The use of plasma aldosterone and urinary sodium to potassium ratio as translatable quantitative biomarkers of mineralocorticoid receptor antagonism

<p>Abstract</p> <p>Background</p> <p>Accumulating evidence supports the role of the mineralocorticoid receptor (MR) in the pathogenesis of diabetic nephropathy. These findings have generated renewed interest in novel MR antagonists with improved selectivity against other nuclear hormone receptors and a potentially reduced risk of hyperkalemia. Characterization of novel MR antagonists warrants establishing translatable biomarkers of activity at the MR receptor. We assessed the translatability of urinary sodium to potassium ratio (Na⁺/K⁺) and plasma aldosterone as biomarkers of MR antagonism using eplerenone (Inspra^®), a commercially available MR antagonist. Further we utilized these biomarkers to demonstrate antagonism of MR by PF-03882845, a novel compound.</p> <p>Methods</p> <p>The effect of eplerenone and PF-03882845 on urinary Na⁺/K⁺and plasma aldosterone were characterized in Sprague-Dawley rats and spontaneously hypertensive rats (SHR). Additionally, the effect of eplerenone on these biomarkers was determined in healthy volunteers. Drug exposure-response data were modeled to evaluate the translatability of these biomarkers from rats to humans.</p> <p>Results</p> <p>In Sprague-Dawley rats, eplerenone elicited a rapid effect on urinary Na⁺/K⁺yielding an EC₅₀that was within 5-fold of the functional <it>in vitro </it>IC₅₀. More importantly, the effect of eplerenone on urinary Na⁺/K⁺in healthy volunteers yielded an EC₅₀that was within 2-fold of the EC₅₀generated in Sprague-Dawley rats. Similarly, the potency of PF-03882845 in elevating urinary Na⁺/K⁺in Sprague-Dawley rats was within 3-fold of its <it>in vitro </it>functional potency. The effect of MR antagonism on urinary Na⁺/K⁺was not sustained chronically; thus we studied the effect of the compounds on plasma aldosterone following chronic dosing in SHR. Modeling of drug exposure-response data for both eplerenone and PF-03882845 yielded EC₅₀values that were within 2-fold of that estimated from modeling of drug exposure with changes in urinary sodium and potassium excretion. Importantly, similar unbound concentrations of eplerenone in humans and SHR rats yielded the same magnitude of elevations in aldosterone, indicating a good translatability from rat to human.</p> <p>Conclusions</p> <p>Urinary Na⁺/K⁺and plasma aldosterone appear to be translatable biomarkers of MR antagonism following administration of single or multiple doses of compound, respectively.</p> <p>Trial Registration</p> <p>For clinical study reference EE3-96-02-004, this study was completed in 1996 and falls out scope for disclosure requirements.</p> <p>Clinical study reference A6141115: <url>http://clinicaltrials.gov</url>, <url>http://NIHclinicaltrails.gov</url>; NCTID: <a href="http://www.clinicaltrials.gov/ct2/show/NCT00990223">NCT00990223</a></p

Truncated glucagon-like peptide-1 and exendin-4 α-conotoxin pl14a peptide chimeras maintain potency and α-helicity and reveal interactions vital for cAMP signaling in vitro

Glucagon-like peptide-1 (GE P-1) signaling through the glucagon-like peptide 1 receptor (GLP-1R) is a key regulator of normal glucose metabolism, and exogenous GLP-1R agonist therapy is a promising avenue for the treatment of type 2 diabetes mellitus. To date, the development of therapeutic GLP-1R agonists has focused on producing drugs with an extended serum half-life. This has been achieved by engineering synthetic analogs of GLP-1 or the more stable exogenous GLP-1R agonist exendin-4 (Ex-4). These synthetic peptide hormones share the overall structure of GLP-1 and Ex-4, with a C-terminal helical segment and a flexible N-terminal tail. Although numerous studies have investigated the molecular determinants underpinning GLP-1 and Ex-4 binding and signaling through the GLP1R, these have primarily focused on the length and composition of the N-terminal tail or on how to modulate the helicity of the full-length peptides. Here, we investigate the effect of C-terminal truncation in GLP-1 and Ex-4 on the cAMP pathway. To ensure helical C-terminal regions in the truncated peptides, we produced a series of chimeric peptides combining the N-terminal portion of GLP-1 or Ex-4 and the C-terminal segment of the helix-promoting peptide alpha-conotoxin p114a. The helicity and structures of the chimeric peptides were confirmed using circular dichroism and NMR, respectively. We found no direct correlation between the fractional helicity and potency in signaling via the cAMP pathway. Rather, the most important feature for efficient receptor binding and signaling was the C-terminal helical segment (residues 22-27) directing the binding of Phe' into a hydrophobic pocket on the GLP-1R

