Analytical pharmacology and allosterism: the importance of quantifying drug parameters in drug discovery

doi:10.1016/j.ddtec.2012.07.006

Drug Discovery Today: Technologies

Volume 10, Issue 2, Summer 2013, Pages e229-e235

https://doi.org/10.1016/j.ddtec.2012.07.006 Get rights and content

Allosteric ligands bind to receptors at sites separate from those binding endogenous ligands; this allows for a wide range of effects from antagonism to potentiation to direct agonism. This paper discusses techniques to quantify complex allosteric behaviors that yield parameters to characterize direct effects (τ_B, the efficacy of allosteric agonists), α and β (the effect of the allosteric ligand on endogenous ligand affinity and efficacy respectively). These parameters are independent of the system used to determine them and thus can be used to predict allosteric effects in all systems.

Graphical abstract

Introduction

The discipline of ‘Analytical Pharmacology’ was defined by James Black as ‘… attempts to interpret ligand interactions with complex biosystems as exposed in different types of bioassay …’ [1, 2]. The benefit of applying analytical pharmacological techniques to drug discovery is that scales of activity can be derived that yield values that are independent of the system in which they are obtained. These then can be used to predict activity in all systems including therapeutic ones. This paper will discuss the application of these approaches to a complex class of ligands that bind to receptors allosterically to modify receptor behavior both directly and in response to interactions with endogenous agonists.

Section snippets

Allosteric modulation

On a molecular level there are two ways that a molecule can interact with seven transmembrane receptors (7TMRs). The first is where the ligand competes with the natural endogenous ligand (i.e. hormone, neurotransmitter) for a binding site on the receptor protein; this will be termed orthosteric interaction. The main feature of this type of mechanism is that there is never a receptor protein species with both the endogenous and second ligand binding at the same instant. From this standpoint,

Allosteric mechanisms lead to unique drug behaviors

There are two main features of allosteric drug action that give rise to unique ligand behaviors: discrete geography of binding and the modification of receptor protein conformation. As a preface to a discussion of how this mechanism results in complex drug behavior it is worth considering the model for allosteric binding [4, 5].where A is the endogenous agonist, B the allosteric ligand, K_A and K_B refer to the equilibrium dissociation constants for the receptor complexes with ligands A and B and

Modeling functional allosteric effects

In view of the notion that an allosteric ligand can stabilize an essentially new receptor conformation, pharmacological models quantifying these effects must have the capability of describing all possible effects on basal signaling and agonist-induced threshold, sensitivity and maximal effect. The most simple model to describe these types of effects is a combination of the Ehlert model (Eq. (1)) for allosteric interaction of a ligand [A], modulator [B] and receptor (R) [4] and the Black/Leff

Negative allosteric antagonism (allosteric antagonists)

The first type of allosteric ligand to be considered is one that reduces the affinity and/or the efficacy of the agonist (allosteric antagonism). As already shown by Eq. (3), fractional values of α denote decreases in the affinity of the probe agonist ligand. If there is no concomitant effect on efficacy (β = 1) then dextral displacement of dose–response curves (to a maximal value of α⁻¹) occurs. From Eq. (5) it can be shown that the maximal response to an agonist A in the presence of an

Positive allosteric modulation (PAM)

For allosteric ligands with values of α and/or β > 1 (referred to as PAMs for positive allosteric modulators), potentiation of agonist effect will be seen. As with allosteric antagonism, α effects will translate to movement of agonist dose–response curves along the concentration axis (sinistral displacement for PAMs as opposed to dextral displacement for antagonists) with no changes in the maximal response. By contrast increases in β will do the same for full agonists (since no further increases

Allosteric agonism

Allosteric changes in receptor conformations can alter the affinity and efficacy of co-binding ligands and because signaling proteins (i.e. G proteins, β-arrestin, among others) are also co-binding ligands, direct agonism can be the result. All 7TMR agonism is allosteric in nature because agonists are modulators of receptor conduit proteins to modify their interactions with cytosolic guests (signaling proteins [19]). Specifically, agonist ligands can be considered as being modulators that

