Estimating prevalence of osteoporosis: examples from industrialized countries.

UNLABELLED: In nine industrialized countries in North America, Europe, Japan, and Australia, country-specific osteoporosis prevalence (estimated from published data) at the total hip or hip/spine ranged from 9 to 38 % for women and 1 to 8 % for men. In these countries, osteoporosis affects up to 49 million individuals. PURPOSE: Standardized country-specific prevalence estimates are scarce, limiting our ability to anticipate the potential global impact of osteoporosis. This study estimated the prevalence of osteoporosis in several industrialized countries (USA, Canada, five European countries, Australia, and Japan) using the World Health Organization (WHO) bone mineral density (BMD)-based definition of osteoporosis: BMD T-score assessed by dual-energy x-ray absorptiometry ≤-2.5. METHODS: Osteoporosis prevalence was estimated for males and females aged 50 years and above using total hip BMD and then either total hip or spine BMD. We compiled published location-specific data, using the National Health and Nutrition Examination Survey (NHANES) III age and BMD reference groups, and adjusted for differences in disease definitions across sources. Relevant NHANES III ratios (e.g., male to female osteoporosis at the total hip) were applied where data were missing for countries outside the USA. Data were extrapolated from geographically similar countries as needed. Population counts for 2010 were used to estimate the number of individuals with osteoporosis in each country. RESULTS: For females, osteoporosis prevalence ranged from 9 % (UK) to 15 % (France and Germany) based on total hip BMD and from 16 % (USA) to 38 % (Japan) when spine BMD data were included. For males, prevalence ranged from 1 % (UK) to 4 % (Japan) based on total hip BMD and from 3 % (Canada) to 8 % (France, Germany, Italy, and Spain) when spine BMD data were included. CONCLUSIONS: Up to 49 million individuals met the WHO osteoporosis criteria in a number of industrialized countries in North America, Europe, Japan, and Australia.

Sex- and age-specific incidence of non-traumatic fractures in selected industrialized countries.

UNLABELLED: Various methodological approaches have estimated the incidence of osteoporosis-related fractures, making comparisons difficult. This study estimated the incidence rates of non-traumatic fractures in 12 countries using standard definitions. Applying these rates to the 2010 population figures of these countries, a total of 5.2 million non-traumatic fractures were estimated, mostly in women. PURPOSE: The purpose of this study was to estimate annual country-, sex-, and age-specific incidence of non-traumatic hip, vertebral, and other fractures for women aged ≥50 and men ≥60 years and the number of fractures expected in 12 countries based on these incidence rates. METHODS: Electronically indexed medical literature and relevant web sites were reviewed to identify studies reporting age- and sex-specific fracture incidence rates to obtain estimates of the proportion of fractures considered to be non-traumatic and to gather relevant census data. From these data, we extrapolated to estimate the number of fractures in 12 countries in North America, Europe, Japan, and Australia. RESULTS: Annual non-traumatic hip fracture incidence rates were highest for women in Sweden, Denmark, and Finland. In women, vertebral fractures were more common than hip fractures. The incidence of vertebral fractures was highest among Scandinavian and Canadian women. In men, Scandinavians had the highest incidence of hip fractures, while Australian men had the highest incidence of vertebral fractures. Hip and vertebral fracture incidence increased steeply with age for both women and men. Age appears to exert less influence on the incidence of fractures at sites other than hip and vertebrae. In 2010, 5.2 million non-traumatic fractures were expected in the 12 countries studied, of which 2.8 million were at the hip or spine. Women accounted for most of the total non-traumatic fracture burden (77 %). CONCLUSIONS: Non-traumatic fractures pose a significant burden, affecting millions of women and men in countries around the world each year.

Muscarinic agonists and antagonists in the treatment of Alzheimer's disease.

