Drug Receptors & Pharmacodynamics: Introduction

Therapeutic and toxic effects of drugs result from their interactions with molecules in the patient. Most drugs act by associating with specific macromolecules in ways that alter the macromolecules' biochemical or biophysical activities. This idea, more than a century old, is embodied in the term receptor: the component of a cell or organism that interacts with a drug and initiates the chain of events leading to the drug's observed effects.

Receptors have become the central focus of investigation of drug effects and their mechanisms of action (pharmacodynamics). The receptor concept, extended to endocrinology, immunology, and molecular biology, has proved essential for explaining many aspects of biologic regulation. Many drug receptors have been isolated and characterized in detail, thus opening the way to precise understanding of the molecular basis of drug action.

The receptor concept has important practical consequences for the development of drugs and for arriving at therapeutic decisions in clinical practice. These consequences form the basis for understanding the actions and clinical uses of drugs described in almost every chapter of this book. They may be briefly summarized as follows:

1.      Receptors largely determine the quantitative relations between dose or concentration of drug and pharmacologic effects. The receptor's affinity for binding a drug determines the concentration of drug required to form a significant number of drug-receptor complexes, and the total number of receptors may limit the maximal effect a drug may produce.

2.      Receptors are responsible for selectivity of drug action. The molecular size, shape, and electrical charge of a drug determine whether—and with what affinity—it will bind to a particular receptor among the vast array of chemically different binding sites available in a cell, tissue, or patient. Accordingly, changes in the chemical structure of a drug can dramatically increase or decrease a new drug's affinities for different classes of receptors, with resulting alterations in therapeutic and toxic effects.

3.      Receptors mediate the actions of pharmacologic agonists and antagonists. Some drugs and many natural ligands, such as hormones and neurotransmitters, regulate the function of receptor macromolecules as agonists; this means that they activate the receptor to signal as a direct result of binding to it. Some agonists activate a single kind of receptor to produce all their biologic functions, whereas others selectively promote one receptor function more than another.

Other drugs act as pharmacologic antagonists; that is, they bind to receptors but do not activate generation of a signal; consequently, they interfere with the ability of an agonist to activate the receptor. The effect of a so-called "pure" antagonist on a cell or in a patient depends entirely on its preventing the binding of agonist molecules and blocking their biologic actions. Other antagonists, in addition to preventing agonist binding, suppress the basal signaling ("constitutive") activity of receptors. Some of the most useful drugs in clinical medicine are pharmacologic antagonists.

Macromolecular Nature of Drug Receptors

Most receptors are proteins, presumably because the structures of polypeptides provide both the necessary diversity and the necessary specificity of shape and electrical charge. Receptors vary greatly in structure and can be identified in many ways. Traditionally, drug binding was used to identify or purify receptors from tissue extracts; consequently, receptors were discovered more recently than the drugs that bind to them. However, advances in molecular biology and genome sequencing have begun to reverse this order. Now receptors are being discovered by predicted structure or sequence homology to other (known) receptors, and drugs that bind to them are developed later using chemical screening methods. This effort has revealed, for many known drugs, a larger diversity of receptors than previously anticipated. It has also identified a number of "orphan" receptors, so-called because their ligands are presently unknown, which may prove to be useful targets for the development of new drugs.

The best-characterized drug receptors are regulatory proteins, which mediate the actions of endogenous chemical signals such as neurotransmitters, autacoids, and hormones. This class of receptors mediates the effects of many of the most useful therapeutic agents. The molecular structures and biochemical mechanisms of these regulatory receptors are described in a later section entitled Signaling Mechanisms & Drug Action.

Other classes of proteins that have been clearly identified as drug receptors include enzymes, which may be inhibited (or, less commonly, activated) by binding a drug (eg, dihydrofolate reductase, the receptor for the antineoplastic drug methotrexate); transport proteins (eg, Na⁺,K⁺ ATPase, the membrane receptor for cardioactive digitalis glycosides); and structural proteins (eg, tubulin, the receptor for colchicine, an anti-inflammatory agent).

This chapter deals with three aspects of drug receptor function, presented in increasing order of complexity: (1) receptors as determinants of the quantitative relation between the concentration of a drug and the pharmacologic response, (2) receptors as regulatory proteins and components of chemical signaling mechanisms that provide targets for important drugs, and (3) receptors as key determinants of the therapeutic and toxic effects of drugs in patients.

Relation between Drug Concentration & Response

The relation between dose of a drug and the clinically observed response may be complex. In carefully controlled in vitro systems, however, the relation between concentration of a drug and its effect is often simple and can be described with mathematical precision. This idealized relation underlies the more complex relations between dose and effect that occur when drugs are given to patients.

Concentration-Effect Curves & Receptor Binding of Agonists

Even in intact animals or patients, responses to low doses of a drug usually increase in direct proportion to dose. As doses increase, however, the response increment diminishes; finally, doses may be reached at which no further increase in response can be achieved. In idealized or in vitro systems, the relation between drug concentration and effect is described by a hyperbolic curve (Figure 2–1A) according to the following equation:

where E is the effect observed at concentration C, E_max is the maximal response that can be produced by the drug, and EC₅₀ is the concentration of drug that produces 50% of maximal effect.

This hyperbolic relation resembles the mass action law, which describes association between two molecules of a given affinity. This resemblance suggests that drug agonists act by binding to ("occupying") a distinct class of biologic molecules with a characteristic affinity for the drug receptor. Radioactive receptor ligands have been used to confirm this occupancy assumption in many drug-receptor systems. In these systems, drug bound to receptors (B) relates to the concentration of free (unbound) drug (C) as depicted in Figure 2–1B and as described by an analogous equation:

in which B_max indicates the total concentration of receptor sites (ie, sites bound to the drug at infinitely high concentrations of free drug) and K_d (the equilibrium dissociation constant) represents the concentration of free drug at which half-maximal binding is observed. This constant characterizes the receptor's affinity for binding the drug in a reciprocal fashion: If the K_d is low, binding affinity is high, and vice versa. The EC₅₀ and K_d may be identical, but need not be, as discussed below. Dose-response data are often presented as a plot of the drug effect (ordinate) against the logarithm of the dose or concentration (abscissa). This mathematical maneuver transforms the hyperbolic curve of Figure 2–1 into a sigmoid curve with a linear midportion (eg, Figure 2–2). This expands the scale of the concentration axis at low concentrations (where the effect is changing rapidly) and compresses it at high concentrations (where the effect is changing slowly), but has no special biologic or pharmacologic significance.

Receptor-Effector Coupling & Spare Receptors

When a receptor is occupied by an agonist, the resulting conformational change is only the first of many steps usually required to produce a pharmacologic response. The transduction process that links drug occupancy of receptors and pharmacologic response is often termed coupling. The relative efficiency of occupancy-response coupling is partially determined by the initial conformational change in the receptor; thus, the effects of full agonists can be considered more efficiently coupled to receptor occupancy than can the effects of partial agonists (described in text that follows). Coupling efficiency is also determined by the biochemical events that transduce receptor occupancy into cellular response. Sometimes the biologic effect of the drug is linearly related to the number of receptors bound. This is often true for drug-regulated ion channels, eg, in which the ion current produced by the drug is directly proportional to the number of receptors (ion channels) bound. In other cases, the biologic response is a more complex function of drug binding to receptors. This is often true for receptors linked to enzymatic signal transduction cascades, eg, in which the biologic response often increases disproportionately to the number of receptors occupied by drug.

Many factors can contribute to nonlinear occupancy-response coupling, and often these factors are only partially understood. The concept of "spare" receptors, regardless of the precise biochemical mechanism involved, can help us to think about these effects. Receptors are said to be "spare" for a given pharmacologic response if it is possible to elicit a maximal biologic response at a concentration of agonist that does not result in occupancy of the full complement of available receptors. Experimentally, spare receptors may be demonstrated by using irreversible antagonists to prevent binding of agonist to a proportion of available receptors and showing that high concentrations of agonist can still produce an undiminished maximal response (Figure 2–2). Thus, the same maximal inotropic response of heart muscle to catecholamines can be elicited even under conditions in which 90% of the adrenoceptors are occupied by a quasi-irreversible antagonist. Accordingly, myocardial cells are said to contain a large proportion of spare adrenoceptors.

