Prepared by: Abraham Daniel C. Cruz, MD Instructor A, Department of Pharmacology, FEU-NRMF Inst.

of Medicine MS (Candidate) Pharmacology, UP Manila College of Medicine (1 Reference Basic and Clinical Pharmacology by Katzung)

By the end of the lecture, the student should be able to:

Understand basic principles of receptor pharmacology and types of drug-receptor interactions (agonist and antagonist) Correlate drug dose and biologic response using the graded and quantal dose-response curves Describe the different receptor types and the specific mechanisms of signalling and regulation that result in a biologic response Describe the different factors that cause variations in drug response

Receptors
component of a cell or organism that interacts with a

drug and initiates the chain of events leading to the drugs observed effects Focus of drug effects and mechanisms of action Applicable in:
endocrinology, immunology, molecular biology to explain biologic regulation Drug development and clinical decision making

Key to understanding drug action and clinical uses

largely determine the quantitative relations between dose or concentration of drug and pharmacologic effects
affinity for drug binding determines the concentration required to

form a significant number of drug-receptor complexes total number of receptors may limit the maximal effect of a drug

responsible for selectivity of drug action

molecular size, shape, and electrical charge of a drug - determine

whetherand with what affinityit will bind to a particular receptor changes in the chemical structure of a drug can increase or decrease a new drug's affinities for different classes of receptors alterations in therapeutic and toxic effects

mediate both pharmacologic agonist and antagonist action

Agonists - activate the receptor to signal as a direct result of binding

to it or through indirect means

Direct bind to receptors directly Indirect

Increase or decrease concentration of

endogenous ligands May bind to:

Enzymes (synthesis or metabolism) Transport proteins

Pharmacologic antagonists
Bind to receptors but do not activate the

generation of a signal interfere with agonist to activate the receptor

Prevent agonist binding Suppress basal signalling (constitutive) activity of receptors

Proteins/polypeptides diverse; specific shape and electrical charge Identification process

Old drug binding purify and identify receptors

from tissue extracts New molecular biology and gene sequencing predict structure or sequence homology to other known receptors (structure activity relati0nship) drug development
Discoveries
Many drugs bind to receptors other than previously known orphan receptors no known ligands; target of research

Types of drug receptors

Regulatory proteins (mediate action of

endogenous chemical signals) Enzymes Transport proteins (ion channels) Structural proteins (tubulin)

Determinants of the quantitative relation between drug concentration and pharmacologic response Regulatory proteins/components for cell signaling mechanisms drug targets Determinant of therapeutic and toxic effects in patients

Hyperbolic
Low dose response

increment increases in direct proportion to dose (linear) Increasing doses response increment diminishes Very high doses no further increase in response

E = effect observed at concentration C Emax = maximal response that can be produced by the drug EC50 - concentration of drug that produces 50% of maximal effect

Hyperbolic action resembles the mass action law (association between two molecules [agonist + receptor] of a given affinity)

B = drug bound to receptors C = free (unbound) drug B max = total number of receptor sites; sites bound to the drug at infinitely high drug concentrations Kd = dissociation constant

HYPERBOLIC CURVE

SIGMOID CURVE AGONIST/DRUG CONCENTRATION IN LOGARITHMIC SCALE

Coupling transduction process that links receptor occupancy and pharmacologic response Determinants of coupling efficiency:

Initial conformational change (based on structure

activity relationship)
full agonists more efficiently coupled compared to partial agonists

Signal transduction
biochemical events that transduce receptor occupancy to a response
Re== relation to number of receptors bound; example Ion channels Non-linear biologic response increased disproportionately to number of receptors bound receptors linked to enzymatic signal transduction cascades

One factor for non-linear occupancy-response coupling Maximal biologic response at agonist concentration that does not result in full occupancy of receptors Spareness

Temporal - Ex. G protein-coupled receptors and second messengers

Elicits response AFTER drug is no longer bound to receptors

Number
Affinity of agonist to receptor (Kd, dissociation constant) Degree of spareness total number of receptors present compared to the number required to elicit a maximal biologic response

Allows for precise evaluation of the effect of drug dosage without considering the biochemical details of the signaling response The Kd of the agonist-receptor interaction determines the fraction of total receptors (B/Bmax) that will be occupied at a given concentration (C) of agonist (regardless of receptor concentration)

Example
One cell, 4 receptors (no spare receptors), 4

effectors
Half maximal response is elicited when an agonist binds 2 receptors (50% of receptors)