Alzheimer's disease (AD) is a neurodegenerative disease characterized by cognitive impairment and personality changes. The development of drugs for the treatment of the cognitive deficits of AD has focused on agents which counteract loss in cholinergic activity. Although symptoms of AD have been successfully treated with acetylcholinesterase inhibitors (tacrine, donepezil. rivastigmine, galanthamine), limited success has been achieved with direct M1 agonists, probably due to their lack of selectivity versus other muscarinic receptor subtypes. Muscarinic M2 antagonists have been reported to increase synaptic levels of acetylcholine after oral administration to rats (e.g. BIBN-99, SCH-57790), but their selectivity versus other muscarinic receptor subtypes is modest. Exploration of a series of piperidinylpiperidines has yielded the potent and selective M2 antagonist SCH-217443. This antagonist has excellent bioavailability in rats and dogs and shows activity in a rat model of cognition.

Facilitation of acetylcholine release and improvement in cognition by a selective M2 muscarinic antagonist, SCH 72788.

Current treatment of Alzheimer's Disease (AD) requires acetylcholinesterase inhibition to increase acetylcholine (ACh) concentrations in the synaptic cleft. Another mechanism by which ACh levels can be increased is blockade of presynaptic M2 muscarinic autoreceptors that regulate ACh release. An antagonist designed for this purpose must be highly selective for M2 receptors to avoid blocking postsynaptic M1 receptors, which mediate the cognitive effects of ACh. Structure-activity studies of substituted methylpiperadines led to the synthesis of 4-[4-[1(S)-[4-[(1,3-benzodioxol-5-yl)sulfonyl]phenyl]ethyl]-3(R)-methyl-1-piperazinyl]-4-methyl-1-(propylsulfonyl)piperidine. This compound, SCH 72788, binds to cloned human M2 receptors expressed in CHO cells with an affinity of 0.5 nM, and its affinity at M1 receptors is 84-fold lower. SCH 72788 is a functional M2 antagonist that competitively inhibits the ability of the agonist oxotremorine-M to inhibit adenylyl cyclase activity. In an in vivo microdialysis paradigm, SCH 72788 increases ACh release from the striatum of conscious rats. The compound is also active in a rodent model of cognition, the young rat passive avoidance response paradigm. The effects of SCH 72788 suggest that M2 receptor antagonists may be useful for treating the cognitive decline observed in AD and other dementias.

Mutagenesis approaches for elucidation of protein structure-function relationships.

Several mutagenesis strategies have been used to allow the identification of specific receptor-ligand interactions to support the docking of ligands within various receptor models. The strengths and limitations of each strategy are discussed in this overview. Large-scale mapping is described using deletion and chimeric receptors. Fine tuning with single residue replacements is also covered along with two-dimensional approaches.

L-750355, a human beta3-adrenoceptor agonist; in vitro pharmacology and profile of activity in vivo in the rhesus monkey.

The profile of in vitro and in vivo biology of a human beta3-adrenoceptor agonist, (S)-N-[4-[2-[[3[(2-amino-5-pyridinyl)oxy]-2-hydroxy-propyl]amino]-eth yl]-phenyl]-4-isopropylbenzenesulfonamide, L-750355, is described. Using cloned human and rhesus beta1-, beta2- and beta3-adrenoceptors, expressed in Chinese hamster ovary (CHO) cells, L-750355 was shown to be a potent, albeit partial, agonist for the human (EC(50)=10 nM; % maximal receptor activation=49%) and rhesus (EC(50)=28 nM; % maximal receptor activation=34%) beta3-adrenoceptors. Furthermore, L-750355 stimulates lipolysis in rhesus adipocytes in vitro. L-750355 is a weak partial agonist (EC(50)=3.2 microM; % maximal receptor activation=33% ) for the human beta1-adrenoceptor but exhibits no agonist activity for rhesus beta1- or beta2-adrenoceptors of either human or rhesus origin. Administration of L-750355 to anesthetized rhesus monkeys, as a series of rising dose intravenous infusions, evokes dose-dependent glycerolemia and tachycardia with no change in mean arterial blood pressure or plasma potassium. The dose-response curve for L-750355-induced glycerolemia lies to the left of that for tachycardia. Propranolol, at a dose (0.3 mg/kg, i.v. ) that attenuates isoproterenol-induced changes in heart rate and glycerolemia, abolished L-750355-induced tachycardia but had no effect on L-750355-induced glycerolemia.

Substituted oxazole benzenesulfonamides as potent human beta3 adrenergic receptor agonists.

