Newer asthma therapies.
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Adrenal Cortex Hormones↗
Biomedical subjects
L J Smith
Publications and source records attributed to L J Smith.
Side-chain conformational disorder in a molten globule: molecular dynamics simulations of the A-state of human alpha-lactalbumin.
Molten globules are compact, partially folded forms of proteins consisting of an ensemble of interconverting conformers with disorder in the side-chain packing across the ensemble. Using insights from experimental data a strategy has been devised to describe this side-chain disorder in a molten globule. Structures of human alpha-lactalbumin have been generated with significantly different side-chain packings to those observed in the native protein by changing all the chi1 torsion angles. Molecular dynamics simulations have been carried out starting from these structures using conditions under which the molten globule is seen experimentally, i.e. at low pH and in the absence of calcium. In each simulation, low energy conformers were generated which are compact and retain substantial secondary structure and an overall native fold, but have very different side-chain contacts; there are all-atom root-mean-square differences of up to 0.5 nm between the different structures. The structures from the different simulations taken together provide an initial description of possible contributors to the ensemble of conformers in the molten globule state which is consistent, at least in general terms, with experimental data.
Calcium↗
Anesthesia case of the month. Anesthesia in a dog with tricuspid insufficiency and atrial fibrillation.
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Analgesics, Opioid↗
Long-term safety and efficacy of zafirlukast in the treatment of asthma: interim results of an open-label extension trial.
BACKGROUND: Current guidelines recommend anti-leukotriene agents as alternative treatments for mild persistent asthma; however, information on their long-term safety and efficacy is needed. OBJECTIVE: To evaluate long-term safety and efficacy of and compliance with oral zafirlukast (Z; 20 mg BID) during the first 39 weeks of ongoing, multicenter, open-label extension (OLE) of a previously reported 13-week, randomized (2:1), double-blind (DB), placebo (P)-controlled trial in mild-to-moderate asthmatic patients treated previously with beta2-agonists alone. METHODS: Patients (12 to 76 years; FEV1 > or = 55% predicted) elected to enter OLE after completing the DB trial. Safety evaluated by adverse events (AEs), laboratory tests, and physical and electrocardiographic examinations. Efficacy assessed by spirometry measurements [FEV1, FEV1 % predicted, personal-best (post-bronchodilator) FEV1] and treatment failure rates. Compliance was calculated as percentage of treatment dispensed. After a visit at OLE week 3 (week 16), patients had visits every 12 weeks. RESULTS: A total of 443 patients (nz-->z = 310, np-->z = 133) entered the OLE. Results through the OLE period showed that 80% of patients overall reported AEs. Of patients randomized to Z and P during DB period, 68% and 67%, respectively, reported AEs during quarter I (Q1); percentage of Z-treated patients reporting AEs during the OLE ranged from 66% (Q2) to 44% (Q4). Review by quarters showed occurrence of AEs in Z-treated groups (Q2-4) was similar to that in P group (Q1). Compared with baseline (week 0), modest yet significant improvements (P < or = .02) in all spirometry measurements were noted in Z --> Z and P --> Z groups at OLE week 3, with sustained effects noted during OLE period. Treatment failure rates during OLE ranged from 7.2% (Q2) to 3.3% (Q4). Mean compliance ranged from 98% (OLE week 3) to 95% (OLE week 39). CONCLUSIONS: Long-term treatment with zafirlukast was safe and well tolerated in asthmatic patients. Sustained efficacy and asthma control and good compliance were observed over extended treatment period. Results demonstrate long-term safety and effectiveness of and compliance with this anti-leukotriene agent.
Adolescent↗
Characterisation of side-chain conformational preferences in a biologically active but unfolded protein.
A combination of experimental NMR 3J alpha beta coupling constant measurements and theoretical predictions from a statistical model for a random coil have been used to characterise the conformations of amino acid side-chains in an unfolded fibronectin binding protein. The statistical model uses the distribution of torsion angles in a data base of native folded protein structures to provide a description of the torsion angle populations of each residue in a random coil. For all but three of the residues studied a close agreement is observed between the experimental 3J alpha beta data and the model predictions (correlation coefficient 0.90; RMSD 0.70 Hz). In these cases the populations about the chi 1 torsion angles in the conformational ensemble defining the fibronectin binding protein are well described by those present in the protein data base. For Phe 69, Asp 92 and Asp 105 however significant deviations are observed between the predictions and experimental data. Each of these side-chains is found to be involved in persistent non-random structural features arising from clustering of hydrophobic groups or interactions between charged side-chains. The analysis demonstrates the detailed insight that can be provided into conformationally disordered states by combining experimental and theoretical approaches.