Pengaruh Harga Terhadap Peningkatan Penjualan Produk Semen Tiga Roda Pada PT. Robcaga Beo Kabupaten Kepulauan Talaud

The development in business world these days is market by the competition between the business company is getting fierce. Especially in managing the company business unit. It is shown by the appearance of a company that offer a good quality product with a compete price on the market. To handle the fierce competition on the market then one from so many effort that the company do is by apply the strategic price. Which on the way of applying that strategi the company try to set a price that can be compete in the market so the increase sale of the product become maximum. With right price and controlled will result the domino effect to a company to build long term relationship with costumer so it can increase the sales volume. This research is a descriptive quantitative research by using the correlation approach and simple regression. To see relation between variable and to measure the impact to the variable itself. So the purpose of this research is to know how far the price effect and to the increase of PT. ROBCAGA in Talaud. According to the sesult of the research, can be shown as following: price has a correlation and significant determination effort to the increase sale of PT. ROBCAGA Talaud. According to the data analysis, coefficient value moment r = 0,685. That show there is a positive relation, and can be categorize as high and strong, also price coefficient determination to the increase sale is by 46,5% and 53,5% by the rest of it depends on the unknown factors that not been analyze in this research

Type I interferon regulation by USP18 is a key vulnerability in cancer

Precise regulation of Type I interferon signaling is crucial for combating infection and cancer while avoiding autoimmunity. Type I interferon signaling is negatively regulated by USP18. USP18 cleaves ISG15, an interferon-induced ubiquitin-like modification, via its canonical catalytic function, and inhibits Type I interferon receptor activity through its scaffold role. USP18 loss-of-function dramatically impacts immune regulation, pathogen susceptibility, and tumor growth. However, prior studies have reached conflicting conclusions regarding the relative importance of catalytic versus scaffold function. Here, we develop biochemical and cellular methods to systematically define the physiological role of USP18. By comparing a patient-derived mutation impairing scaffold function (I60N) to a mutation disrupting catalytic activity (C64S), we demonstrate that scaffold function is critical for cancer cell vulnerability to Type I interferon. Surprisingly, we discovered that human USP18 exhibits minimal catalytic activity, in stark contrast to mouse USP18. These findings resolve human USP18's mechanism-of-action and enable USP18-targeted therapeutics

Cyclic alpha-conotoxin peptidomimetic chimeras as potent GLP-1R agonists

Type 2 diabetes mellitus (T2DM) results from compromised pancreatic beta-cell function, reduced insulin production, and lowered insulin sensitivity in target organs resulting in hyperglycemia. The GLP-1 hormone has two biologically active forms, GLP-1-(7-37) and GLP-1-(7-36)amide, which are equipotent at the glucagon-like peptide-1 receptor (GLP-1R). These peptides are central both to normal glucose metabolism and dysregulation in T2DM. Several structurally modified GLP-1 analogues are now approved drugs, and a number of other analogues are in clinical trials. None of these compounds is orally bioavailable and all require parenteral delivery. Recently, a number of smaller peptidomimetics containing 11-12 natural and unnatural amino acids have been identified that have similar insulin regulating profiles as GLP-1. The alpha-conotoxins are a class of disulfide rich peptide venoms isolated from cone snails, and are known for their highly constrained structures and resistance to enzymatic degradation. In this study, we examined whether 11-residue peptidomimetics incorporated into alpha-conotoxin scaffolds, forming monocyclic or bicyclic compounds constrained by disulfide bonds and/or backbone cyclization, could activate the GLP-1 receptor (GLP-1R). Several compounds showed potent (nanomolar) agonist activity at GLP-1R, as evaluated via cAMP signaling. In addition, HPLC retention times and in silica calculations suggested that mono- and bicyclic compounds had more favorable n-octanol/water partition coefficients according to the virtual partition coefficient model (vLogP), while maintaining a smaller radius of gyration compared to corresponding uncyclized peptidomimetics. Our findings suggest that cyclic peptidomimetics provide a potential avenue for future design of potent, compact ligands targeting GLP-1R and possessing improved physicochemical properties. (C) 2015 Elsevier Masson SAS. All rights reserved

Helixconstraints and amino acid substitution in GLP-1 increase cAMP and insulin secretion but not beta-arrestin 2 signaling