One cell, 40 receptors, 4 effectors Half maximal response is elicited when an agonist binds 2 receptors (5% of receptors) Therefore: lower agonist concentration is required to reach half maximal response increased tissue sensitivity

Receptor Antagonists
Pharmacologic antagonists Bind to receptors but do not activate them Prevent agonists (drugs or endogenous molecules) from activating receptors inverse agonists reduce receptor activity

below basal levels; (-) bound ligand

binds to the same receptor as an agonist but induces a pharmacological response opposite to that agonist

Reversible vs. irreversible competitive antagonists

BOND TYPE van der Waals

MECHANISM

BOND STRENGTH

Shifting electron density in areas of a molecule, or in a + molecule as a whole, results in the generation of transient positive or negative charges. These areas interact with transient areas of opposite charge on another molecule. Hydrogen atoms bound to nitrogen or oxygen become more positively polarized, allowing them to bond to more negatively polarized atoms such as oxygen, nitrogen, or sulfur. ++

Hydrogen

Ionic

Atoms with an excess of electrons (imparting an overall negative charge on the atom) are attracted to atoms with a deficiency of electrons (imparting an overall positive charge on the atom).
Two bonding atoms share electrons.

+++

Covalent

++++

In the presence of a fixed concentration of agonist

increasing antagonist concentration progressively inhibit

the agonist response

Effects can be surmounted by sufficiently high agonist concentrations Emax remains the same for any fixed concentration of antagonist Increases agonist concentration required for a given degree of response

Shifts concentration-effect curve to the right

Schild Equation

Used primarily to determine Ki (dissociation

constant)

Therapeutic implications
degree of inhibition produced by a competitive antagonist

depends on the concentration of antagonist (ex. Interindividual variation in drug clearance)

Clinical response to a competitive antagonist depends on

the concentration of agonist that is competing for binding to receptors

Ex. Beta-adrenoreceptor blockers vs. norepinephrine (endogenous) blockade may be overcome in situations that increase NE (exercise, postural changes, stress)

(or nearly irreversible) covalent bond or tight binding to receptor unavailable for agonist binding Remaining unoccupied receptors are too low to elicit a response despite high agonist levels
EXCEPTION: presence of spare receptors (but

requires higher agonist doses)

need not be in the unbound form to elicit a response once bound to receptors Duration of action is dependent on the rate of turnover of receptor molecules and not its elimination rate Advantage: prevent responses to varying high and low agonist concentrations Disadvantage: if overdose occurs, a physiologic antagonist must be given (acts on another receptor but elicits the opposite response)

Bind to a site on the receptor protein separate from the agonist binding site
Prevent receptor activation WITHOUT blocking

agonist binding Actions are reversible if anatagonists do not bind covalently

Bind on a separate site on the receptor protein and alter receptor function without inactivating receptor Example: benzodiazepines and GABAA receptor enhance the net activating effect of GABA on channel conductance

produce a lower response at full receptor occupancy

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 Failure to produce a maximal response is not due to decreased affinity for binding to receptors (even at high concentrations that saturate binding to all receptors) competitively inhibit the responses produced by full agonists (Figure 24C)

many drugs used clinically as antagonists are in fact weak partial agonists

Chemical antagonism
Does not involve receptors; chemical interactions between

two substances (ionic binding, etc.) that render one of the drugs unavailable for receptor binding (ex. Protamine + Heparin)

Physiologic antagonism
Antagonism between endogenous regulatory pathways

mediated by different receptors Effects are less specific and less easy to control Ex. Glucocorticoids hyperglycemia; insulin hypoglycemia

Ligands steroids (glucocorticoids, mineralocorticoids, sex steroids, vitamin D) and thyroid hormones Stimulate transcription of genes by binding to specific target DNA sequences (response elements)

Therapeutic consequences
Effects are produced after a lag period of 30

minutes to several hours (time required for protein synthesis) effects are NOT immediate Effects persist for hours to days after agonist concentration has been reduced to zero due to slow turnover of newly synthesized proteins

Ligands - insulin, epidermal growth factor (EGF), platelet-derived growth factor (PDGF), atrial natriuretic peptide (ANP), transforming growth factor-alpha (TGF-alpha ), and many other trophic hormones

down-regulation
Limits intensity and duration of action ligands Occurs by accelerated endocytosis and eventual

degradation of receptors after ligand binding

Occurs faster than de novo synthesis of receptors decreased effector cell responsiveness