Adhesins, Bacterial↗
Duration of action and hemodynamic properties of mivacurium chloride in dogs anesthetized with halothane.
OBJECTIVE: To describe onset and duration of neuromuscular blockade induced by mivacurium chloride and its associated hemodynamic effects at 3 dosages in healthy dogs. ANIMALS: 7 Labrador Retrievers. PROCEDURE: Anesthesia was induced with thiopental and maintained with halothane in oxygen, and dogs were mechanically ventilated to end-tidal P(CO)2 between 35 and 40 mm Hg. Core temperature, end-tidal P(CO)2, and halothane concentration were kept constant throughout the experiment. Neuromuscular function was assessed by evaluation of the train-of-four response to a supramaximal electrical stimulus of 2 Hz applied to the ulnar nerve every 10 seconds. Blood for determination of plasma cholinesterase activity was obtained prior to administration of mivacurium, a bolus of which was administered IV, using a randomized Latin-square design for dosages of 0.01, 0.02, and 0.05 mg/kg of body weight. RESULTS: All dogs had typical plasma cholinesterase activity. After administration of mivacurium, differences were not evident between groups in heart rate, systolic, mean, or diastolic blood pressure, change at any time in heart rate, systolic, mean, or diastolic blood pressure, or pH. Interval from onset to 100% neuromuscular blockade was 3.92+/-1.70, 2.42+/-0.53, and 1.63+/-0.25 minutes at dosages of 0.01, 0.02, and 0.05 mg/kg, respectively. Duration of measurable neuromuscular blockade was 33.72+/-12.73, 65.38+/-12.82, and 151.0+/-38.50 minutes, respectively. Time of onset and duration of effect differed significantly among dosages. CONCLUSIONS AND CLINICAL RELEVANCE: Mivacurium provides good hemodynamic stability at the dosages tested. In dogs, this drug has a rapid onset and long duration of effect.
Anesthesia↗
Pharmacokinetic variables of mivacurium chloride after intravenous administration in dogs.
OBJECTIVE: To determine pharmacokinetic variables of mivacurium chloride after IV administration in dogs. ANIMALS: 5 healthy Labrador Retrievers. PROCEDURE: Anesthesia was induced with thiopental and maintained with halothane in oxygen. Dogs were ventilated mechanically to an end-tidal P(CO)2 value between 35 and 40 mm Hg. Heart rate, direct blood pressure, and arterial pH were recorded throughout the experiment. Core temperature, end-tidal P(CO)2, and halothane concentration were kept constant throughout the experiment. Paired blood samples for determination of plasma cholinesterase activity were collected prior to administration of a bolus of mivacurium (0.05 mg/kg of body weight), which was administered IV during a 2-second period. Arterial blood samples were obtained for determination of plasma mivacurium concentration 0, 1, 3, 5, 10, 30, 60, 120, 150, and 180 minutes after administration of mivacurium. Blood was collected into tubes containing EDTA and 0.25% echothiophate. Mivacurium concentration was determined, using reversed-phase high-performance liquid chromatography. RESULTS: For the trans-trans isomer, mean +/- SEM volume of distribution was 0.18+/-0.024 L/kg, median half-life was 34.9 minutes (range, 26.7 to 53.5 minutes), and clearance was 12+/-2 ml/min/kg. For the cis-trans isomer, values were 0.31+/-0.05 L/kg, 43.4 minutes (range, 31.5 to 69.3 minutes), and 15+/-2 ml/min/kg, respectively. Values for the cis-cis isomer were not calculated, because it was not detectable in plasma 60 minutes after mivacurium administration in all 5 dogs. CONCLUSIONS AND CLINICAL RELEVANCE: The transtrans and cis-trans isomers of mivacurium have a long half-life and slow clearance in healthy dogs anesthetized with halothane.
Animals↗
Structural and dynamical characterization of a biologically active unfolded fibronectin-binding protein from Staphylococcus aureus.
