effects against colon cancer development in epithelial and stromal areas in rats . DB00472 ( FLX ) is a drug commonly used as antidepressant . However , its effects on tumorigenesis remain controversial . Aiming to evaluate the effects of FLX treatment on early malignant changes , we analyzed serotonin ( 5-HT ) metabolism and recognition , aberrant crypt foci ( Q9NQ94 ) , proliferative process , microvessels , vascular endothelial growth factor ( P15692 ) , and cyclooxygenase-2 ( P35354 ) expression in colon tissue . Male Wistar rats received a daily FLX-gavage ( 30mgkg(-1) ) and , a single dose of 1,2 dimethylhydrazine ( Q03001 ; i.p. , 125mgkg(-1) ) . After 6 weeks of FLX-treatment , our results revealed that FLX and nor-fluoxetine ( N-FLX ) are present in colon tissue , which was related to significant increase in serotonin ( 5-HT ) levels ( P < 0.05 ) possibly through a blockade in P31645 mRNA ( serotonin reuptake transporter ; P < 0.05 ) resulting in lower 5-hydroxyindoleacetic acid ( 5-HIAA ) levels ( P < 0.01 ) and , P28335 receptor mRNA expressions . FLX-treatment decreased dysplastic Q9NQ94 development ( P < 0.01 ) and proliferative process ( P < 0.001 ) in epithelia . We observed a significant decrease in the development of malignant microvessels ( P < 0.05 ) , P15692 ( P < 0.001 ) , and P35354 expression ( P < 0.01 ) . These findings suggest that FLX may have oncostatic effects on carcinogenic colon tissue , probably due to its modulatory activity on 5-HT metabolism and/or its ability to reduce colonic malignant events . Purine receptor Q15077 mediates cellular response to γ-ray-induced DNA damage . We previously showed that nucleotide P2 receptor agonists such as DB00171 and UTP amplify γ-ray-induced focus formation of phosphorylated histone H2A variant P16104 ( γ P16104 ) , which is considered to be an indicator of DNA damage so far , by activating purine Q15077 and Q9H244 receptors . Therefore , we hypothesized that these P2 receptors play a role in inducing the repair response to γ-ray-induced DNA damage . In the present study , we tested this idea by using human lung cancer A549 cells . First , reverse-transcription polymerase chain reaction ( RT-PCR ) showed that Q15077 receptor is highly expressed in A549 cells , but Q9H244 receptor is only weakly expressed . Next , colony formation assay revealed that Q15077 receptor antagonist MRS2578 markedly reduced the survival rate of γ-ray-exposed A549 cells . The survival rate was also significantly reduced in Q15077 -knock-down cells , compared with scramble siRNA-transfected cells . Since it has reported that phosphorylation of P27361 /2 after activation of P00533 via Q15077 and Q9H244 receptors is involved in the repair response to γ-ray-induced DNA damage , we next examined whether γ-ray-induced phosphorylation of P27361 /2 was also inhibited by MRS2578 in A549 cells . We found that it was . Taken together , these findings indicate that purinergic signaling through Q15077 receptor , followed by P27361 /2 activation , promotes the cellular repair response to γ-ray-induced DNA damage . Purinergic receptors contribute to early mesangial cell transformation and renal vessel hypertrophy during angiotensin II-induced hypertension . Chronic P03950 II infusions lead to increases in intrarenal P03950 II levels , hypertension , and tissue injury . Increased blood pressure also elicits increases in renal interstitial fluid ( Q9HBH0 ) DB00171 concentrations that stimulate cell proliferation . We evaluated the contribution of purinergic receptor activation to P03950 II-induced renal injury in rats by treating with clopidogrel , a Q9H244 receptor blocker , or with PPADS , a nonselective P2 receptor blocker . alpha-Actin expression in mesangial cells , afferent arteriolar wall thickness ( AAWT ) , cortical cell proliferation , and macrophage infiltration were used as early markers of renal injury . DB00758 and PPADS did not alter blood pressure , renin or kidney