|
|
|
|
|
|
|
|
|
|
|
 | Signet home |  | news | | people | | meetings |  |
Registration
|
|
||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
TBC.
Plasma protein binding is an important consideration in the development of new drugs. Current methods for determining the degree of protein binding utilise LC-MS to directly measure the free concentration of drug in the presence of serum albumin. Typically only a single concentration of drug is used and the result is returned as percentage bound. For poorly bound compounds this is a sufficient ranking tool, but as compounds approach 98 % bound it ceases to be meaningful. For such classes it would be useful to quantify the equilibrium dissociation constant (Kd) of the drug for serum albumin, in the same way as is reported for receptors. This is often not possible for highly bound drugs using LC-MS due to limits of quantification.
Radioligand binding and high throughput functional assays are common, robust techniques that are widely employed in the drug discovery process. We have explored the potential of utilising these assays to estimate the Kd of drugs at serum albumin. Initially we generated mathematical models that allowed us to simulate the effects of introducing plasma protein on both receptor binding and signalling, to assist in experimental design. We then used these equations to directly fit experimental data (e.g. families of curves in the absence and presence of increasing concentrations of protein) to estimate the affinity of drug at plasma protein. This talk will outline our experiences to date and discuss the potential future application of this method.
TBC.
In the seventies Ariëns, one of pharmacology's godfathers, described a receptor as a beautiful lady whom he wrote letters to, got an answer now and then, accepted she never showed up, but hoping she would one day. Recently, after years of trial-and-error, a number of GPCR crystal structures has been determined: no ladies, but of an intriguing beauty for sure. We were involved in the structure elucidation of the human A2A adenosine receptor in complex with a high affinity antagonist, ZM241385, which was determined to 2.6 Å resolution. Four disulfide bridges in the extracellular domain combined with a subtle repacking of the transmembrane helices relative to the beta-adrenergic and rhodopsin receptor structures define a pocket capable of binding the antagonist in an extended conformation perpendicular to the membrane plane. The binding site highlights an integral role for the extracellular loops together with the helical core in ligand recognition by this class of GPCRs. These findings question the use of rhodopsin or any other single receptor structure in homology modeling studies. On the other hand, we learned that virtual screening of an individual receptor structure (the A2A receptor in our case) can indeed help in finding novel ligands for that particular receptor.
Measuring relative efficacies of agonists for different signalling outputs Many people working in the biomedical sciences are unclear as to the meaning of the term “efficacy” in relation to drug action. This is significant because it can influence the conclusions people arrive at with regard to the mechanism of action of drugs. For example many people conclude that if two agonists produce the same maximum response then they have the same efficacy, whereas if quantified their actual efficacies could in fact be very different. In addition, we can correlate the relative efficacies of agonists at a receptor for more than one signaling output to determine whether the two measures correlate. This is a useful way to detect whether or not biased agonism (also called ligand-directed signaling or functional selectivity) is present. In this talk I will illustrate these ideas by using our recent data measuring the efficacy of a series of ligands at MOPr to activate G protein and recruit arrestin.
TBC.
TBC.
TBC.
TBC.
Lloyd Bridge
Stephen Hill
John King
Lauren May
Markus Owen
Maintained by Markus Owen | © Signet - Cell Signalling Network 2008 |
Icons from FamFamFam Silk