How can we account for the phenomenon of spare receptors? In the example of the adrenoceptor, receptor activation promotes binding of guanosine triphosphate (GTP) to an intermediate signaling protein and activation of the signaling intermediate may greatly outlast the agonist-receptor interaction (see the following section on G Proteins & Second Messengers). In such a case, the "spareness" of receptors is temporal. Maximal response can be elicited by activation of relatively few receptors because the response initiated by an individual ligand-receptor binding event persists longer than the binding event itself.

In other cases, in which the biochemical mechanism is not understood, we imagine that the receptors might be spare in number. If the concentration or amount of cellular components other than the receptors limits the coupling of receptor occupancy to response, then a maximal response can occur without occupancy of all receptors. Thus, the sensitivity of a cell or tissue to a particular concentration of agonist depends not only on the affinity of the receptor for binding the agonist (characterized by the K_d) but also on the degree of spareness—the total number of receptors present compared with the number actually needed to elicit a maximal biologic response.

The concept of spare receptors is very useful clinically because it allows one to think precisely about the effects of drug dosage without needing to consider biochemical details of the signaling response. The K_d of the agonist-receptor interaction determines what fraction (B/B_max) of total receptors will be occupied at a given free concentration (C) of agonist regardless of the receptor concentration:

Imagine a responding cell with four receptors and four effectors. Here the number of effectors does not limit the maximal response, and the receptors are not spare in number. Consequently, an agonist present at a concentration equal to the K_d will occupy 50% of the receptors, and half of the effectors will be activated, producing a half-maximal response (ie, two receptors stimulate two effectors). Now imagine that the number of receptors increases 10-fold to 40 receptors but that the total number of effectors remains constant. Most of the receptors are now spare in number. As a result, a much lower concentration of agonist suffices to occupy 2 of the 40 receptors (5% of the receptors), and this same low concentration of agonist is able to elicit a half-maximal response (two of four effectors activated). Thus, it is possible to change the sensitivity of tissues with spare receptors by changing the receptor concentration.

Competitive & Irreversible Antagonists

Receptor antagonists bind to receptors but do not activate them. The primary action of antagonists is to prevent agonists (other drugs or endogenous regulatory molecules) from activating receptors. Some antagonists (so-called "inverse agonists"), also reduce receptor activity below basal levels observed in the absence of bound ligand. Antagonists are divided into two classes depending on whether or not they reversibly compete with agonists for binding to receptors.

In the presence of a fixed concentration of agonist, increasing concentrations of a reversible competitive antagonist progressively inhibit the agonist response; high antagonist concentrations prevent response completely. Conversely, sufficiently high concentrations of agonist can surmount the effect of a given concentration of the antagonist; that is, the E_max for the agonist remains the same for any fixed concentration of antagonist (Figure 2–3A). Because the antagonism is competitive, the presence of antagonist increases the agonist concentration required for a given degree of response, and so the agonist concentration-effect curve is shifted to the right.

The concentration (C') of an agonist required to produce a given effect in the presence of a fixed concentration ([I]) of competitive antagonist is greater than the agonist concentration (C) required to produce the same effect in the absence of the antagonist. The ratio of these two agonist concentrations (dose ratio) is related to the dissociation constant (K_i) of the antagonist by the Schild equation:

Pharmacologists often use this relation to determine the K_i of a competitive antagonist. Even without knowledge of the relation between agonist occupancy of the receptor and response, the K_i can be determined simply and accurately. As shown in Figure 2–3, concentration response curves are obtained in the presence and in the absence of a fixed concentration of competitive antagonist; comparison of the agonist concentrations required to produce identical degrees of pharmacologic effect in the two situations reveals the antagonist's K_i. If C' is twice C, for example, then [I] = K_i.

For the clinician, this mathematical relation has two important therapeutic implications:

1.      The degree of inhibition produced by a competitive antagonist depends on the concentration of antagonist. The competitive -adrenoceptor antagonist propranolol provides a useful example. Patients receiving a fixed dose of this drug exhibit a wide range of plasma concentrations, owing to differences among individuals in clearance of propranolol. As a result, inhibitory effects on physiologic responses to norepinephrine and epinephrine (endogenous adrenergic receptor agonists) may vary widely, and the dose of propranolol must be adjusted accordingly.

2.      Clinical response to a competitive antagonist depends on the concentration of agonist that is competing for binding to receptors. Here also propranolol provides a useful example: When this drug is administered at moderate doses sufficient to block the effect of basal levels of the neurotransmitter norepinephrine, resting heart rate is decreased. However, increase in the release of norepinephrine and epinephrine that occurs with exercise, postural changes, or emotional stress may suffice to overcome this competitive antagonism. Accordingly, the same dose of propranolol may have little effect under these conditions, thereby altering therapeutic response.

Some receptor antagonists bind to the receptor in an irreversible or nearly irreversible fashion, either by forming a covalent bond with the receptor or by binding so tightly that, for practical purposes, the receptor is unavailable for binding of agonist. After occupancy of some proportion of receptors by such an antagonist, the number of remaining unoccupied receptors may be too low for the agonist (even at high concentrations) to elicit a response comparable to the previous maximal response (Figure 2–3B). If spare receptors are present, however, a lower dose of an irreversible antagonist may leave enough receptors unoccupied to allow achievement of maximum response to agonist, although a higher agonist concentration will be required (Figure 2–2B and C; see Receptor-Effector Coupling & Spare Receptors).

Therapeutically, irreversible antagonists present distinct advantages and disadvantages. Once the irreversible antagonist has occupied the receptor, it need not be present in unbound form to inhibit agonist responses. Consequently, the duration of action of such an irreversible antagonist is relatively independent of its own rate of elimination and more dependent on the rate of turnover of receptor molecules.

Phenoxybenzamine, an irreversible -adrenoceptor antagonist, is used to control the hypertension caused by catecholamines released from pheochromocytoma, a tumor of the adrenal medulla. If administration of phenoxybenzamine lowers blood pressure, blockade will be maintained even when the tumor episodically releases very large amounts of catecholamine. In this case, the ability to prevent responses to varying and high concentrations of agonist is a therapeutic advantage. If overdose occurs, however, a real problem may arise. If the -adrenoceptor blockade cannot be overcome, excess effects of the drug must be antagonized "physiologically," ie, by using a pressor agent that does not act via receptors.

Antagonists can function noncompetitively in a different way; that is, by binding to a site on the receptor protein separate from the agonist binding site and thereby preventing receptor activation without blocking agonist binding. Although these drugs act noncompetitively, their actions are reversible if they do not bind covalently. Some drugs, often called allosteric modulators, bind to a separate site on the receptor protein and alter receptor function without inactivating the receptor. For example, benzodiazepines bind noncompetitively to ion channels activated by the neurotransmitter -aminobutyric acid (GABA), enhancing the net activating effect of GABA on channel conductance.

Partial Agonists

Based on the maximal pharmacologic response that occurs when all receptors are occupied, agonists can be divided into two classes: partial agonists produce a lower response, at full receptor occupancy, than do full agonists. Partial agonists produce concentration-effect curves that resemble those observed with full agonists in the presence of an antagonist that irreversibly blocks some of the receptor sites (compare Figures 2–2 [curve D] and 2–4B). It is important to emphasize that the failure of partial agonists to produce a maximal response is not due to decreased affinity for binding to receptors. Indeed, a partial agonist's inability to cause a maximal pharmacologic response, even when present at high concentrations that saturate binding to all receptors, is indicated by the fact that partial agonists competitively inhibit the responses produced by full agonists (Figure 2–4C). Many drugs used clinically as antagonists are actually weak partial agonists. Partial agonism can be useful in some clinical circumstances. For example, buprenorphine, a partial agonist of -opioid receptors, is a generally safer analgesic drug than morphine because it produces less respiratory depression in overdose. Buprenorphine is effectively antianalgesic when administered to morphine-dependent individuals, however, and may precipitate a drug withdrawal syndrome.

