Elsevier

Drug Discovery Today

Volume 7, Issue 4, 15 February 2002, Pages 235-246
Drug Discovery Today

Review
Target validation of G-protein coupled receptors

https://doi.org/10.1016/S1359-6446(01)02131-6Get rights and content

Abstract

G-protein coupled receptors (GPCRs) represent possibly the most important target class of proteins for drug discovery. Over 30% of clinically marketed drugs are active at this receptor family. These drugs exhibit their activity at <10% of all known GPCRs. A major challenge for the pharmaceutical industry is to associate the many novel GPCRs with disease to identify the drugs of the future. This process consists of a collection of experimental paradigms that together can be loosely labelled ‘target validation’.

Section snippets

In silico target validation

GPCRs can be classified into three major families according to sequence homology. Family A represents the largest subgroup of receptors and includes catecholamine, neuropeptide, chemokine, glycoprotein, lipid and nucleotide receptors. Family B contains receptors for a large number of peptides such as calcitonin-gene-related peptide (CGRP) and calcitonin, and family C is the metabotropic family containing the metabotropic glutamate receptors (mGluRs), γ-amino butyric acid (GABAB) receptors and

Biological target validation

Target validation occurs throughout the process of drug discovery and there are many levels of experiments that can add weight to a target's ability to modify disease. A truly validated drug target is one that improves disease outcome when it is modulated in humans. However, this level of validation is expensive and time consuming and confidence can generally be gained in a target well before clinical experiments are performed. Evidence of a target's involvement in disease is gathered from a

Expression profiling

Expression profiling of GPCRs is one of the earliest steps in target validation. The goal of such studies is to identify the tissues and cell types in which the receptor is expressed and to determine if the pattern of expression is altered in disease. Put simply, if a receptor is only expressed in the brain it might be a target in CNS disorders but is unlikely to be a target in respiratory disease. The analysis of the pattern of receptor mRNA expression can be performed by northern blotting or

Reverse pharmacology

One direct means of revealing the physiological function of an orphan GPCR is to identify either its endogenous ‘natural ligand’ or a synthetic, surrogate ligand. This ligand–receptor pairing can then be used to investigate the G-protein coupling and effector-regulating specificities of the receptor in question, and mutagenesis studies can be employed to identify key amino acid residues within the receptor that are involved in ligand binding. Such functional information and tissue distribution

Commonly used methods for the identification of ligands at orphan GPCRs

The choice of assay is of paramount importance for the success of a ligand fishing strategy. For an orphan receptor the G-protein signalling pathway is often unknown. Thus to maximize the chance of success the assay system must be as generic as possible to allow for the detection of a wide range of signalling mechanisms, but also be amenable to HTS such that the activity of a large number of ligands can be readily measured. Such assay systems in mammalian cells rely mainly on measuring changes

Successful identification of ligands at orphan GPCRs

Once receptor expression has been achieved in a suitable system, screening for ligands can commence. The ability to marry orphan receptors with their endogenous ligand partner arguably provides the most powerful means of validating the role of a receptor in the pathophysiology of disease. The number of ligand–orphan GPCR pairings have increased exponentially over the past two years. Indeed, there are often multiple publications describing such pairings [such as Histamine H4 and

Characterization of liganded GPCRs

Several approaches have been adopted to derive evidence that a particular receptor–ligand combination is involved in disease pathology. In the following paragraphs we discuss the application of five complementary approaches that have been used to validate GPCRs as drug targets in animal models of disease: (1) studies of the effects of the natural ligand; (2) studies of the effects of synthetic antagonists; (3) the use of antagonist antibodies; (4) the application of antisense DNA technology and

Closing remarks

In recent times, genome mining has led to the identification of a large number of novel GPCRs. Such receptors are likely to respond to ligands from a broad spectrum of chemical classes, ranging from small molecules to large peptides. All of these orphan GPCRs are potential therapeutic targets. Based upon the historical success of GPCR drugs, the pharmaceutical industry is investing billions of dollars in the race to validate orphan GPCRs as drug targets. As outlined in this review, the

Acknowledgements

The authors would like to express their thanks to Tony Arleth, Diane Cousens, Mike Pedrick, Mick Hurle, Steve Foord, Darren Moore, Paul Murdock and Phil Szekeres, who have provided data presented in this review. We would also like to thank the many members of the Gene Expression and Protein Biochemistry, Systems Research and Screening Sciences departments across GlaxoSmithKline for discussions over many years that have resulted in the generation of the target validation strategies outlined here.

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