As a part of our investigation into the development of orally bioavailable beta3 adrenergic receptor agonists, we have identified a series of substituted oxazole derivatives that are potent beta3 agonists with excellent selectivity against other beta receptors. Several of these compounds showed excellent oral bioavailability in dogs. One example, cyclopentylethyloxazole 5f is a potent beta3 agonist (EC50 = 14 nM, 84% activation) with 340-fold and 160-fold selectivity over beta1 and beta2 receptors, respectively, and has 38% oral bioavailability in dogs.

Discovery of an orally bioavailable alkyl oxadiazole beta3 adrenergic receptor agonist.

5-n-Pentyl oxadiazole substituted benzenesulfonamide 8 is a potent and selective beta3 adrenergic receptor agonist (beta3 EC50 = 23 nM, beta1 IC50 = 3000 nM, beta2 IC50 = 3000 nM). The compound has high oral bioavailability in dogs (62%) and rats (36%) and is among the most orally bioavailable beta3 adrenergic receptor agonists reported to date.

Synthesis and SAR of benzyl and phenoxymethylene oxadiazole benzenesulfonamides as selective beta3 adrenergic receptor agonist antiobesity agents.

Benzyl and phenoxymethylene substituted oxadiazoles are potent and orally bioavailable beta3 adrenergic receptor (AR) agonists. The 4-trifluormethoxy substituted 5-benzyl oxadiazole 5f has an EC50 of 8 nM in the beta3 AR agonist assay with 100-fold selectivity over beta1 and beta2 AR binding inhibition activity. Its oral bioavailability in dogs is 30 +/- 4%, with a half-life of 3.8 +/- 0.4 h. In the anesthetized rhesus, 5f evoked a dose-dependent glycerolemia (ED50Gly = 0.15 mg/kg). Under these conditions a heart rate increase of 15% was observed at a dose level of 10 mg/kg.

Molecular characterization of the melanin-concentrating hormone/receptor complex: identification of critical residues involved in binding and activation.

A molecular model of the human melanin-concentrating hormone (MCH) peptide was constructed and docked into a helical, bacteriorhodopsin-based model of the recently identified human MCH receptor. From this hormone-receptor complex, potential sites of agonist-receptor interaction were identified, and site-directed mutagenesis was used to substitute residues predicted to reside within the receptor binding pocket. Substitution of Asp(123)(3.32) in the third transmembrane domain of the receptor resulted in a loss of detectable (125)I-MCH binding and of MCH-stimulated Ca(2+) flux; cell surface expression of the mutant receptor was not affected. Arg(11) and Arg(14) of the MCH ligand were identified as potential sites of interaction with Asp(123)(3.32). [Ala(14)]-MCH was equipotent to native MCH in its ability to bind to and activate the wild-type MCH receptor, whereas [Ala(11)]-MCH displayed a 3000-fold reduction in binding affinity and a complete loss of measurable functional activity. Furthermore, [Lys(11)]-MCH and [D-Arg(11)]-MCH displayed reduced affinity for the receptor. [Lys(11)]-MCH was observed to be a partial agonist, eliciting approximately 67% of the native peptide's activity in a Ca(2+) flux assay, and [D-Arg(11)]-MCH was determined to be a functional antagonist with a K(b) valve of 15.8 microM. These data provide evidence that a basic moiety with specific stereochemical requirements at this site is needed for receptor activation. We conclude that both Asp(123)(3.32) in the MCH receptor and Arg(11) in the MCH peptide are required for the formation of the MCH peptide/receptor complex and propose that they form a direct interaction that is critical for receptor function.

Activation of the NPY Y5 receptor regulates both feeding and energy expenditure.

Intracerebroventricular (ICV) administration of neuropeptide Y (NPY) has been shown to decrease energy expenditure, induce hypothermia, and stimulate food intake. Recent evidence has suggested that the Y5 receptor may be a significant mediator of NPY-stimulated feeding. The present study attempts to further characterize the role of NPY Y5-receptor subtypes in feeding and energy expenditure regulation. Satiated Long-Evans rats with temperature transponders implanted in the interscapular brown adipose tissue (BAT) displayed a dose-dependent decrease in BAT temperature and an increase in food intake after ICV infusion of NPY. Similar effects were induced by ICV administration of peptide analogs of NPY that activate the Y5 receptor, but not by analogs that activate Y1, Y2, or Y4 receptors. Furthermore, ICV infusion of the Y5 selective agonist D-[Trp(32)]-NPY significantly reduced oxygen consumption and energy expenditure of rats as measured by indirect calorimetry. These data suggest that the NPY Y5-receptor subtype not only mediates the feeding response of NPY but also contributes to brown fat temperature and energy expenditure regulation.