Other Mechanisms of Drug Antagonism

Not all the mechanisms of antagonism involve interactions of drugs or endogenous ligands at a single type of receptor, and some types of antagonism do not involve a receptor at all. For example, protamine, a protein that is positively charged at physiologic pH, can be used clinically to counteract the effects of heparin, an anticoagulant that is negatively charged. In this case, one drug acts as a chemical antagonist of the other simply by ionic binding that makes the other drug unavailable for interactions with proteins involved in blood clotting.

Another type of antagonism is physiologic antagonism between endogenous regulatory pathways mediated by different receptors. For example, several catabolic actions of the glucocorticoid hormones lead to increased blood sugar, an effect that is physiologically opposed by insulin. Although glucocorticoids and insulin act on quite distinct receptor-effector systems, the clinician must sometimes administer insulin to oppose the hyperglycemic effects of a glucocorticoid hormone, whether the latter is elevated by endogenous synthesis (eg, a tumor of the adrenal cortex) or as a result of glucocorticoid therapy.

In general, use of a drug as a physiologic antagonist produces effects that are less specific and less easy to control than are the effects of a receptor-specific antagonist. Thus, for example, to treat bradycardia caused by increased release of acetylcholine from vagus nerve endings, the physician could use isoproterenol, a -adrenoceptor agonist that increases heart rate by mimicking sympathetic stimulation of the heart. However, use of this physiologic antagonist would be less rational—and potentially more dangerous—than would use of a receptor-specific antagonist such as atropine (a competitive antagonist at the receptors at which acetylcholine slows heart rate).

Signaling Mechanisms & Drug Action

Until now we have considered receptor interactions and drug effects in terms of equations and concentration-effect curves. We must also understand the molecular mechanisms by which a drug acts. Such understanding allows us to ask basic questions with important clinical implications:

       Why do some drugs produce effects that persist for minutes, hours, or even days after the drug is no longer present?

       Why do responses to other drugs diminish rapidly with prolonged or repeated administration?

       How do cellular mechanisms for amplifying external chemical signals explain the phenomenon of spare receptors?

       Why do chemically similar drugs often exhibit extraordinary selectivity in their actions?

       Do these mechanisms provide targets for developing new drugs?

Most transmembrane signaling is accomplished by a small number of different molecular mechanisms. Each type of mechanism has been adapted, through the evolution of distinctive protein families, to transduce many different signals. These protein families include receptors on the cell surface and within the cell, as well as enzymes and other components that generate, amplify, coordinate, and terminate postreceptor signaling by chemical second messengers in the cytoplasm. This section first discusses the mechanisms for carrying chemical information across the plasma membrane and then outlines key features of cytoplasmic second messengers.

Five basic mechanisms of transmembrane signaling are well understood (Figure 2–5). Each uses a different strategy to circumvent the barrier posed by the lipid bilayer of the plasma membrane. These strategies use (1) a lipid-soluble ligand that crosses the membrane and acts on an intracellular receptor; (2) a transmembrane receptor protein whose intracellular enzymatic activity is allosterically regulated by a ligand that binds to a site on the protein's extracellular domain; (3) a transmembrane receptor that binds and stimulates a protein tyrosine kinase; (4) a ligand-gated transmembrane ion channel that can be induced to open or close by the binding of a ligand; or (5) a transmembrane receptor protein that stimulates a GTP-binding signal transducer protein (G protein), which in turn modulates production of an intracellular second messenger.

Although the five established mechanisms do not account for all the chemical signals conveyed across cell membranes, they do transduce many of the most important signals exploited in pharmacotherapy.

Intracellular Receptors for Lipid-Soluble Agents

Several biologic ligands are sufficiently lipid-soluble to cross the plasma membrane and act on intracellular receptors. One class of such ligands includes steroids (corticosteroids, mineralocorticoids, sex steroids, vitamin D), and thyroid hormone, whose receptors stimulate the transcription of genes by binding to specific DNA sequences near the gene whose expression is to be regulated. Many of the target DNA sequences (called response elements) have been identified.

These "gene-active" receptors belong to a protein family that evolved from a common precursor. Dissection of the receptors by recombinant DNA techniques has provided insights into their molecular mechanism. For example, binding of glucocorticoid hormone to its normal receptor protein relieves an inhibitory constraint on the transcription-stimulating activity of the protein. Figure 2–6 schematically depicts the molecular mechanism of glucocorticoid action: In the absence of hormone, the receptor is bound to hsp90, a protein that appears to prevent normal folding of several structural domains of the receptor. Binding of hormone to the ligand-binding domain triggers release of hsp90. This allows the DNA-binding and transcription-activating domains of the receptor to fold into their functionally active conformations, so that the activated receptor can initiate transcription of target genes.

The mechanism used by hormones that act by regulating gene expression has two therapeutically important consequences:

1.      All of these hormones produce their effects after a characteristic lag period of 30 minutes to several hours—the time required for the synthesis of new proteins. This means that the gene-active hormones cannot be expected to alter a pathologic state within minutes (eg, glucocorticoids will not immediately relieve the symptoms of acute bronchial asthma).

2.      The effects of these agents can persist for hours or days after the agonist concentration has been reduced to zero. The persistence of effect is primarily due to the relatively slow turnover of most enzymes and proteins, which can remain active in cells for hours or days after they have been synthesized. Consequently, it means that the beneficial (or toxic) effects of a gene-active hormone usually decrease slowly when administration of the hormone is stopped.

Ligand-Regulated Transmembrane Enzymes Including Receptor Tyrosine Kinases

This class of receptor molecules mediates the first steps in signaling by insulin, epidermal growth factor (EGF), platelet-derived growth factor (PDGF), atrial natriuretic peptide (ANP), transforming growth factor- (TGF-), and many other trophic hormones. These receptors are polypeptides consisting of an extracellular hormone-binding domain and a cytoplasmic enzyme domain, which may be a protein tyrosine kinase, a serine kinase, or a guanylyl cyclase (Figure 2–7). In all these receptors, the two domains are connected by a hydrophobic segment of the polypeptide that crosses the lipid bilayer of the plasma membrane.

The receptor tyrosine kinase signaling pathway begins with binding of ligand, typically a polypeptide hormone or growth factor, to the receptor's extracellular domain. The resulting change in receptor conformation causes receptor molecules to bind to one another, which in turn brings together the tyrosine kinase domains, which become enzymatically active, and phosphorylate one another as well as additional downstream signaling proteins. Activated receptors catalyze phosphorylation of tyrosine residues on different target signaling proteins, thereby allowing a single type of activated receptor to modulate a number of biochemical processes.

Insulin, for example, uses a single class of receptors to trigger increased uptake of glucose and amino acids and to regulate metabolism of glycogen and triglycerides in the cell. Similarly, each of the growth factors initiates in its specific target cells a complex program of cellular events ranging from altered membrane transport of ions and metabolites to changes in the expression of many genes.

Inhibitors of receptor tyrosine kinases are finding increased use in neoplastic disorders in which excessive growth factor signaling is often involved. Some of these inhibitors are monoclonal antibodies (eg, trastuzumab, cetuximab), which bind to the extracellular domain of a particular receptor and interfere with binding of growth factor. Other inhibitors are membrane-permeant "small molecule" chemicals (eg, gefitinib, erlotinib), which inhibit the receptor's kinase activity in the cytoplasm.

