HuBio 543 September 6, 2007 Frank F. Vincenzi Pharmacodynamics HuBio 543 September 6, 2007 Frank F. Vincenzi

Learning Objectives Quantification of drug receptor interactions and responses Potency Schild equation and regression Competitive and non-competitive antagonism Spare receptors Kd, EC50, pD2, pA2 Receptors, signal transduction, transmembrane signaling Agonist, antagonist, partial agonist, inverse agonist, multiple receptor states Intrinsic activity, efficacy, SAR Desensitization, up and down regulation

Typical concentration-effect curve (plotted arithmetically)

A slide rule (logarithmic scale) In the scale L, numbers show the distance from the origin where the length of rule is assumed to be 1(one). The scales D and C are related to the fanction y = log x, where the base of it is 10. In scales D and C, the numbers are placed at points whose distance from the origin is ther logarithm. The scale L gives the mantissa of the logarithm of the number standing above it in the scale C. For example, log 3.0 = 0.48 by this slide rule. The scale CI has the same graduation as the scale C, but it is in the reverse order from right to left. A numbers in the scale C gives the inverse of the number located below it in the scale C. For example, the inverse of 3.0 is 0.33 by this slide rule. As in this example, a slide rule requires a rough calculation to determine the number of digits before the dismal point.

Typical log concentration-effect curve (graded ‘dose-response’ curve)

Drug (D) - Receptor (R) Interaction k1 D + R DR k2 Kd = ([D] * [R]) / [DR] = k2/k1 Kd = dissociation constant k1 = association rate constant k2 = dissociation rate constant

Several ways to express agonist potency &/or apparent affinity of agonists EC50 (effective concentration, 50%, M) Kd (apparent dissociation constant, M) pD2 (negative log of molar concentration (M) of the drug giving a response, which when compared to the maximum, gives a ratio of 2) (i.e., negative log of half maximal concentration)

The classical concentration-effect relationship and the laws of mass action Effect = (Effectmax * conc)/(conc + EC50) In the previous data slide EC50 ~ 3 x 10-9 M Thus, the apparent Kd of ACh ~ 3 x 10-9 M IF (NOTE, BIG IF) EC50 = Kd then Bound drug = (Bmax * conc)/(conc + Kd)

Binding of a radioligand to tissue samples Adapted from Schaffhauser et al., 1998

Scatchard analysis of binding of 125iodocyanopindolol to beta-receptors in human heart Adapted from Heitz et al., 1983

Acetylcholine (ACh): One drug with different affinities for two different receptors (adapted from Clark, 1933)

ACh: Different affinities for different receptors Muscarinic receptors EC50 = apparent Kd ~ 3 x 10-8 M, pD2 ~7.5 Nicotinic receptor EC50 = apparent Kd ~ 3 x 10-6 M, pD2 ~5.5 In these experiments, affinity of ACh for muscarinic receptors is apparently ~100 times greater than for nicotinic receptors. ACh is 100 times more potent as a muscarinic agonist than as a nicotinic agonist. So, when injected as a drug, muscarinic effects normally predominate, unless the muscarinic receptors are blocked. (No problem for nerves releasing ACh locally onto nicotinic receptors, however).

Properties of an agonist (e. g Properties of an agonist (e.g., ACh) (on receptors lacking spontaneous activity) Accessibility Affinity Intrinsic activity > 0

Different affinities of related agonist drugs for the same receptor: Different potencies (adapted from Ariëns et al., 1964)

Properties of an antagonist (on receptors lacking spontaneous activity) Accessibility Affinity Intrinsic activity = 0

Pharmacological antagonism in an intact animal

Properties of a partial agonist (on receptors lacking spontaneous activity) Accessibility Affinity 0 < Intrinsic activity < 1

Theoretical concentration-effect curves for a full and partial agonist of a given receptor

Multiple receptor conformational states: How to understand agonists, partial agonists and antagonists

Simple case: receptor has little or no spontaneous activity in the absence of added drug ‘inactive’ R ‘active’ R

An agonist binds more tightly to the ‘active’ state of the receptor: Equilibrium shifts to the active state

A competitive antagonist binds equally tightly to the ‘inactive’ and active states of the receptor: No change in equilibrium

A partial agonist binds to both the ‘inactive’ and ‘active’ states of the receptor: Partial shift of equilibrium

Multiple receptor states: How to understand inverse agonists (in this LESS SIMPLE case, the receptor has spontaneous (often called constituitive) activity in the absence of added drug)

The less simple case: Some receptors are ‘active’ even in the absence of added drug

Inverse agonists bind more tightly to the resting state of the spontaneously active receptor: Equilibrium shifts toward the inactive state

Receptor activation by agonists, inverse agonists, etc. Newman-Tancredi et al., 1997

How to quantify drug antagonism Schild Equation (C’/C) = 1 + ([I]/Ki) Schild plot or Schild regression log(C’/C - 1) vs. log [I] pA2 = -log([I] giving a dose ratio of 2) Where [I] = Kd of antagonist at its receptor.

Antagonism of acetylcholine by atropine Adapted from Altiere et al., 1994

Schild plot of antagonism of acetylcholine by atropine Adapted from Altiere et al., 1994

Antagonism of acetylcholine by pirenzepine Adapted from Altiere et al., 1994

Schild plot: Antagonism of acetylcholine by two different antagonists 3 atropine pirenzepine 2 1 -10 -9 -8 -7 -6 -5 log [antagonist] (M) Adapted from Altiere et al., 1994

Different pA2 values (affinities) for different receptors of some clinically useful drugs: The basis of therapeutic selectivity

Evidence for the existence of spare receptors

How nature achieves neurotransmitter sensitivity without a loss of speed: Spare receptors:

Drug (D) - Receptor (R) Interaction k1 D + R DR k2 Kd = ([D] * [R]) / [DR] = k2/k1 Kd = dissociation constant k1 = association rate constant k2 = dissociation rate constant