Potent, elective human beta3 adrenergic receptor agonists containing a substituted indoline-5-sulfonamide pharmacophore.

A series of compounds possessing an N-substituted indoline-5-sulfonamide pharmacophore was prepared and evaluated for their human beta3 adrenergic receptor agonist activity. The SAR of a wide range of urea and heterocyclic substituents is discussed. 4-Octyl thiazole compound 8c was the most potent and selective compound in the series, with 2800-fold selectivity over beta1 binding and 1400-fold selectivity over beta2 binding.

L-770,644: a potent and selective human beta3 adrenergic receptor agonist with improved oral bioavailability.

L-770,644 (9c) is a potent and selective agonist of the human beta3 adrenergic receptor (EC50 = 13 nM). It shows good oral bioavailability in both dogs and rats (%F = 27), and is a full agonist for glycerolemia in the rhesus monkey (ED50 = 0.21 mg/kg). Based on its desirable in vitro and in vivo properties, L-770,644 was chosen for further preclinical evaluation.

Human beta3 adrenergic receptor agonists containing cyclic ureidobenzenesulfonamides.

Human beta3 adrenergic receptor agonists containing 5-membered ring ureas were shown to be potent partial agonists with excellent selectivity over beta1 and beta2 binding. L-760,087 (4a) and L-764,646 (5a) (beta3 EC50 = 18 and 14 nM, respectively) stimulate lipolysis in rhesus monkeys (ED50 = 0.2 and 0.1 mg/kg, respectively) with minimal effects on heart rate. Oral absorption in dogs is improved over other urea analogs.

Human beta3 adrenergic receptor agonists containing imidazolidinone and imidazolone benzenesulfonamides.

The cyclopentylpropylimidazolidinone L-766,892 is a potent beta3 AR agonist (EC50 5.7 nM, 64% activation) with 420- and 130-fold selectivity over binding to the beta1 and beta2 ARs, respectively. In anesthetized rhesus monkeys, L-766,892 elicited dose-dependent hyperglycerolemia (ED50 0.1 mg/kg) with minimal effects on heart rate.

SCH 57790: a novel M2 receptor selective antagonist.

As a decrease in cholinergic neurons has been observed in Alzheimer's Disease (AD), therapeutic approaches to AD include inhibition of acetylcholinesterase to increase acetylcholine levels. Evidence suggests that acetylcholine release in the CNS is modulated by negative feedback via presynaptic M2 receptors, blockade of which should provide another means of increasing acetylcholine release. Structure-activity studies of [4-(phenylsulfonyl)phenyl]methylpiperazines led to the synthesis of 4-cyclohexyl-alpha-[4-[[4-methoxyphenyl]sulfinyl]-phenyl]-1-piperazin eacetonitrile. This compound, SCH 57790, binds to cloned human M2 receptors expressed in CHO cells with an affinity of 2.78 nM; the affinity at M1 receptors is 40-fold lower. SCH 57790 is an antagonist at M2 receptors expressed in CHO cells, as the compound blocks the inhibition of adenylyl cyclase activity mediated by the muscarinic agonist oxotremorine. This compound should be useful in assessing the potential of M2 receptor blockade for enhancement of cognition.

Presenilins interact with armadillo proteins including neural-specific plakophilin-related protein and beta-catenin.

Missense substitutions in the presenilin 1 (PS1) and presenilin 2 (PS2) proteins are associated with early-onset familial Alzheimer's disease. We have used yeast-two-hybrid and coimmunoprecipitation methods to show that the large cytoplasmic loop domains of PS1 and PS2 interact specifically with three members of the armadillo protein family, including beta-catenin, p0071, and a novel neuronal-specific armadillo protein--neural plakophilin-related armadillo protein (NPRAP). The PS1:NPRAP interaction occurs between the arm repeats of NPRAP and residues 372-399 at the C-terminal end of the large cytoplasmic loop of PS1. The latter residues contain a single arm-like domain and are highly conserved in the presenilins, suggesting that they form a functional armadillo protein binding site for the presenilins.