The intensity and duration of action of EGF, PDGF, and other agents that act via receptor tyrosine kinases are limited by a process called receptor down-regulation. Ligand binding often induces accelerated endocytosis of receptors from the cell surface, followed by the degradation of those receptors (and their bound ligands). When this process occurs at a rate faster than de novo synthesis of receptors, the total number of cell-surface receptors is reduced (down-regulated), and the cell's responsiveness to ligand is correspondingly diminished. A well-understood example is the EGF receptor tyrosine kinase, which undergoes rapid endocytosis and is trafficked to lysosomes after EGF binding; genetic mutations that interfere with this process cause excessive growth factor–induced cell proliferation and are associated with an increased susceptibility to certain types of cancer. Endocytosis of other receptor tyrosine kinases, most notably receptors for nerve growth factor, serves a very different function. Internalized nerve growth factor receptors are not rapidly degraded and are translocated in endocytic vesicles from the distal axon, where receptors are activated by nerve growth factor released from the innervated tissue, to the cell body. In the cell body, the growth factor signal is transduced to transcription factors regulating the expression of genes controlling cell survival. This process effectively transports a critical survival signal from its site of release to its site of signaling effect, and does so over a remarkably long distance—up to 1 meter in certain sensory neurons.

A number of regulators of growth and differentiation, including TGF-, act on another class of transmembrane receptor enzymes that phosphorylate serine and threonine residues. ANP, an important regulator of blood volume and vascular tone, acts on a transmembrane receptor whose intracellular domain, a guanylyl cyclase, generates cGMP (see below). Receptors in both groups, like the receptor tyrosine kinases, are active in their dimeric forms.

Cytokine Receptors

Cytokine receptors respond to a heterogeneous group of peptide ligands, which include growth hormone, erythropoietin, several kinds of interferon, and other regulators of growth and differentiation. These receptors use a mechanism (Figure 2–8) closely resembling that of receptor tyrosine kinases, except that in this case, the protein tyrosine kinase activity is not intrinsic to the receptor molecule. Instead, a separate protein tyrosine kinase, from the Janus-kinase (JAK) family, binds noncovalently to the receptor. As in the case of the EGF receptor, cytokine receptors dimerize after they bind the activating ligand, allowing the bound JAKs to become activated and to phosphorylate tyrosine residues on the receptor. Phosphorylated tyrosine residues on the receptor's cytoplasmic surface then set in motion a complex signaling dance by binding another set of proteins, called STATs (signal transducers and activators of transcription). The bound STATs are themselves phosphorylated by the JAKs, two STAT molecules dimerize (attaching to one another's tyrosine phosphates), and finally the STAT/STAT dimer dissociates from the receptor and travels to the nucleus, where it regulates transcription of specific genes.

Ligand- and Voltage-Gated Channels

Many of the most useful drugs in clinical medicine act by mimicking or blocking the actions of endogenous ligands that regulate the flow of ions through plasma membrane channels. The natural ligands are acetylcholine, serotonin, GABA, and glutamate. All of these agents are synaptic transmitters.

Each of their receptors transmits its signal across the plasma membrane by increasing transmembrane conductance of the relevant ion and thereby altering the electrical potential across the membrane. For example, acetylcholine causes the opening of the ion channel in the nicotinic acetylcholine receptor (AChR), which allows Na⁺ to flow down its concentration gradient into cells, producing a localized excitatory postsynaptic potential—a depolarization.

The AChR is one of the best characterized of all cell-surface receptors for hormones or neurotransmitters (Figure 2–9). One form of this receptor is a pentamer made up of four different polypeptide subunits (eg, two chains plus one , one , and one chain, all with molecular weights ranging from 43,000 to 50,000). These polypeptides, each of which crosses the lipid bilayer four times, form a cylindrical structure that is 8 nm in diameter. When acetylcholine binds to sites on the subunits, a conformational change occurs that results in the transient opening of a central aqueous channel through which sodium ions penetrate from the extracellular fluid into the cell.

The time elapsed between the binding of the agonist to a ligand-gated channel and the cellular response can often be measured in milliseconds. The rapidity of this signaling mechanism is crucially important for moment-to-moment transfer of information across synapses. Ligand-gated ion channels can be regulated by multiple mechanisms, including phosphorylation and endocytosis. In the central nervous system, these mechanisms contribute to synaptic plasticity involved in learning and memory.

Voltage-gated ion channels do not bind neurotransmitters directly but are controlled by membrane potential; such channels are also important drug targets. For example, verapamil inhibits voltage-gated calcium channels that are present in the heart and in vascular smooth muscle, producing antiarrhythmic effects and reducing blood pressure.

G Proteins & Second Messengers

Many extracellular ligands act by increasing the intracellular concentrations of second messengers such as cyclic adenosine-3',5'-monophosphate (cAMP), calcium ion, or the phosphoinositides (described below). In most cases, they use a transmembrane signaling system with three separate components. First, the extracellular ligand is specifically detected by a cell-surface receptor. The receptor in turn triggers the activation of a G protein located on the cytoplasmic face of the plasma membrane. The activated G protein then changes the activity of an effector element, usually an enzyme or ion channel. This element then changes the concentration of the intracellular second messenger. For cAMP, the effector enzyme is adenylyl cyclase, a membrane protein that converts intracellular adenosine triphosphate (ATP) to cAMP. The corresponding G protein, G_s, stimulates adenylyl cyclase after being activated by hormones and neurotransmitters that act via specific G_s-coupled receptors. There are many examples of such receptors, including adrenoceptors, glucagon receptors, thyrotropin receptors, and certain subtypes of dopamine and serotonin receptors.

G_s and other G proteins use a molecular mechanism that involves binding and hydrolysis of GTP (Figure 2–10). This mechanism allows the transduced signal to be amplified. For example, a neurotransmitter such as norepinephrine may encounter its membrane receptor for only a few milliseconds. When the encounter generates a GTP-bound G_s molecule, however, the duration of activation of adenylyl cyclase depends on the longevity of GTP binding to G_s rather than on the receptor's affinity for norepinephrine. Indeed, like other G proteins, GTP-bound G_s may remain active for tens of seconds, enormously amplifying the original signal. This mechanism also helps explain how signaling by G proteins produces the phenomenon of spare receptors. The family of G proteins contains several functionally diverse subfamilies (Table 2–1), each of which mediates effects of a particular set of receptors to a distinctive group of effectors. Note that an endogenous ligand (eg, norepinephrine, acetylcholine, serotonin, many others not listed in Table 2–1) may bind and stimulate receptors that couple to different subsets of G proteins. The apparent promiscuity of such a ligand allows it to elicit different G protein-dependent responses in different cells. For instance, the body responds to danger by using catecholamines (norepinephrine and epinephrine) both to increase heart rate and to induce constriction of blood vessels in the skin, by acting on G_s-coupled adrenoceptors and G_q-coupled ₁ adrenoceptors, respectively. Ligand promiscuity also offers opportunities in drug development (see Receptor Classes & Drug Development in the following text).

Receptors coupled to G proteins (GPCRs)  make up a family of "seven-transmembrane" (7-TM) or "serpentine" receptors, so called because the receptor polypeptide chain "snakes" across the plasma membrane seven times (Figure 2–11). Receptors for adrenergic amines, serotonin, acetylcholine (muscarinic but not nicotinic), many peptide hormones, odorants, and even visual receptors (in retinal rod and cone cells) all belong to the serpentine family. All were derived from a common evolutionary precursor. Several serpentine receptors exist as dimers or larger complexes. These complexes include homodimers (complexes of two identical GPCRs and heterodimers (complexes of different GPCRs). The GABA_B and metabotropic glutamate receptors are composed of two related serpentine receptor subunits in an obligate heterodimer; the presence of both subunits is required for normal signaling function in vivo. A number of other serpentine receptors can exist as homo- or heterodimers and, in experimental systems, some of these complexes have distinct pharmacologic properties that suggest interesting opportunities for drug development. The physiologic significance of most of these oligomeric complexes is not yet known. Serpentine receptors signal by a different mechanism than receptor tyrosine kinases and cytokine receptors, however, and it is thought that dimerization is not essential for activation of many serpentine receptors.

All serpentine receptors transduce signals across the plasma membrane in essentially the same way. Often the agonist ligand—eg, a catecholamine or acetylcholine—is bound in a pocket enclosed by the transmembrane regions of the receptor (as in Figure 2–11). The resulting change in conformation of these regions is transmitted to cytoplasmic loops of the receptor, which in turn activate the appropriate G protein by promoting replacement of GDP by GTP, as described above. Amino acids in the third cytoplasmic loop of the serpentine receptor polypeptide are generally thought to play a key role in mediating receptor interaction with G proteins (shown by arrows in Figure 2–11). The structural basis for binding of an inverse agoinist to the ₂ receptor was determined recently using X-ray crystallography.