A 130-residue fragment (D1-D4) taken from a fibronectin-binding protein of Staphylococcus aureus, which contains four fibronectin-binding repeats and is unfolded but biologically active at neutral pH, has been studied extensively by NMR spectroscopy. Using heteronuclear multidimensional techniques, the conformational properties of D1-D4 have been defined at both a global and a local level. Diffusion studies give an average effective radius of 26.2 +/- 0.1 A, approximately 75% larger than that expected for a globular protein of this size. Analysis of chemical shift, 3JHNalpha coupling constant, and NOE data show that the experimental parameters agree well overall with values measured in short model peptides and with predictions from a statistical model for a random coil. Sequences where specific features give deviations from these predictions for a random coil have however been identified. These arise from clustering of hydrophobic side chains and electrostatic interactions between charged groups. 15N relaxation studies demonstrate that local fluctuations of the chain are the dominant motional process that gives rise to relaxation of the 15N nuclei, with a persistence length of approximately 7-10 residues for the segmental motion. The consequences of the structural and dynamical properties of this unfolded protein for its biological role of binding to fibronectin have been considered. It is found that the regions of the sequence involved in binding have a high propensity for populating extended conformations, a feature that would allow a number of both charged and hydrophobic groups to be presented to fibronectin for highly specific binding.
Adhesins, Bacterial↗
Side-chains in native and random coil protein conformations. Analysis of NMR coupling constants and chi1 torsion angle preferences.
The behaviour of amino acid side-chains in proteins in solution has been characterised by analysing NMR 3JHalphaH beta coupling constants and crystallographic chi1 torsion angles. Side-chains both in the core of native folded proteins and in situations where there is an absence of close packing including the random coil state have been considered. An analysis of experimental 3JHalphaH beta coupling constant data for ten proteins shows that in the core of native proteins a very close similarity is observed between the chi1 conformations adopted in solution and in crystals. There is clear evidence, however, for significant motional averaging about the chi1 torsion angles in solution. Using a model of a Gaussian distribution about the average torsion angles the extent of these fluctuations has been quantified; the standard deviation for the motion is 26 degrees, the fluctuations about chi1 in the protein core being similar in size to those found for main-chain phi torsion angles in solution. From the distribution of chi1 torsion angles in a data base of protein crystal structures, torsion angle populations and coupling constants have been predicted for a random coil polypeptide. Significant variations in the chi1 distributions for different amino acids give differences in the predicted coupling constants; for 3JHalphaH beta, for example, values of 5.1 and 5.7 Hz are predicted for serine compared with 4.9 and 9.9 Hz for leucine. Experimental data for short unstructured peptides show an excellent agreement with the predictions, indicating that the overall chi1 distributions in protein crystals reflect the local preferences of the amino acids. Predictions from the protein data base therefore provide an important framework for interpreting experimental data for non-native protein conformations and for residues on the surface of folded proteins.
Crystallography↗
Molecular dynamics simulations of peptide fragments from hen lysozyme: insight into non-native protein conformations.
Molecular dynamics simulations of four peptides taken from the hen lysozyme sequence have been used to generate models for non-native protein conformations. Comparisons between the different peptides and with experimental data for denatured lysozyme and peptide fragments provides insight into the characteristics of the conformational ensembles populated in these non-native states and the dependence of their structural features on the amino acid sequence. For the denatured conformers populated local contacts dominate in determining the properties observed in the trajectories, all four peptides showing similar characteristics. These include a significant increase in the number of main-chain O(i)-NH(i+2) hydrogen bonds and hydrogen bonds involving side-chain groups, this increase compensating to a large extent for the loss of hydrogen bonds involved in helical or beta-sheet secondary structure in the native fold, and the generation of a population of collapsed states with local clusterings of hydrophobic groups. The hydrophobic clusters enable at least partial burial of many side-chains exposed by the loss of tertiary contacts on denaturation and provide models that may explain the experimentally observed protection of amides from hydrogen exchange and the existence of residual secondary structure in non-native species of lysozyme. The results suggest that this approach has an important role to play in aiding the interpretation of experimental data for conformationally disordered non-native states of proteins.
Animals↗
A Philadelphia negative chronic myelogenous leukemia with the chimeric BCR/ABL gene on chromosome 9 and a b3-a2 splice junction.
Approximately 5% of patients with chronic myelogenous leukemia (CML) do not reveal the Philadelphia (Ph) chromosome cytogenetically and are termed Ph-negative CML cases. We report one such case, which appeared normal by routine banding techniques. The BCR/ABL rearrangement was detected by reverse transcriptase-polymerase chain reaction and Southern blotting analysis, which suggested a b3-a2 splice junction. Dual color fluorescence in situ hybridization (FISH) with BCR and ABL DNA probes showed that the chimeric fusion gene was localized on chromosome 9q34, rather than at the typical location on chromosome 22q11. The BCR/ABL rearrangement was detected in 75% of the patient's bone-marrow population, whereas the remaining 25% of the cells appeared normal. The use of dual color FISH in the diagnosis of CML is extremely valuable not only in identifying cases of Ph-negative CML, but also in quantifying the proportion of transformed cell populations. This information ultimately results in an enhancement of our ability to monitor therapy, follow disease progression, and determine transplant eligibility.