The GalR2 galanin receptor mediates galanin-induced jejunal contraction, but not feeding behavior, in the rat: differentiation of central and peripheral effects of receptor subtype activation.

The neuropeptide galanin mediates a diverse array of physiological functions through activation of specific receptors. Roles of the three recently cloned galanin receptors (GalRs) in rat intestinal contraction and food intake were examined using GalR-selective ligands and the results were compared with the pharmacological profiles of defined GalRs. The action profile of these ligands in jejunal contraction resembled only that of GalR2 and only a high level of GalR2 mRNA was detected in the tissue, supporting GalR2 as the receptor mediating jejunal contraction. The action profile for food intake in rats excluded GalR2, GalR3 and the putative pituitary galanin receptor as the 'feeding receptor', suggesting that either GalR1 or an unidentified GalR is responsible for mediating this function.

Evidence for hydrophobic interaction between galanin and the GalR1 galanin receptor and GalR1-mediated ligand internalization: fluorescent probing with a fluorescein-galanin.

Galanin is a neuropeptide that activates specific receptors to modulate several physiological functions including food intake, nociception, and learning and memory. The molecular nature of the interaction between galanin and its receptors and the fate of the galanin/receptor complex after the binding event are not understood. A fluorescein-N-galanin (F-Gal) was generated to measure the interaction between galanin and rat GalR1 galanin receptor (rGalR1) and rGalR1-mediated ligand internalization using flow cytometry in transfected Chinese hamster ovary (CHO) cells. Like galanin, F-Gal bound rGalR1 with high affinity and stimulated intracellular signaling events. Fluorescence quenching by soluble KI of rGalR1-bound F-Gal revealed a highly protected environment around the fluorescein, suggesting that the N-terminal portion of galanin, which constitutes the binding site of galanin for the receptor, binds to a protected hydrophobic binding pocket within the receptor. Exposure to F-Gal stimulated rapid (t1/2 approximately 10 min) and extensive (78%) internalization of surface F-Gal into rGalR1/CHO cells at 37 degreesC but not at 0 degreesC. In addition, the internalization did not occur in parental CHO cells at either 0 or 37 degreesC and was inhibited by addition of 0.25 M sucrose in the medium, indicating a GalR1-mediated energy-requiring endocytic process. These results revealed a hydrophobic interaction between galanin and the GalR1 receptor, which is in contrast to those of other G protein-coupled receptors that mainly require hydrophilic interaction with their peptide ligands near or outside the plasma membrane surface, and illustrated that the initial binding interaction is followed by rapid cellular internalization of the agonist/GalR1 complex.

Differential effects of intracerebroventricular glucagon-like peptide-1 on feeding and energy expenditure regulation.

Intracerebroventricular (i.c.v.) administration of glucagon-like peptide-1-(7-37) amide (GLP-1) has been shown to modulate food and water intake. The present studies further characterize the effects of i.c.v. GLP-1 in the regulation of energy balance in lean and obese animals. In both obese and lean Zucker rats, a single i.c.v. infusion of GLP-1 (1-30 microg) resulted in a dose-dependent reduction of food intake and decrease in respiratory quotient relative to the saline control during the first 2 h of the nocturnal cycle. In obese Zucker rats, the food intake was reduced by 73 +/- 11% of the control at the 30 microg dose, whereas a modest 45 +/- 18% reduction was observed in lean rats. Despite the large reduction in food intake seen with GLP-1, there was no compensatory decrease in nocturnal oxygen consumption in the obese Zucker rats. Interestingly, low doses of GLP-1 (1 microg) in lean Zucker rats, which had minimal effects on food intake, caused a 19 +/- 7% increase in O2 consumption during the first 2 h of the nocturnal cycle. These data suggest that central GLP-1 may be an important factor controlling negative energy balance in both the lean and obese Zucker rats.