Receptor Regulation

G protein-mediated responses to drugs and hormonal agonists often attenuate with time (Figure 2–12, top). After reaching an initial high level, the response (eg, cellular cAMP accumulation, Na⁺ influx, contractility, etc) diminishes over seconds or minutes, even in the continued presence of the agonist. This "desensitization" is often rapidly reversible; a second exposure to agonist, if provided a few minutes after termination of the first exposure, results in a response similar to the initial response.

The mechanism mediating rapid desensitization of G protein-coupled receptors often involves receptor phosphorylation, as illustrated by rapid desensitization of the adrenoceptor (Figure 2–12, top). The agonist-induced change in conformation of the receptor causes it to bind, activate, and serve as a substrate for a family of specific receptor kinases, called G protein-coupled receptor kinases (GRKs). The activated GRK then phosphorylates serine residues in the receptor's carboxyl terminal tail. The presence of phosphoserines increases the receptor's affinity for binding a third protein, -arrestin. Binding of -arrestin to cytoplasmic loops of the receptor diminishes the receptor's ability to interact with G_s, thereby reducing the agonist response (ie, stimulation of adenylyl cyclase). Upon removal of agonist, GRK activation is terminated, and the desensitization process can be reversed by cellular phosphatases.

For the adrenoceptor, and many other serpentine receptors, -arrestin binding also accelerates endocytosis of receptors from the plasma membrane. Endocytosis of receptors promotes their dephosphorylation, by a receptor phosphatase that is present at high concentration on endosome membranes, and receptors then return to the plasma membrane. This helps explain the ability of cells to recover receptor-mediated signaling responsiveness very efficiently after agonist-induced desensitization. Several serpentine receptors—including the adrenoceptor if it is persistently activated—instead traffic to lysosomes after endocytosis and are degraded. This process effectively attenuates (rather than restores) cellular responsiveness, similar to the process of down-regulation described above for the epidermal growth factor receptor. Thus, depending on the particular receptor and duration of activation, endocytosis can contribute to either rapid recovery or prolonged attenuation of cellular responsiveness (Figure 2–12).

Well-Established Second Messengers

Cyclic Adenosine Monophosphate (cAMP)

Acting as an intracellular second messenger, cAMP mediates such hormonal responses as the mobilization of stored energy (the breakdown of carbohydrates in liver or triglycerides in fat cells stimulated by -adrenomimetic catecholamines), conservation of water by the kidney (mediated by vasopressin), Ca²⁺ homeostasis (regulated by parathyroid hormone), and increased rate and contractile force of heart muscle (-adrenomimetic catecholamines). It also regulates the production of adrenal and sex steroids (in response to corticotropin or follicle-stimulating hormone), relaxation of smooth muscle, and many other endocrine and neural processes.

cAMP exerts most of its effects by stimulating cAMP-dependent protein kinases (Figure 2–13). These kinases are composed of a cAMP-binding regulatory (R) dimer and two catalytic (C) chains. When cAMP binds to the R dimer, active C chains are released to diffuse through the cytoplasm and nucleus, where they transfer phosphate from ATP to appropriate substrate proteins, often enzymes. The specificity of the regulatory effects of cAMP resides in the distinct protein substrates of the kinases that are expressed in different cells. For example, liver is rich in phosphorylase kinase and glycogen synthase, enzymes whose reciprocal regulation by cAMP-dependent phosphorylation governs carbohydrate storage and release.

When the hormonal stimulus stops, the intracellular actions of cAMP are terminated by an elaborate series of enzymes. cAMP-stimulated phosphorylation of enzyme substrates is rapidly reversed by a diverse group of specific and nonspecific phosphatases. cAMP itself is degraded to 5'-AMP by several cyclic nucleotide phosphodiesterases (PDE; Figure 2–13). Competitive inhibition of cAMP degradation is one way caffeine, theophylline, and other methylxanthines produce their effects (see Chapter 20).

Calcium and Phosphoinositides

Another well-studied second messenger system involves hormonal stimulation of phosphoinositide hydrolysis (Figure 2–14). Some of the hormones, neurotransmitters, and growth factors that trigger this pathway bind to receptors linked to G proteins, whereas others bind to receptor tyrosine kinases. In all cases, the crucial step is stimulation of a membrane enzyme, phospholipase C (PLC), which splits a minor phospholipid component of the plasma membrane, phosphatidylinositol-4,5-bisphosphate (PIP₂), into two second messengers, diacylglycerol (DAG) and inositol-1,4,5-trisphosphate (IP₃ or InsP₃). Diacylglycerol is confined to the membrane, where it activates a phospholipid- and calcium-sensitive protein kinase called protein kinase C. IP₃ is water-soluble and diffuses through the cytoplasm to trigger release of Ca²⁺ by binding to ligand-gated calcium channels in the limiting membranes of internal storage vesicles. Elevated cytoplasmic Ca²⁺ concentration resulting from IP₃-promoted opening of these channels promotes the binding of Ca²⁺ to the calcium-binding protein calmodulin, which regulates activities of other enzymes, including calcium-dependent protein kinases.

With its multiple second messengers and protein kinases, the phosphoinositide signaling pathway is much more complex than the cAMP pathway. For example, different cell types may contain one or more specialized calcium- and calmodulin-dependent kinases with limited substrate specificity (eg, myosin light-chain kinase) in addition to a general calcium- and calmodulin-dependent kinase that can phosphorylate a wide variety of protein substrates. Furthermore, at least nine structurally distinct types of protein kinase C have been identified.

As in the cAMP system, multiple mechanisms damp or terminate signaling by this pathway. IP₃ is inactivated by dephosphorylation; diacylglycerol is either phosphorylated to yield phosphatidic acid, which is then converted back into phospholipids, or it is deacylated to yield arachidonic acid; Ca²⁺ is actively removed from the cytoplasm by Ca²⁺ pumps.

These and other nonreceptor elements of the calcium-phosphoinositide signaling pathway are of considerable importance in pharmacotherapy. For example, lithium ion, used in treatment of bipolar (manic-depressive) disorder, affects the cellular metabolism of phosphoinositides (see Chapter 29).

Cyclic Guanosine Monophosphate (cGMP)

Unlike cAMP, the ubiquitous and versatile carrier of diverse messages, cGMP has established signaling roles in only a few cell types. In intestinal mucosa and vascular smooth muscle, the cGMP-based signal transduction mechanism closely parallels the cAMP-mediated signaling mechanism. Ligands detected by cell-surface receptors stimulate membrane-bound guanylyl cyclase to produce cGMP, and cGMP acts by stimulating a cGMP-dependent protein kinase. The actions of cGMP in these cells are terminated by enzymatic degradation of the cyclic nucleotide and by dephosphorylation of kinase substrates.

Increased cGMP concentration causes relaxation of vascular smooth muscle by a kinase-mediated mechanism that results in dephosphorylation of myosin light chains (see Figure 12–2). In these smooth muscle cells, cGMP synthesis can be elevated by two transmembrane signaling mechanisms utilizing two different guanylyl cyclases. Atrial natriuretic peptide, a blood-borne peptide hormone, stimulates a transmembrane receptor by binding to its extracellular domain, thereby activating the guanylyl cyclase activity that resides in the receptor's intracellular domain. The other mechanism mediates responses to nitric oxide (NO; see Chapter 19), which is generated in vascular endothelial cells in response to natural vasodilator agents such as acetylcholine and histamine. After entering the target cell, nitric oxide binds to and activates a cytoplasmic guanylyl cyclase (see Figure 19–2). A number of useful vasodilating drugs, such as nitroglycerin and sodium nitroprusside used in treating cardiac ischemia and acute hypertension, act by generating or mimicking nitric oxide. Other drugs produce vasodilation by inhibiting specific phosphodiesterases, thereby interfering with the metabolic breakdown of cGMP. One such drug is sildenafil, used in treating erectile dysfunction (see Chapter 12).