Glucagon-like peptide (GLP-1) is an endogenous hormone that induces insulin secretion from pancreatic islets and modified forms are used to treat diabetes mellitus type 2. Understanding how GLP-1 interacts with its receptor (GLP-1R) can potentially lead to more effective drugs. Modeling and NMR studies of the N-terminus of GLP-1 suggest a β-turn between residues Glu9-Phe12 and a kinked alpha helix between Val16-Gly37. N-terminal turn constraints attenuated binding affinity and activity (compounds 1–8). Lys-Asp (i, i+4) crosslinks in the middle and at the C-terminus increased alpha helicity and cAMP stimulation without much effect on binding affinity or beta-arrestin 2 recruitment (compounds 9–18). Strategic positioning of helix-inducing constraints and amino acid substitutions (Tyr16, Ala22) increased peptide helicity and produced ten-fold higher cAMP potency (compounds 19–28) over GLP-1(7–37)-NH. The most potent cAMP activator (compound 23) was also the most potent inducer of insulin secretion

Ethnicity and Cutaneous Melanoma in the City of Sao Paulo, Brazil: A Case-Control Study

Background: Over the last century the incidence of cutaneous melanoma has increased worldwide, a trend that has also been observed in Brazil. The identified risk factors for melanoma include the pattern of sun exposure, family history, and certain phenotypic features. In addition, the incidence of melanoma might be influenced by ethnicity. Like many countries, Brazil has high immigration rates and consequently a heterogenous population. However, Brazil is unique among such countries in that the ethnic heterogeneity of its population is primarily attributable to admixture. This study aimed to evaluate the contribution of European ethnicity to the risk of cutaneous melanoma in Brazil. Methodology/Principal Findings: We carried out a hospital-based case-control study in the metropolitan area of Sao Paulo, Brazil. We evaluated 424 hospitalized patients (202 melanoma patients and 222 control patients) regarding phenotypic features, sun exposure, and number of grandparents born in Europe. Through multivariate logistic regression analysis, we found the following variables to be independently associated with melanoma: grandparents born in Europe-Spain (OR = 3.01, 95% CI: 1.03-8.77), Italy (OR = 3.47, 95% CI: 1.41-8.57), a Germanic/Slavic country (OR = 3.06, 95% CI: 1.05-8.93), or &gt;= 2 European countries (OR = 2.82, 95% CI: 1.06-7.47); eye color-light brown (OR = 1.99, 95% CI: 1.14-3.84) and green/blue (OR = 4.62; 95% CI 2.22-9.58); pigmented lesion removal (OR = 3.78; 95% CI: 2.21-6.49); no lifetime sunscreen use (OR = 3.08; 95% CI: 1.03-9.22); and lifetime severe sunburn (OR = 1.81; 95% CI: 1.03-3.19). Conclusions: Our results indicate that European ancestry is a risk factor for cutaneous melanoma. Such risk appears to be related not only to skin type, eye color, and tanning capacity but also to others specific characteristics of European populations introduced in the New World by European immigrants.Fundacao de Amparo a Pesquisa do Estado de Sao Paulo - FAPESP [06-52041-9, 5-56069-2]Fundacao de Amparo a Pesquisa do Estado de Sao Paulo (FAPESP)Conselho Nacional de Desenvolvimento Cientifico e Tecnologico - Brasil (National Counsel of Technological and Scientific Development - Brazil) - CNPq [478239/03-3]Conselho Nacional de Desenvolvimento Cientifico e Tecnologico Brasil (National Counsel of Technological and Scientific Development Brazil) CNP

By Any Other Name: Heterologous Replacement of the Escherichia coli RNase P Protein Subunit Has In Vivo Fitness Consequences

Bacterial RNase P is an essential ribonucleoprotein composed of a catalytic RNA component (encoded by the rnpB gene) and an associated protein moiety (encoded by rnpA). We construct a system that allows for the deletion of the essential endogenous rnpA copy and for its simultaneous replacement by a heterologous version of the gene. Using growth rate as a proxy, we explore the effects on fitness of heterologous replacement by increasingly divergent versions of the RNase P protein. All of the heterologs tested complement the loss of the endogenous rnpA gene, suggesting that all existing bacterial versions of the rnpA sequence retain the elements required for functional interaction with the RNase P RNA. All replacements, however, exact a cost on organismal fitness, and particularly on the rate of growth acceleration, defined as the time required to reach maximal growth rate. Our data suggest that the similarity of the heterolog to the endogenous version — whether defined at the sequence, structure or codon usage level — does not predict the fitness costs of the replacement. The common assumption that sequence similarity predicts functional similarity requires experimental confirmation and may prove to be an oversimplification