P03950 II content . alpha-Actin expression increased from control of 0.6 +/- 0.4 % of mesangial area to 6.3 +/- 1.9 % in P03950 II-infused rats and this response was prevented by clopidogrel ( 0.4 +/- 0.2 % ) and PPADS . The increase in AAWT from 4.7 +/- 0.1 to 6.0 +/- 0.1 mm in P03950 II rats was also prevented by clopidogrel ( 4.8 +/- 0.1 mm ) and PPADS . P03950 II infusion led to interstitial macrophage infiltration ( 105 +/- 16 vs. 62 +/- 4 cell/mm(2) ) and tubular proliferation ( 71 +/- 15 vs. 20 +/- 4 cell/mm(2) ) and these effects were prevented by clopidogrel ( 52 +/- 4 and 36 +/- 3 cell/mm(2) ) and PPADS . Q9HBH0 DB00171 levels were higher in P03950 II-infused rats than in control rats ( 11.8 +/- 1.9 vs. 5.6 +/- 0.6 nmol/l , P < 0.05 ) . The results suggest that activation of vascular and glomerular purinergic P2 receptors may contribute to the mesangial cell transformation , renal inflammation , and vascular hypertrophy observed in P03950 II-dependent hypertension . Establishment of pemetrexed-resistant non-small cell lung cancer cell lines . DB00642 ( P15941 ) , a multitargeted antifolate with manageable toxicity , is active against non-squamous non-small cell lung cancer ; however , most patients eventually acquire resistance to P15941 . To elucidate the resistant mechanism , we established P15941 -resistant lung adenocarcinoma cell lines . Two parental cell lines , PC-9 and A549 , were treated with step-wise increasing concentrations of P15941 . Growth inhibition was determined by the 3-[4,5-dimethyl-thizol-2-yl]-2,5-diphenyltetrazolium bromide assay . Expression of the genes encoding thymidylate synthase ( TS ) , dihydrofolate reductase ( P00374 ) , and glycinamide ribonucleotide formyltransferase ( GARFT ) was analyzed by quantitative real-time reverse transcriptase polymerase chain reaction . The four PC-9 sublines were more resistant than the PC-9 cell line to P15941 ( 2.2- , 2.9- , 8.4- , and 14.3-fold , respectively ) . The four A549 sublines also showed more resistance to P15941 ( 7.8- , 9.6- , 42.3- , and 42.4-fold , respectively ) than the parent cell line . All resistant sublines showed cross-resistance to cisplatin , but not to docetaxel , vinorelbine , 5-fluorouracil , or the active metabolite of irinotecan , SN-38 . All P15941 -resistant sublines expressed more TS than the parental cells , by polymerase chain reaction and Western blotting . P00374 was significantly increased in the four P15941 -resistant A549 sublines . GARFT did not correlate with resistance to P15941 . In summary , P15941 -resistant cells remained sensitive to docetaxel , vinorelbine , 5-fluorouracil , and irinotecan . TS expression appeared to be associated with resistance to P15941 . [ Functional characteristics of calcium-sensitive adenylyl cyclase of ciliate Tetrahymena pyriformis ] . DB01373 -sensitive forms of adenylyl cyclase ( AC ) were revealed in most vertebrates and invertebrates and also in some unicellular organisms , in particular ciliates . We have shown for the first time that calcium cations influence the AC activity of ciliate Tetrahymena pyriformis . These cations at the concentrations of 0.2-20 microM stimulated the enzyme activity , and maximum of catalytic effect was observed at 2 microM Ca2+ . DB01373 cations at a concentrations of 100 microM or higher inhibited the AC activity . P62158 antagonists W-5 and W-7 at the concentrations of 20-100 microM inhibited the catalytic effect induced by 5 microM Ca2+ and blocked the effect at higher concentrations of Ca2+ . DB00477 , another calmodulin antagonist , reduced Ca2+-stimulated AC activity only at the concentrations of 200-1000 microM . AC stimulating effects of serotonin , P01133 and DB02527 increased in the presence of 5 microM Ca2+ . AC stimulating effects of P01133 , DB02527 and insulin decreased in the presence of 100 microM Ca2+ , and AC stimulating effect of DB02527 decreased also in the presence of calmodulin antagonists ( 1 mM ) . At