Interplay among Signaling Mechanisms

The calcium-phosphoinositide and cAMP signaling pathways oppose one another in some cells and are complementary in others. For example, vasopressor agents that contract smooth muscle act by IP₃-mediated mobilization of Ca²⁺, whereas agents that relax smooth muscle often act by elevation of cAMP. In contrast, cAMP and phosphoinositide second messengers act together to stimulate glucose release from the liver.

Phosphorylation: A Common Theme

Almost all second messenger signaling involves reversible phosphorylation, which performs two principal functions in signaling: amplification and flexible regulation. In amplification, rather like GTP bound to a G protein, the attachment of a phosphoryl group to a serine, threonine, or tyrosine residue powerfully amplifies the initial regulatory signal by recording a molecular memory that the pathway has been activated; dephosphorylation erases the memory, taking a longer time to do so than is required for dissociation of an allosteric ligand. In flexible regulation, differing substrate specificities of the multiple protein kinases regulated by second messengers provide branch points in signaling pathways that may be independently regulated. In this way, cAMP, Ca²⁺, or other second messengers can use the presence or absence of particular kinases or kinase substrates to produce quite different effects in different cell types. Inhibitors of protein kinases have great potential as therapeutic agents, particularly in neoplastic diseases. Trastuzumab, an antibody that antagonizes growth factor receptor signaling (discussed earlier), is a useful therapeutic agent for breast cancer. Another example of this general approach is imatinib, a small molecule inhibitor of the cytoplasmic tyrosine kinase Abl, which is activated by growth factor signaling pathways. Imatinib appears to be very effective for treating chronic myelogenous leukemia, which is caused by a chromosomal translocation event that produces an active Bcr/Abl fusion protein in hematopoietic cells.

Receptor Classes & Drug Development

The existence of a specific drug receptor is usually inferred from studying the structure-activity relationship of a group of structurally similar congeners of the drug that mimic or antagonize its effects. Thus, if a series of related agonists exhibits identical relative potencies in producing two distinct effects, it is likely that the two effects are mediated by similar or identical receptor molecules. In addition, if identical receptors mediate both effects, a competitive antagonist will inhibit both responses with the same K_i; a second competitive antagonist will inhibit both responses with its own characteristic K_i. Thus, studies of the relation between structure and activity of a series of agonists and antagonists can identify a species of receptor that mediates a set of pharmacologic responses.

Exactly the same experimental procedure can show that observed effects of a drug are mediated by different receptors. In this case, effects mediated by different receptors may exhibit different orders of potency among agonists and different K_i values for each competitive antagonist.

Wherever we look, evolution has created many different receptors that function to mediate responses to any individual chemical signal. In some cases, the same chemical acts on completely different structural receptor classes. For example, acetylcholine uses ligand-gated ion channels (nicotinic AChRs) to initiate a fast excitatory postsynaptic potential (EPSP) in postganglionic neurons. Acetylcholine also activates a separate class of G protein-coupled receptors (muscarinic AChRs), which modulate responsiveness of the same neurons to the fast EPSP. In addition, each structural class usually includes multiple subtypes of receptor, often with significantly different signaling or regulatory properties. For example, many biogenic amines (eg, norepinephrine, acetylcholine, and serotonin) activate more than one receptor, each of which may activate a different G protein, as previously described (see also Table 2–1). The existence of many receptor classes and subtypes for the same endogenous ligand has created important opportunities for drug development. For example, propranolol, a selective antagonist of adrenoceptors, can reduce an accelerated heart rate without preventing the sympathetic nervous system from causing vasoconstriction, an effect mediated by ₁ receptors.

The principle of drug selectivity may even apply to structurally identical receptors expressed in different cells, eg, receptors for steroids such as estrogen (Figure 2–6). Different cell types express different accessory proteins, which interact with steroid receptors and change the functional effects of drug-receptor interaction. For example, tamoxifen acts as an antagonist on estrogen receptors expressed in mammary tissue but as an agonist on estrogen receptors in bone. Consequently, tamoxifen may be useful not only in the treatment and prophylaxis of breast cancer but also in the prevention of osteoporosis by increasing bone density (see Chapters 40 and 42). Tamoxifen may also create complications in postmenopausal women, however, by exerting an agonist action in the uterus, stimulating endometrial cell proliferation.

New drug development is not confined to agents that act on receptors for extracellular chemical signals. Pharmaceutical chemists are now determining whether elements of signaling pathways distal to the receptors may also serve as targets of selective and useful drugs. For example, clinically useful agents might be developed that act selectively on specific G proteins, kinases, phosphatases, or the enzymes that degrade second messengers.

Relation between Drug Dose & Clinical Response

We have dealt with receptors as molecules and shown how receptors can quantitatively account for the relation between dose or concentration of a drug and pharmacologic responses, at least in an idealized system. When faced with a patient who needs treatment, the prescriber must make a choice among a variety of possible drugs and devise a dosage regimen that is likely to produce maximal benefit and minimal toxicity. To make rational therapeutic decisions, the prescriber must understand how drug-receptor interactions underlie the relations between dose and response in patients, the nature and causes of variation in pharmacologic responsiveness, and the clinical implications of selectivity of drug action.

Dose & Response in Patients

Graded Dose-Response Relations

To choose among drugs and to determine appropriate doses of a drug, the prescriber must know the relative pharmacologic potency and maximal efficacy of the drugs in relation to the desired therapeutic effect. These two important terms, often confusing to students and clinicians, can be explained by referring to Figure 2–15, which depicts graded dose-response curves that relate the dose of four different drugs to the magnitude of a particular therapeutic effect.

Potency

Drugs A and B are said to be more potent than drugs C and D because of the relative positions of their dose-response curves along the dose axis of Figure 2–15. Potency refers to the concentration (EC₅₀) or dose (ED₅₀) of a drug required to produce 50% of that drug's maximal effect. Thus, the pharmacologic potency of drug A in Figure 2–15 is less than that of drug B, a partial agonist because the EC₅₀ of A is greater than the EC₅₀ of B. Potency of a drug depends in part on the affinity (K_d) of receptors for binding the drug and in part on the efficiency with which drug-receptor interaction is coupled to response. Note that some doses of drug A can produce larger effects than any dose of drug B, despite the fact that we describe drug B as pharmacologically more potent. The reason for this is that drug A has a larger maximal efficacy (as described below).

For clinical use, it is important to distinguish between a drug's potency and its efficacy. The clinical effectiveness of a drug depends not on its potency (EC₅₀), but on its maximal efficacy (see below) and its ability to reach the relevant receptors. This ability can depend on its route of administration, absorption, distribution through the body, and clearance from the blood or site of action. In deciding which of two drugs to administer to a patient, the prescriber must usually consider their relative effectiveness rather than their relative potency. Pharmacologic potency can largely determine the administered dose of the chosen drug.

For therapeutic purposes, the potency of a drug should be stated in dosage units, usually in terms of a particular therapeutic end point (eg, 50 mg for mild sedation, 1 mcg/kg/min for an increase in heart rate of 25 bpm). Relative potency, the ratio of equi-effective doses (0.2, 10, etc), may be used in comparing one drug with another.

Maximal Efficacy

This parameter reflects the limit of the dose-response relation on the response axis. Drugs A, C, and D in Figure 2–15 have equal maximal efficacy, whereas all have greater maximal efficacy than drug B. The maximal efficacy (sometimes referred to simply as efficacy) of a drug is obviously crucial for making clinical decisions when a large response is needed. It may be determined by the drug's mode of interactions with receptors (as with partial agonists* or by characteristics of the receptor-effector system involved.

Thus, diuretics that act on one portion of the nephron may produce much greater excretion of fluid and electrolytes than diuretics that act elsewhere. In addition, the practical efficacy of a drug for achieving a therapeutic end point (eg, increased cardiac contractility) may be limited by the drug's propensity to cause a toxic effect (eg, fatal cardiac arrhythmia) even if the drug could otherwise produce a greater therapeutic effect.