Similar to tyrosine kinases Different phosphorylation system JAK-STAT

Janus kinase family Signal transducers and activators of transcription

Ligands interleukins, luekotrienes, GH, prolactin

Natural ligands synaptic neurotransmitters

Ach, 5-HT, GABA, Glutamate

Increase transmembrane conductance of a relevant ion alters electrical potential across the membrane May be:
Directly linked G protein coupled 2nd messenger system

Regulated by phosphorylation and endocytosis

For signal amplification Effect is magnified relative to drug-receptor binding (intensity and duration) Transmembrane signaling system w/ 3 components Cell-surface receptor G-protein Changes the activity of an effector element Effector element (enzyme or ion channel) Changes the concentration of intracellular second messenger

Second messengers
cAMP, calcium ion, phosphoinositides

Example receptors
beta-adrenoceptors, glucagon receptors,

thyrotropin receptors, and certain subtypes of dopamine and serotonin receptors

7-TM receptors seven transmembrane serpentine

Receptor polypeptide chain snakes across the plasma

membrane seven times

Ligands
adrenergic amines Serotonin Acetylcholine
(muscarinic but not nicotinic)

many peptide hormones, odorants, and even visual

receptors (in retinal rod and cone cells)

Desensitization
-adrenoreceptor (example) Phosphorylation of the carboxy terminal (OH

residues) after prolonged exposure to agonist

By G protein-coupled receptor kinases (GRKs) Results in
Increased binding of -arrestin prevents receptor from binding to G protein Increased receptor endocytosis

REVERSED when agonist is removed for a few minutes

Produced by increased activity of adenylyl cyclase mediates:

mobilization of stored energy (-adrenomimetic

catecholamines) conservation of water by the kidney (vasopressin) Ca2+ homeostasis (parathyroid hormone) increased rate and contractile force of heart muscle (adrenomimetic catecholamines regulates the production of adrenal and sex steroids (in response to corticotropin or follicle-stimulating hormone) relaxation of smooth muscle endocrine and neural processes

Termination of action
After cessation of hormonal stimulus Dephosphorylation of enzyme substrates
By specific and non-specific phosphatases

Degradation of cAMP to 5'-AMP

By several cyclic nucleotide phosphodiesterases (PDE) Competitive inhibition MOA of caffeine, theophylline (methylxanthines)

Hormones, NT, growth factors G protein coupled or through tyrosine kinases Crucial step: stimulation of phospholipase C
Splits PIP2 to DAG
Confined to membrane Activates protein kinase C (phospholipid- and calcium-sensitive)

IP3
Water soluble diffuses to the cytoplasm Triggers calcium ion release from storage vesicles binds to calmodulin regulates calcium-dependent protein kinases

More complex than cAMP pathway One cell may contain multiple calcium- and calmodulin-dependent kinases Termination of action
IP3 dephosphorylation DAG Phosphorylation phosphatidic acid converted back to phospholipids Deacetylation arachidonic acid Calcium actively removed by pumps

Ligands stimulate guanylyl cyclase to produce cGMP Stimulate cGMP-dependent protein kinases Termination of action

Enzymatic degradation
Dephosphorylation of kinase substrates

Examples
Vascular smooth muscle relaxation (thru MLC

dephosphorylation) NO (from nitrates) activates guanylyl cyclase vasodilation PDE inhibitors (ex. Sidenafil) inhibit cGMP breakdown

Amplification
attachment of a phosphoryl group records a molecular

memory that the pathway has been activated dephosphorylation erases the memory takes a longer time compared ligand dissociation

Flexible regulation
branch points in signaling pathways that may be

independently regulated 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

The response to a drug is proportional to the concentration of the receptors that are bound (occupied) by the drug Two major types
1. Graded 2. Quantal

ASSUMPTION: response to a drug is proportional to the concentration of receptors that are bound by the drug response = [DR] = [D]______ max response [Ro] [D] + Kd

 Assumption: response is proportional to

receptor occupancy TWO IMPORTANT PARAMETERS: Potency (EC50) concentration at which the drug elicits 50% of its maximal response

Efficacy (Emax)- maximal response produced by the drug (also known as maximal efficacy)

Shows increasing responses of to increasing doses of a drug

Graph of the fraction of a population that shows a specified response to increasing doses of drug For evaluating an all-or-none response
Ex. Prevent seizures, relief of headache, etc.

median effective dose (ED50)

dose at which 50% of individuals exhibit the specified

quantal effect

median toxic dose (TD50)

dose required to produce a particular toxic effect in 50% of

animals median lethal dose (LD50) - if the toxic effect is death of the animal

*may also give information on the potency between 2 drugs given a specified quantal response