the same time , stimulating effect of D-glucose in the presence of Ca2+ and calmodulin antagonists did not change essentially . The data obtained speak in favor of the presence of calcium-sensitive forms of AC in ciliate T. pyriformis which mediate enzyme stimulation by P01133 , DB02527 , insulin , and serotonin . Suppression of tumor growth and metastasis by a P17948 antagonizing peptide identified from a phage display library . Although the P15692 -Flk-1-pathway has been known as the major driving force of angiogenesis , new evidence has shown that P17948 /Flt-1 plays important roles during the neovascularization under pathological conditions including tumor , atherosclerosis and arthritis . In search of Flt-1 receptor antagonizing peptides , we screened a phage display 12-mer-peptide library with recombinant Flt-1 protein . Seven candidate peptides were identified that specifically bound to P15692 receptor Flt-1 , of which peptide F56 ( WHSDMEWWYLLG ) almost abolished P15692 binding to receptor Flt-1 in vitro . In vivo , F56 fused with P00374 ( P00374 -F56 ) inhibited angiogenesis in a P62158 assay . Moreover , P00374 -F56 significantly inhibited the growth of nodules of human gastric cancer cell line MGC-803 in BALB/c nude mice . Histological analyses showed that necrosis of the implanted tumor was markedly enhanced following treatment with P00374 -F56 . In the severe combined immunodeficiency disease ( SCID ) mouse model for studying metastasis of the human breast cancer cell line BICR-H1 , synthetic peptide F56 significantly inhibited tumor growth and lung metastases . Taken together , our results have demonstrated that peptide F56 , as a Flt-1 receptor antagonist , fulfilled the antiangiogenic and antimetastatic effects by specifically interfering with the interaction between P15692 and receptor Flt-1 . Thus , short peptide F56 may have clinical potential in tumor therapy . [ Effects of chronic fluoxetine treatment on catalepsy and immune response in mice genetically predisposed to freezing reaction : the role of P08908 and 5- Q13049 receptors and tph2 and P31645 genes ] . ASC/Icg ( Antidepressant Sensitive Catalepsy ) mouse strain selected for high predisposition to pinch-induced catalepsy is characterized by depressive-like behavior and impaired immune response . Chronic treatment with SSRI fluoxetine attenuated catalepsy manifestation and normalized a decreased number of rosette-forming cells ( P41440 ) in spleen in ASC mice . Chronic fluoxetine administration had no effect on catalepsy and P41440 number in mice of parental cataleptic CBA/Lac strain . DB00472 failed to alter P08908 receptor functional activity in mice of both strains and diminished 5- Q13049 receptor functional activity in CBA but not in ASC mice . No effect on cortical P08908 and 5- Q13049 receptor mRNA levels and on P08908 receptor , tph2 ( tryptophan hydroxylase-2 ) and P31645 ( serotonin transporter ) mesencephalic gene expression was observed in ASC mice . Other possible serotonergic mechanisms of fluoxetine effect on catalepsy and immune response in mice with depressive-like state are discussed . Order of genes on human chromosome 5q with respect to 5q interstitial deletions . Using ( a ) somatic cell hybrids retaining partial chromosome 5 and ( b ) clinical samples from patients with acquired deletions of the long arm of chromosome 5 , combined with chromosome 5-linked DNA probes , some of which exhibited RFLPs , we have determined the order of a series of genes on chromosome 5 . The order established is 5pter ---- MLVI-2 ---- cen ---- P07686 ---- P00374 ---- Pi227- --- cp12.6 ---- ( P05113 , P05112 ) ---- P08700 ---- P04141 ---- P05230 ---- ( P07333 , P09619 ) ---- ( treC,ADRBR ) ---- ( Q5SW96 - Q13585 , P09603 ) ---- qter . The suggested order and orientation for the closely linked P08700 / P04141 gene pair is cen ---- 5' P08700 3' ---- 5' P04141 3' ---- qter , on the basis of analysis of the P04141 rearrangement in HL60 