*Note that "maximal efficacy," used in a therapeutic context, does not have exactly the same meaning that the term denotes in the more specialized context of drug-receptor interactions described earlier in this chapter. In an idealized in vitro system, efficacy denotes the relative maximal efficacy of agonists and partial agonists that act via the same receptor. In therapeutics, efficacy denotes the extent or degree of an effect that can be achieved in the intact patient. Thus, therapeutic efficacy may be affected by the characteristics of a particular drug-receptor interaction, but it also depends on a host of other factors as noted in the text.

Shape of Dose-Response Curves

Although the responses depicted in curves A, B, and C of Figure 2–15 approximate the shape of a simple Michaelis-Menten relation (transformed to a logarithmic plot), some clinical responses do not. Extremely steep dose-response curves (eg, curve D) may have important clinical consequences if the upper portion of the curve represents an undesirable extent of response (eg, coma caused by a sedative-hypnotic). Steep dose-response curves in patients can result from cooperative interactions of several different actions of a drug (eg, effects on brain, heart, and peripheral vessels, all contributing to lowering of blood pressure).

Quantal Dose-Effect Curves

Graded dose-response curves of the sort described above have certain limitations in their application to clinical decision making. For example, such curves may be impossible to construct if the pharmacologic response is an either-or (quantal) event, such as prevention of convulsions, arrhythmia, or death. Furthermore, the clinical relevance of a quantitative dose-response relation in a single patient, no matter how precisely defined, may be limited in application to other patients, owing to the great potential variability among patients in severity of disease and responsiveness to drugs.

Some of these difficulties may be avoided by determining the dose of drug required to produce a specified magnitude of effect in a large number of individual patients or experimental animals and plotting the cumulative frequency distribution of responders versus the log dose (Figure 2–16). The specified quantal effect may be chosen on the basis of clinical relevance (eg, relief of headache) or for preservation of safety of experimental subjects (eg, using low doses of a cardiac stimulant and specifying an increase in heart rate of 20 bpm as the quantal effect), or it may be an inherently quantal event (eg, death of an experimental animal). For most drugs, the doses required to produce a specified quantal effect in individuals are lognormally distributed; that is, a frequency distribution of such responses plotted against the log of the dose produces a gaussian normal curve of variation (colored areas, Figure 2–16). When these responses are summated, the resulting cumulative frequency distribution constitutes a quantal dose-effect curve (or dose-percent curve) of the proportion or percentage of individuals who exhibit the effect plotted as a function of log dose.

The quantal dose-effect curve is often characterized by stating the median effective dose (ED₅₀), which is the dose at which 50% of individuals exhibit the specified quantal effect. (Note that the abbreviation ED₅₀ has a different meaning in this context from its meaning in relation to graded dose-effect curves, described in previous text). Similarly, the dose required to produce a particular toxic effect in 50% of animals is called the median toxic dose (TD₅₀). If the toxic effect is death of the animal, a median lethal dose (LD₅₀) may be experimentally defined. Such values provide a convenient way of comparing the potencies of drugs in experimental and clinical settings: Thus, if the ED₅₀s of two drugs for producing a specified quantal effect are 5 and 500 mg, respectively, then the first drug can be said to be 100 times more potent than the second for that particular effect. Similarly, one can obtain a valuable index of the selectivity of a drug's action by comparing its ED₅₀s for two different quantal effects in a population (eg, cough suppression versus sedation for opioid drugs).

Quantal dose-effect curves may also be used to generate information regarding the margin of safety to be expected from a particular drug used to produce a specified effect. One measure, which relates the dose of a drug required to produce a desired effect to that which produces an undesired effect, is the therapeutic index. In animal studies, the therapeutic index is usually defined as the ratio of the TD₅₀ to the ED₅₀ for some therapeutically relevant effect. The precision possible in animal experiments may make it useful to use such a therapeutic index to estimate the potential benefit of a drug in humans. Of course, the therapeutic index of a drug in humans is almost never known with real precision; instead, drug trials and accumulated clinical experience often reveal a range of usually effective doses and a different (but sometimes overlapping) range of possibly toxic doses. The clinically acceptable risk of toxicity depends critically on the severity of the disease being treated. For example, the dose range that provides relief from an ordinary headache in the majority of patients should be very much lower than the dose range that produces serious toxicity, even if the toxicity occurs in a small minority of patients. However, for treatment of a lethal disease such as Hodgkin's lymphoma, the acceptable difference between therapeutic and toxic doses may be smaller.

Finally, note that the quantal dose-effect curve and the graded dose-response curve summarize somewhat different sets of information, although both appear sigmoid in shape on a semilogarithmic plot (compare Figures 2–15 and 2–16). Critical information required for making rational therapeutic decisions can be obtained from each type of curve. Both curves provide information regarding the potency and selectivity of drugs; the graded dose-response curve indicates the maximal efficacy of a drug, and the quantal dose-effect curve indicates the potential variability of responsiveness among individuals.

Variation in Drug Responsiveness

Individuals may vary considerably in their response to a drug; indeed, a single individual may respond differently to the same drug at different times during the course of treatment. Occasionally, individuals exhibit an unusual or idiosyncratic drug response, one that is infrequently observed in most patients. The idiosyncratic responses are usually caused by genetic differences in metabolism of the drug or by immunologic mechanisms, including allergic reactions.

Quantitative variations in drug response are in general more common and more clinically important. An individual patient is hyporeactive  or hyperreactive  to a drug in that the intensity of effect of a given dose of drug is diminished or increased compared with the effect seen in most individuals. (Note:  The term hypersensitivity usually refers to allergic or other immunologic responses to drugs.) With some drugs, the intensity of response to a given dose may change during the course of therapy; in these cases, responsiveness usually decreases as a consequence of continued drug administration, producing a state of relative tolerance to the drug's effects. When responsiveness diminishes rapidly after administration of a drug, the response is said to be subject to tachyphylaxis.

Even before administering the first dose of a drug, the prescriber should consider factors that may help in predicting the direction and extent of possible variations in responsiveness. These include the propensity of a particular drug to produce tolerance or tachyphylaxis as well as the effects of age, sex, body size, disease state, genetic factors, and simultaneous administration of other drugs.

Four general mechanisms may contribute to variation in drug responsiveness among patients or within an individual patient at different times.

Alteration in Concentration of Drug that Reaches the Receptor

Patients may differ in the rate of absorption of a drug, in distributing it through body compartments, or in clearing the drug from the blood (see Chapter 3). By altering the concentration of drug that reaches relevant receptors, such pharmacokinetic differences may alter the clinical response. Some differences can be predicted on the basis of age, weight, sex, disease state, and liver and kidney function, and by testing specifically for genetic differences that may result from inheritance of a functionally distinctive complement of drug-metabolizing enzymes (see Chapters 3 and 4). Another important mechanism influencing drug availability is active transport of drug from the cytoplasm, mediated by a family of membrane transporters encoded by the so-called multidrug resistance (MDR) genes. For example, up-regulation of MDR gene–encoded transporter expression is a major mechanism by which tumor cells develop resistance to anticancer drugs.

Variation in Concentration of an Endogenous Receptor Ligand

This mechanism contributes greatly to variability in responses to pharmacologic antagonists. Thus, propranolol, a -adrenoceptor antagonist, markedly slows the heart rate of a patient whose endogenous catecholamines are elevated (as in pheochromocytoma) but does not affect the resting heart rate of a well-trained marathon runner. A partial agonist may exhibit even more dramatically different responses: Saralasin, a weak partial agonist at angiotensin II receptors, lowers blood pressure in patients with hypertension caused by increased angiotensin II production and raises blood pressure in patients who produce normal amounts of angiotensin.

Alterations in Number or Function of Receptors

Experimental studies have documented changes in drug response caused by increases or decreases in the number of receptor sites or by alterations in the efficiency of coupling of receptors to distal effector mechanisms. In some cases, the change in receptor number is caused by other hormones; for example, thyroid hormones increase both the number of receptors in rat heart muscle and cardiac sensitivity to catecholamines. Similar changes probably contribute to the tachycardia of thyrotoxicosis in patients and may account for the usefulness of propranolol, a -adrenoceptor antagonist, in ameliorating symptoms of this disease.