TI= TD50 / ED50

Lower TI, higher probability of toxicity and loss of efficacy Higher TI, less likely to cause adverse effects

idiosyncratic drug response

Unusual, infrequently observed; due to: genetic differences in metabolism of the drug immunologic mechanisms (allergic reactions)

hyporeactive or hyperreactive
intensity of effect of a given dose of drug is

diminished or increased in comparison to the effect seen in most individuals

hypersensitivity
allergic or other immunologic responses to drugs

Tolerance (relative)
intensity of response to a given dose may change during

the course of therapy decreased responsiveness due to continued drug administration

Tachyphylaxis
Rapid diminution of responsiveness after administration

Other factors that predict direction and extent of variation in response

age, sex, body size, disease state, genetic factors, and

simultaneous administration of other drugs.

Alteration of drug concentration that reaches receptors

PK changes due to age, sex, disease state, kidney

and liver function

Variation in concentration of endogenous receptor ligand Alteration in function or number of receptors Changes in components of response distal to the receptor

Pharmacokinetic vs Pharmacodynamic
Additive 1+1=2 Potentiation 1+0>1 Synergism 1+1>2 Antagonism as described above

No drug causes only a single, specific effect Drugs are only selective in their actions
bind to one or a few types of receptor more tightly

than to others these receptors control discrete processes that result in distinct effects

May be mediated by
Same receptor-effector mechanism Same receptor but different tissue or effector

mechanism Different receptor types

By the end of the lecture, the student should be able to:

Understand basic principles of receptor pharmacology and types of drug-receptor interactions (agonist and antagonist) Correlate drug dose and biologic response using the graded and quantal dose-response curves Describe the different receptor types and the specific mechanisms of signalling and regulation that result in a biologic response Describe the different factors that cause variations in drug response

Drug receptors can be divided into five groups These groups have different locations and effects See table below

Most agonist (receptor-activating) and antagonist (receptor-blocking) drugs bind to their receptors with weak, reversible bonds A few antagonists bind with strong, covalent bonds, resulting in irreversible action.

When an agonist drug is applied in increasing doses to a responsive system and the increments are recorded, a graded doseresponse curve is observed Binding of drugs to its receptors follows a similar curve

Two properties derived from this curve

EC50 or ED50 Concentration or dose for half-maximal effect Measure of potency of the drug and its affinity for its receptor Emax maximum effect Measure of the maximum response that can be expected from the drug in this system Kd and Bmax corresponding measures for

concentration-binding plots

When a specific intensity or drug response is defined and the dose required to produce that intensity of response are measured in a large population of subjects, biologic variation results in a spread of these doses over a range

The defined response may be a therapeutic or a toxic effect. Comparison of the median dose to produce a toxic effect versus the median dose to produce a therapeutic effect may be carried out to determine a therapeutic index (TI)
TI = TD50/ED50

The therapeutic index is sometimes usedin research to compare the safety of different members of a family of drugs

A drug that binds to a drug receptor without activating it acts as an antagonist A drug that binds to any portion of a receptor molecule and inactivates it through a chemical action will antagonize the action of agonists

Competitive pharmacologic antagonism

Reversible binding to the receptor site that can be

surmounted by an agonist will increase the measured EC50 of the agonist drug but have no effect on the Emax (see panel A below)

Irreversible pharmacologic antagonism

Irreversible binding that cannot be surmounted by

any concentration of agonist will decrease the Emax, but will not affect the EC5o for agonists (see panel B below) If spare receptors are present, the EC50 will be shifted to the right by low doses of the antagonist until all the spare receptors are blocked Further increases in the antagonist dose will then decrease the Emax

Physiologic antagonism
Binding of an agonist drug to a receptor that

produces effects opposite to the effects of another agonist drug acting at a second receptor Effects on Emax and EC50 are dose- and drugdependent

Physiologic antagonists may be very effective and rapid acting Ex. Epinephrine is the drug of choice for anaphylaxis
Epinephine acts as a physiologic antagonist to

leukotrienes and other mediators that do not act at epinephrine receptors

Chemical antagonism
A drug that chemically binds an agonist drug and

prevents it from acting on its receptors is a chemical antagonist

Partial agonist effect

Drugs within a chemical family bind to the same

receptor but not all produce the same maximum effect (Emax) Partial agonists - drugs that produce less than the full effect observed for that receptor system, even when given in doses that fully saturate receptors
Bind reversibly to the same receptors Act like competitive pharmacologic antagonists when combined with full agonists