DNA . The map position of the GRL locus , which was consistent with both somatic cell hybrid and 5q- analyses , was telomeric to P04141 and centromeric to P07333 / P09619 , near P05230 . Long-range restriction-enzyme analysis of 5q- DNAs did not detect rearrangements of 5q-linked probes except in HL60 DNA , but it did reveal putative long-range RFLPs of several loci . RFLPs for GRL , Pi227 , cp12.6 , P08700 , and P07333 can detect deletions in bone marrow and in leukemia cells from patients with acquired 5q deletions . [ Presynaptic P08908 receptors in immunomodulation ] . It is shown that a selective agonist of P08908 receptors 8-OH-DPAT in a low dose ( 0.1 mg/kg ) , which is known to affect mainly the presynaptic P08908 receptors increased the immune response at the peak of reactions ( the forth or fifth day after immunization with sheep red blood cells - SRBC ) in CBA mice and Wistar rats . Treatment of the animals with the drug 15 min prior to antigen injection raised the number of plaque-forming cells ( lgM- P27918 ) and rosette-forming cells ( P41440 ) in the spleen . The preliminary blockade of P08908 receptor with a selective antagonist of P08908 receptors WAY-100635 ( 0.1 mg/kg ) prevented the immunostimulating effect of 5-HT 1A receptors agonist 8-OH-DPAT , whereas WAY-100635 administration alone in the same dose did n't change the immune response . Activation of P08908 receptors under conditions of electrical lesion of 5-HTergic neurons of the nucleus raphe was unable to enhance the immune reactions , as it did in sham-operated rats . The data obtained indicate that the somatodendric P08908 autoreceptors are involved in immunomodulation . Growth factors expression in patients with erosive esophagitis . Although the pathogenesis and treatment of erosive esophagitis ( EE ) is well recognized , little is known about the cellular and molecular mechanisms of mucosal healing in EE patients . In this pilot study , we enrolled typical EE patients to evaluate what kinds of growth factors and their receptors were activated in their injured esophageal mucosa . Forty endoscopically proved EE patients were consecutively enrolled . Messenger RNA expressions , which includes keratinocyte growth factor ( KGF ) and its receptor ( P21802 ) , epidermal growth factor ( P01133 ) and its receptor ( P00533 ) , hepatocyte growth factor ( P14210 ) and its receptor ( HGFR ) , basic fibroblast growth factor ( P09038 ) , vascular endothelial growth factor ( P15692 ) , and cyclooxygenase ( P36551 ) -1 and P35354 , were measured using real-time polymerase chain reaction ( PCR ) . Data were compared between the injured EE mucosa and their normal esophageal mucosa above EE . The mRNA expressions of P14210 , HGFR , P01133 , P15692 , and P35354 , but not P00533 , KGF , P21802 , P09038 , and P23219 , were significantly increased in the injured mucosa of EE patients compared with those of normal mucosa ( P < 0.05 ) . The study found that P14210 , HGFR , P01133 , P15692 , and , P35354 are activated in the injured mucosa of EE patients ; their activation might be involved in mucosal repair and ulcer healing of EE . Glucocorticoid-induced surface expression of annexin 1 blocks beta2-integrin adhesion of human eosinophils to intercellular adhesion molecule 1 surrogate protein . BACKGROUND : Glucocorticoids attenuate the population of eosinophils and T lymphocytes in asthmatic airways . The decrease in airway eosinophilia is caused both by accelerated cell death and by induction of blockade of integrin adhesion . In this study , we examined the hypothesis that annexin 1 surface expression , which is upregulated by the glucocorticoid receptor , prevents integrin adhesion essential to cell migration by blocking intracellular translocation of cytosolic group IV phospholipase A2 ( P47712 ) . OBJECTIVE : To examine the relationship of the glucocorticoid on annexin 1 expression and the effect of blockade of annexin 1 activity on adhesion of human