Allosteric ligands have conditional activity

Unlike conventional ligands that bind to orthosteric (binding site for endogenous ligand) sites, the binding of allosteric ligands is subject to the type and concentration of co-binding ligands. This can be seen directly from the equation yielding the concentration of ligand–receptor complex (expressed as a fraction of total receptor) for binding: $\frac{[BR]}{[R_{t}]} = \frac{[B] / K_{B} (1 + α [A] / K_{A})}{[B] / K_{B} (1 + α [A] / K_{A}) + [A] / K_{A} + 1}$ where [A] is the co-binding ligand, K_B and K_A refer to the equilibrium dissociation constants of

Conclusions

Allosteric ligands have several properties that distinguish them from orthosteric ligands; these are the result of the unique mechanism of allosteric ligands that lead to saturation of effect and probe-dependence [24]. Because allosteric effects are conditional (i.e. they depend on the nature and presence of ligands co-binding to the receptor), they can be very complex and they can vary with the basal activity and sensitivity of the system. For this reason it is especially important that

References (24)

T.P. Kenakin et al.
Analytical pharmacology: the impact of numbers on pharmacology
Trends Pharmacol. Sci.
(2011)
K. Leach
Allosteric GPCR modulators: taking advantage of permissive receptor pharmacology
Trends Pharmacol. Sci.
(2007)
T.P. Kenakin
Ligand detection in the allosteric world
J. Biomol. Screen.
(2010)
K.J. Gregory
Identification of orthosteric and allosteric site mutations in M2 muscarinic acetylcholine receptors that contribute to ligand-selective signaling bias
J. Biol. Chem.
(2010)
J.W. Black
A personal view of pharmacology
Ann. Rev. Pharmacol.
(1996)
T.P. Kenakin
New concepts in drug discovery: collateral efficacy and permissive antagonism
Nat. Rev. Drug Discov.
(2005)
F.J. Ehlert
Estimation of the affinities of allosteric ligands using radioligand binding and pharmacological null methods
Mol. Pharmacol.
(1988)
J.M. Stockton
Modification of the binding properties of muscarinic receptors by gallamine
Mol. Pharmacol.
(1983)
O. Arunlakshana et al.
Some quantitative uses of drug antagonists
Br. J. Pharmacol.
(1959)
C. Watson
The CCR5 receptor-based mechanism of action of 873140, a potent allosteric non-competitive HIV entry-inhibitor
Mol. Pharmacol.
(2005)

K. Maeda

Spirodiketopiperazine-based CCR5 inhibitor which preserves CC-chemokine/CCR5 interactions and exerts potent activity against R5 human immunodeficiency virus type 1 in vitro

J. Virol.

(2004)

J.W. Black et al.

Operational models of pharmacological agonist

Proc. R. Soc. Lond. [Biol.]

(1983)

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Allosteric modulationAnalytical pharmacology and allosterism: the importance of quantifying drug parameters in drug discovery

Graphical abstract

Introduction

Section snippets

Allosteric modulation

Allosteric mechanisms lead to unique drug behaviors

Modeling functional allosteric effects

Negative allosteric antagonism (allosteric antagonists)

Positive allosteric modulation (PAM)

Allosteric agonism

Allosteric ligands have conditional activity

Conclusions

Trends Pharmacol. Sci.

Trends Pharmacol. Sci.

J. Biomol. Screen.

J. Biol. Chem.

A personal view of pharmacology

Ann. Rev. Pharmacol.

New concepts in drug discovery: collateral efficacy and permissive antagonism

Nat. Rev. Drug Discov.

Estimation of the affinities of allosteric ligands using radioligand binding and pharmacological null methods

Mol. Pharmacol.

Modification of the binding properties of muscarinic receptors by gallamine

Mol. Pharmacol.

Some quantitative uses of drug antagonists

Br. J. Pharmacol.

The CCR5 receptor-based mechanism of action of 873140, a potent allosteric non-competitive HIV entry-inhibitor

Mol. Pharmacol.

Spirodiketopiperazine-based CCR5 inhibitor which preserves CC-chemokine/CCR5 interactions and exerts potent activity against R5 human immunodeficiency virus type 1 in vitro

J. Virol.

Operational models of pharmacological agonist

Proc. R. Soc. Lond. [Biol.]

Allosteric modulation
Analytical pharmacology and allosterism: the importance of quantifying drug parameters in drug discovery