In other cases, the agonist ligand itself induces a decrease in the number (eg, down-regulation) or coupling efficiency (eg, desensitization) of its receptors. These mechanisms (discussed previously under Signaling Mechanisms & Drug Actions) may contribute to two clinically important phenomena: first, tachyphylaxis or tolerance to the effects of some drugs (eg, biogenic amines and their congeners), and second, the "overshoot" phenomena that follow withdrawal of certain drugs. These phenomena can occur with either agonists or antagonists. An antagonist may increase the number of receptors in a critical cell or tissue by preventing down-regulation caused by an endogenous agonist. When the antagonist is withdrawn, the elevated number of receptors can produce an exaggerated response to physiologic concentrations of agonist. Potentially disastrous withdrawal symptoms can result for the opposite reason when administration of an agonist drug is discontinued. In this situation, the number of receptors, which has been decreased by drug-induced down-regulation, is too low for endogenous agonist to produce effective stimulation. For example, the withdrawal of clonidine (a drug whose ₂-adrenoceptor agonist activity reduces blood pressure) can produce hypertensive crisis, probably because the drug down-regulates ₂ adrenoceptors (see Chapter 11).

Genetic factors also can play an important role in altering the number or function of specific receptors. For example, a specific genetic variant of the _2C adrenoceptor—when inherited together with a specific variant of the ₁ adrenoceptor—confers increased risk for developing heart failure, which may be reduced by early intervention using antagonist drugs. The identification of such genetic factors, part of the rapidly developing field of pharmacogenetics, holds promise for clinical diagnosis and in the future may help physicians design the most appropriate pharmacologic therapy for individual patients.

Another interesting example of genetic determination of effects on drug response is seen in the treatment of cancers involving excessive growth factor signaling. Somatic mutations affecting the tyrosine kinase domain of the epidermal growth factor receptor confer enhanced sensitivity to kinase inhibitors such as gefitinib in certain lung cancers. This effect enhances the antineoplastic effect of the drug and, because the somatic mutation is specific to the tumor and not present in the host, the therapeutic index of these drugs can be significantly enhanced in patients whose tumors harbor such mutations.

Changes in Components of Response Distal to the Receptor

Although a drug initiates its actions by binding to receptors, the response observed in a patient depends on the functional integrity of biochemical processes in the responding cell and physiologic regulation by interacting organ systems. Clinically, changes in these postreceptor processes represent the largest and most important class of mechanisms that cause variation in responsiveness to drug therapy.

Before initiating therapy with a drug, the prescriber should be aware of patient characteristics that may limit the clinical response. These characteristics include the age and general health of the patient and—most importantly—the severity and pathophysiologic mechanism of the disease. The most important potential cause of failure to achieve a satisfactory response is that the diagnosis is wrong or physiologically incomplete. Drug therapy is always most successful when it is accurately directed at the pathophysiologic mechanism responsible for the disease.

When the diagnosis is correct and the drug is appropriate, an unsatisfactory therapeutic response can often be traced to compensatory mechanisms in the patient that respond to and oppose the beneficial effects of the drug. Compensatory increases in sympathetic nervous tone and fluid retention by the kidney, for example, can contribute to tolerance to antihypertensive effects of a vasodilator drug. In such cases, additional drugs may be required to achieve a useful therapeutic result.

Clinical Selectivity: Beneficial versus Toxic Effects of Drugs

Although we classify drugs according to their principal actions, it is clear that no drugcauses only a single, specific effect. Why is this so? It is exceedingly unlikely that any kind of drug molecule will bind to only a single type of receptor molecule, if only because the number of potential receptors in every patient is astronomically large. Even if the chemical structure of a drug allowed it to bind to only one kind of receptor, the biochemical processes controlled by such receptors would take place in many cell types and would be coupled to many other biochemical functions; as a result, the patient and the prescriber would probably perceive more than one drug effect. Accordingly, drugs are only selective—rather than specific—in their actions, because they bind to one or a few types of receptor more tightly than to others and because these receptors control discrete processes that result in distinct effects.

It is only because of their selectivity that drugs are useful in clinical medicine. Selectivity can be measured by comparing binding affinities of a drug to different receptors or by comparing ED₅₀s for different effects of a drug in vivo. In drug development and in clinical medicine, selectivity is usually considered by separating effects into two categories: beneficial or therapeutic effects versus toxic effects. Pharmaceutical advertisements and prescribers occasionally use the term side effect, implying that the effect in question is insignificant or occurs via a pathway that is to one side of the principal action of the drug; such implications are frequently erroneous.

Beneficial and Toxic Effects Mediated By the Same Receptor-Effector Mechanism

Much of the serious drug toxicity in clinical practice represents a direct pharmacologic extension of the therapeutic actions of the drug. In some of these cases (eg, bleeding caused by anticoagulant therapy; hypoglycemic coma due to insulin), toxicity may be avoided by judicious management of the dose of drug administered, guided by careful monitoring of effect (measurements of blood coagulation or serum glucose) and aided by ancillary measures (avoiding tissue trauma that may lead to hemorrhage; regulation of carbohydrate intake). In still other cases, the toxicity may be avoided by not administering the drug at all, if the therapeutic indication is weak or if other therapy is available.

In certain situations, a drug is clearly necessary and beneficial but produces unacceptable toxicity when given in doses that produce optimal benefit. In such situations, it may be necessary to add another drug to the treatment regimen. In treating hypertension, for example, administration of a second drug often allows the prescriber to reduce the dose and toxicity of the first drug (see Chapter 11).

Beneficial and Toxic Effects Mediated by Identical Receptors But in Different Tissues or by Different Effector Pathways

Many drugs produce both their desired effects and adverse effects by acting on a single receptor type in different tissues. Examples discussed in this book include: digitalis glycosides, which act by inhibiting Na⁺,K⁺ ATPase in cell membranes; methotrexate, which inhibits the enzyme dihydrofolate reductase; and glucocorticoid hormones.

Three therapeutic strategies are used to avoid or mitigate this sort of toxicity. First, the drug should always be administered at the lowest dose that produces acceptable benefit. Second, adjunctive drugs that act through different receptor mechanisms and produce different toxicities may allow lowering the dose of the first drug, thus limiting its toxicity (eg, use of other immunosuppressive agents added to glucocorticoids in treating inflammatory disorders). Third, selectivity of the drug's actions may be increased by manipulating the concentrations of drug available to receptors in different parts of the body, for example, by aerosol administration of a glucocorticoid to the bronchi in asthma.

Beneficial and Toxic Effects Mediated by Different Types of Receptors

Therapeutic advantages resulting from new chemical entities with improved receptor selectivity were mentioned earlier in this chapter and are described in detail in later chapters. Such drugs include - and -selective adrenoceptor agonists and antagonists, H₁ and H₂ antihistamines, nicotinic and muscarinic blocking agents, and receptor-selective steroid hormones. All these receptors are grouped in functional families, each responsive to a small class of endogenous agonists. The receptors and their associated therapeutic uses were discovered by analyzing effects of the physiologic chemical signals—catecholamines, histamine, acetylcholine, and corticosteroids.

Several other drugs were discovered by exploiting therapeutic or toxic effects of chemically similar agents observed in a clinical context. Examples include quinidine, the sulfonylureas, thiazide diuretics, tricyclic antidepressants, opioid drugs, and phenothiazine antipsychotics. Often the new agents turn out to interact with receptors for endogenous substances (eg, opioids and phenothiazines for endogenous opioid and dopamine receptors, respectively). It is likely that other new drugs will be found to do so in the future, perhaps leading to the discovery of new classes of receptors and endogenous ligands for future drug development.

Thus, the propensity of drugs to bind to different classes of receptor sites is not only a potentially vexing problem in treating patients, it also presents a continuing challenge to pharmacology and an opportunity for developing new and more useful drugs.

References