eosinophils in vitro . To determine the relationship between annexin 1surface expression and nuclear membrane translocation of P47712 . METHODS : Eosinophils isolated from human peripheral blood were pretreated with fluticasone propionate ( FP ) , and beta2-integrin adhesion was measured after stimulation with P05113 or eotaxin . Effects of FP on P47712 expression , phosphorylation , and translocation were determined . The role of annexin 1 was examined by using annexin 1 blocking antibody and/or mimetic peptides . RESULTS : DB00588 decreased stimulated eosinophil adhesion and caused 4-fold increase in annexin 1 expression on the plasma membrane . Inhibition of adhesion by FP was blocked with annexin 1 blocking antibody . Annexin 1 N-terminal mimetic peptide also blocked beta2-integrin adhesion . Translocation of P47712 to the nuclear membrane was significantly blocked by incubation with FP . Blockade was reversed with annexin 1 blocking antibody . CONCLUSION : Blockade of beta2-integrin adhesion by glucocorticoid is regulated by annexin 1 , which blocks P47712 translocation to nuclear membrane . DB06813 : a novel synthetic antifolate for relapsed or refractory peripheral T-cell lymphoma and other potential uses . PURPOSE : The pharmacology , pharmacokinetics , clinical trials , adverse effects , dosage , and economic considerations of pralatrexate ( DB06813 ) are reviewed . SUMMARY : Peripheral T-cell lymphoma ( PTCL ) comprises approximately 15-20 % of all aggressive lymphomas and 5-10 % of all non-Hodgkin 's lymphomas . Advanced PTCL is often refractory to traditional first-line chemotherapy regimens . DB06813 was developed as a synthetic folate analog antimetabolite that competitively inhibits dihydrofolate reductase ( P00374 ) . This results in the depletion of thymidine , leading to interference with deoxyribonucleic acid synthesis and cancer cell death . DB06813 has a higher potency than methotrexate and edatrexate ( EDX ) . The efficacy and safety of DB06813 have been demonstrated in the PROPEL trial , a prospective phase II trial in patients with relapsed or refractory PTCL . Patients with prior stem cell transplantation receiving DB06813 also had similar response rates ( RRs ) . DB06813 was investigated on the treatment of relapsed or refractory cutaneous T-cell lymphoma , previously treated advanced non-small cell lung cancer and other solid malignancies . DB06813 has similar side effects to other P00374 inhibitors . The most common side effect of DB06813 is mucositis . The recommended dose of DB06813 is 30 mg/m(2) weekly once for 6 weeks in 7-week cycle until disease progresses or unacceptable toxicity for PTCL and may require dose reduction or discontinuation . Patients should be supplemented with oral folic acid and intramuscular vitamin B(12) injections . CONCLUSION : DB06813 provides clinical benefit to patients with relapsed or refractory PTCL with durable complete and partial responses in patients who had not responded to multiple prior treatment regimens . Development of stable cell lines expressing different subtypes of GABAA receptors . The experiments reported here were motivated by our interest to express in stably-transfected cells large amounts of recombinant rat GABAA receptors . For this , we developed an original two step selection strategy , in which the first step consisted of transfecting P29320 293 cells with rat GABAA receptor alpha and beta subunits . G 418 resistant colonies isolated at this step were screened for [ 3H ] muscimol binding to select for those that coexpressed alpha- and beta-subunits . The best alpha and beta subunit expressing colony was then supertransfected with a plasmid coding for the gamma rat GABAA receptor subunit and a mutant P00374 gene . After a second round of selection , this time in presence of methotrexate , those colonies that coexpressed ternary alpha beta gamma GABAA receptor combinations were distinguished using [ 3H ] flumazenil as a probe . This strategy was applied to the isolation of 3 GABAA receptor clones , alpha 1 beta 2 gamma 2s , alpha 3 beta 2 gamma 2s and alpha 5 beta 3 gamma 2s , that expressed relatively high levels of these proteins . These 3 cell lines exhibited pharmacological and functional properties similar to cells transiently-transfected with equivalent subunit combinations . These cell lines therefore provide attractive models with which to evaluate the intrinsic activity and potency of compounds at recombinant GABAA receptor subtypes . Spline-fitting with a genetic algorithm : a method for developing classification structure-activity relationships . Classification methods allow for the development of structure-activity relationship models when the target property is categorical rather than continuous . We describe a classification method which fits descriptor splines to activities , with descriptors selected using a genetic algorithm . This method , which we identify as SFGA , is compared to the well-established techniques of recursive partitioning ( RP ) and soft independent modeling by class analogy ( SIMCA ) using five series of compounds : cyclooxygenase-2 ( P35354 ) inhibitors , benzodiazepine receptor ( BZR ) ligands , estrogen receptor ( ER ) ligands , dihydrofolate reductase ( P00374 ) inhibitors , and monoamine oxidase ( MAO ) inhibitors . Only 1-D and 2-D descriptors were used . Approximately 40 % of compounds in each series were assigned to a test set , " cherry-picked " from the complete set such that they lie outside the training set as much as possible . SFGA produced models that were more predictive for all but the P00374 set , for which SIMCA was most predictive . RP gave the least predictive models for all but the MAO set . A similar trend was observed when using training and test sets to which compounds were randomly assigned and when gradually eliminating compounds from the ( designed ) training set . The stability of models was examined for the random and reduced sets , where stability means that classification statistics and the selected descriptors are similar for models derived from different sets . Here , SIMCA produced the most stable models , followed by SFGA and RP . We show that a consensus approach that combines all three methods outperforms the single best model for all data sets . Single agent and combination studies of pralatrexate and molecular correlates of sensitivity . BACKGROUND : DB06813 is a dihydrofolate reductase ( P00374 ) inhibitor with high affinity for reduced folate carrier 1 ( P41440 -1 ) and folylpolyglutamate synthetase ( Q05932 ) , resulting in extensive internalization and accumulation in tumour cells . DB06813 is approved in the US for the treatment of relapsed or refractory peripheral T-cell lymphoma and is being investigated in various malignancies . Here , we evaluated molecular correlates of sensitivity to pralatrexate and explored combinations with a variety of anticancer agents . METHODS : Antiproliferative effects of pralatrexate were evaluated in 15 human-cancer cell lines using the MTT assay . Gene expression was evaluated using qRT-PCR . RESULTS : DB06813 and methotrexate had a similar pattern of cytotoxicity , pralatrexate being more potent . DB06813 potentiated the effects of platinum drugs , antimetabolites and P00533 inhibitors . Dose- and time-dependent cytotoxicity of pralatrexate correlated with high mRNA expression of Q05932 . Acquired resistance to pralatrexate was associated with decreased P41440 -1 expression , whereas methotrexate resistance correlated with increased P00374 expression , suggesting different mechanisms of acquired resistance . CONCLUSION : DB06813 was more potent than methotrexate in a panel of solid tumour lines . Our findings support the further clinical development of pralatrexate in combination with certain cytotoxics and targeted therapies , and suggest that P41440 -1 , Q05932 and P00374 may be potential biomarkers of outcome .