Blotting, Northern↗
A single dose of zafirlukast reduces LTD4-induced bronchoconstriction in patients on maintenance inhaled corticosteroid therapy.
BACKGROUND: Previous studies demonstrated that leukotriene receptor antagonists (LTRAs) are effective in reducing asthma symptoms and the airway response to inhaled leukotriene D4 (LTD4) in asthmatic patients receiving inhaled beta2-agonists alone. OBJECTIVE: To investigate the efficacy of a single 20-mg dose of the oral LTRA zafirlukast in reducing the airway response to inhaled LTD4 in mild-to-moderate asthmatic patients receiving inhaled beta2-agonists and inhaled corticosteroids (ICS). METHODS: In this double-blind, crossover trial, six patients on maintenance ICS (median dose 800 microg/day; range 336 to 1600 microg/day), who had a 20% decrease in FEV1 following inhalation of a maximal concentration of 50 microg/mL LTD4, received either zafirlukast or placebo on each of two study days. Two hours after dosing, patients underwent bronchoprovocation challenges with increasing concentrations of LTD4 (0.1 to 1000 microg/mL) at 10-minute intervals until either the patient's FEV1 decreased by 20% or the maximum concentration of LTD4 was given. Spirometric tests were done just before (baseline) and throughout the challenge phase until the patient's FEV1 returned to within 5% of baseline. Blood samples were collected two hours after dosing to determine plasma concentrations of zafirlukast. RESULTS: Compared with placebo, zafirlukast produced a 1.82-unit increase in logPC20FEV1 and a 1.88-unit increase in logPD20FEV1, representing a 66-fold higher concentration and a 76-fold higher dose of LTD4, respectively, to produce a 20% decrease in FEV1 (P < .001). Mean time to recovery after challenge was 36.7 versus 51.7 minutes when patients received zafirlukast and placebo, respectively. No correlation between clinical effects and plasma drug levels was observed. CONCLUSIONS: This trial demonstrated that asthmatic patients on maintenance ICS can respond to exogenously administered LTD4 and that zafirlukast reduced the airway response to LTD4 in these patients.
Administration, Inhalation↗
The prospects for long-term intervention in asthma with antileukotrienes.
The antileukotriene drugs are the first new therapeutic agents approved for the treatment of asthma in more than 20 years. The currently available compounds are orally active and either prevent the cysteinyl leukotrienes from binding to and activating the cysLT-1 receptor in the lung (leukotriene receptor antagonists, LTRAs) or inhibit leukotriene synthesis (leukotriene synthesis inhibitors, LTSIs). The antileukotrienes acutely bronchodilate the airways of patients with baseline bronchoconstriction, although usually not as well as beta-agonists. When used regularly they produce rapid improvements of pulmonary function and symptoms in patients with mild-to-moderate asthma, and probably of those with more severe asthma as well, which persists for the duration of treatment. Yet, individual responses to the antileukotrienes are variable and not predictable based on clinical criteria. Recent studies suggest they reduce asthmatic airway inflammation. The few comparator studies with other asthma medications indicate they are equal or more effective than cromolyn but equal or less effective than low-to-moderate doses of inhaled corticosteroids. Initial experience with the antileukotrienes reveals limited toxicity and a favourable therapeutic-to-toxic ratio. The exact role of the antileukotrienes in the treatment of asthma remains to be determined, as does the relative potency of the various agents.
Adult↗
Transfusion-acquired hepatitis C virus infection in a Canadian population, October 1983 through June 1985.
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Adult↗
A risk-benefit assessment of antileukotrienes in asthma.
The antileukotriene drugs are the first new therapeutic agents approved for the treatment of asthma in more than 20 years. The currently available compounds are orally active and either prevent the cysteinyl leukotrienes from binding to and activating the cysLT-1 receptor in the lung (leukotriene receptor antagonists) or inhibit leukotriene synthesis (leukotriene synthesis inhibitors). Studies performed in individuals without asthma and patients with asthma reveal that antileukotrienes prevent the bronchoconstriction produced by exercise, cold-air, allergen, aspirin (acetylsalicylic acid) and sulphur dioxide. Except for the setting of aspirin sensitivity where the antileukotrienes are nearly uniformly effective, individual responses to them are variable with complete protection in some, no protection in others and a modest degree of protection in the majority. The antileukotrienes bronchodilate the airways of patients with baseline bronchoconstriction, although usually not as well as beta-agonists. When given for weeks to months they rapidly improve pulmonary function and symptoms in patients with mild-to-moderate asthma, and probably in patients with more severe asthma as well, and these improvements persist for the duration of treatment. Here too, their beneficial effects are variable and not predictable based on clinical criteria. Recent studies suggest they can reduce asthma-induced airway inflammation and are equal or more effective than sodium cromoglycate, but equal or less effective than low-to-moderate dosages of inhaled corticosteroids. Initial experience with the antileukotrienes reveals limited toxicity and what appears to be a favourable therapeutic-to-toxic ratio. However, exposure of more patients with differing characteristics for longer periods of time is needed to substantiate this initial impression. The exact role of the antileukotrienes in the treatment of asthma remains to be determined, as does the relative potency of the various agents.
Anti-Asthmatic Agents↗
NMR analysis of main-chain conformational preferences in an unfolded fibronectin-binding protein.
A 130-residue fragment of the Staphylococcus aureus fibronectin-binding protein has been found to exist in a highly unfolded conformation at neutral pH. Measurement of experimental NMR 3JHNalpha coupling constants provides evidence for individual residues having distinct main-chain conformational preferences that are dependent both on the amino acid concerned and on neighbouring residues in the sequence. Analysis shows that these variations in the populations of individual residues can be explained in detail in terms of statistical distributions of conformational states derived from the protein data base. In particular, when the preceding residue has a beta-branched or aromatic side-chain, a significant increase occurs in the population of the less sterically restricted b region of phi,psi space. The results indicate that the local structure of the fibronectin binding protein in solution, under conditions where it displays full activity, approximates very closely to a statistical random coil structure. This may be an important feature in the biological role of this and other polypeptides involved in protein-protein interactions.
Adhesins, Bacterial↗
Modulation of c-fos expression in the rat striatum by diazepam.
The benzodiazepine agonist, diazepam, inhibits cAMP production in the rat brain. Since cAMP influences c-fos activity, we examined the effects of diazepam on expression of this immediate early gene, as indicated by Western blot analysis. Intraperitoneal administration of diazepam increased Fos protein levels in the striatum, but not in the hippocampus. In contrast, pretreatment with diazepam blocked the potent inducing effect of amphetamine, in both brain regions. Similar induction and blockade effects were also observed for a 90 kDa Fos related antigen (Fra), in the striatum and hippocampus. The possible mechanisms underlying the modulatory effects of diazepam on c-fos expression in the brain are discussed.
Amphetamine↗
Fat absorption after small intestinal transplantation in the rat.
BACKGROUND: Intestinal transplantation is now used for patients with severe malabsorption, however, little data exists quantifying the ability of the graft to absorb fat. This study tested the hypothesis that intestinal transplantation would not affect the lymphatic or venous uptake of fatty acids. METHODS: A syngeneic rat model of intestinal transplantation (SIT) with caval drainage of the graft was used. Control animals underwent intestinal division and reanastomosis (n=15 in each group). The animals were followed for 6 weeks, and fat absorption in vivo was quantified. The animals were anesthetized, sampling catheters were placed in the jugular and superior mesenteric veins and in the mesenteric lymphatic duct, and a feeding tube was passed into the duodenum. Animals were allowed to recover, and a steady-state duodenal infusion of lauric (C12:0) and palmitic (C16:0) fatty acid emulsion was begun. A radiolabeled pulse of lauric (C12:0) and palmitic (C16:0) fatty acid was then given, and the subsequent appearance in the lymphatic and venous systems was quantified. RESULTS: In vivo absorption of dietary fat was preserved, but after transplantation the mesenteric lymphatic flow and cumulative lymphatic appearance of both labels was significantly reduced (flow reduced from 4.8+/-1.1 in controls to 1.0+/-0.29 ml/hr in transplant animals, whereas lauric acid absorption was 33+/-11.4% in controls vs. 7.5+/-2.5% in transplant animals). There was a modest increase in the jugular venous appearance of the fatty acids (2.0+/-1.1% in transplant animals vs. 0.75+/-0.55% in controls for lauric acid; P<0.05 for all comparisons). Absorption of lauric and palmitic acids was very similar, and there was no preferential absorption detected in the portal venous system. Dye studies demonstrated lymphatic recannulization around the vascular anastomosis, into the retroperitoneum. CONCLUSIONS: These results suggest that in this model of SIT, fat absorption via the mesenteric duct is reduced, but that compensatory collaterals form into the retroperitoneal lymphatics. There was no evidence of any significant increase in portal venous uptake of fatty acids after SIT, nor of preferential absorption of medium-chain fatty acids. These results may have implications for patients after SIT.
Animals↗