Dose-Response

PHC7410 Principles of Toxicology

Thomas A. Kocarek, Ph.D.
Institute of Environmental Health Sciences
Office: IBio, Room 2126
Phone: 577-6580
e-mail: t.kocarek@wayne.edu
 Outline

I. General concepts
II. Types of dose-response relationships
III. Comparison of dose-response relationships
IV. Receptor theory
V. Interactions of chemicals
 Suggested Reading

• Casarett and Doull’s Toxicology, 7th Ed., Ch. 2,

Principles of Toxicology, pp. 19-28.
 I. General Concepts

Dose-Response
“This relationship is the most fundamental and
pervasive concept in toxicology.”
Casarett and Doull’s Toxicology
 Assumptions

1. Response is due to the chemical administered

2. Response is related to dose
a) there is a receptor site with which the chemical interacts
b) production and degree of response are related to the
concentration of the chemical at the receptor
c) concentration at receptor is related to the dose administered
(pharmacokinetics/toxicokinetics)
3. Can quantify a toxic response
Important to consider how doses are expressed
II. Types of dose-response relationships
1. Graded - continuous
• Relates intensity of an effect to dose
• Log dose usually gives more linear representation
of data
• potency - position along dose axis - potency in vivo is
influenced by pharmacokinetic properties (i.e., absorption,
distribution, metabolism and excretion) as well as by affinity
for receptor
• efficacy - maximal effect
• variability - differences in magnitude of response among
individuals in same population given the same dose of drug
• ED50 - dose that gives 50% of the maximal
response
Graded Dose-Response Curves
 Types of dose-response relationships

2. Quantal - all or none response

• Relates fraction/percentage of population
responding to dose
• e.g., LD50 - statistically-derived single dose of a substance
that can be expected to cause death in 50% of animals
• Quantal dose-response curves often exhibit a
normal frequency distribution
• Biological variation
• left of curve: hypersusceptible
• right of curve: resistant
Quantal Dose-Response Curves
 III. Comparison of dose-response
relationships
• Therapeutic index
• ratio of the dose required to produce a toxic effect and the dose
needed to elicit a desired therapeutic response
• generally use median doses (i.e., TI = LD50/ED50)
• tells nothing about shapes of curves
• Margin of safety
• for drug: LD1/ED99
• for non-drug: difference between an estimated exposed dose to
a human population and the highest non-toxic dose determined
in experimental animals
• Potency vs. efficacy
• potency - position of curve along dose axis
• efficacy - maximal effect
 IV. Receptor Theory
• Receptor
– specific recognition site with which a chemical
interacts to initiate a biological response
– usually macromolecular entity (protein)
– e.g., neurotransmitter receptors, ion channels,
enzymes
• Occupancy Theory
– first quantitative treatment of receptor theory
– attempt to understand and quantify interaction
between drug and receptor when little was known
about the mechanisms through which receptors
produce their biological effects
 Receptor Theory

A.J. Clark - 1926

• Reaction between drugs and receptors
described by law of mass action
• Assumptions:
1. Magnitude of pharmacological effect directly
proportional to fraction of receptors occupied.
Maximal response obtained only when all receptors
occupied.
2. One molecule of ligand interacts with one receptor.
3. Quantity of drug bound to receptor is negligible
relative to the total amount of drug in bathing
medium.
 EA [ RA]
Eq. 1 
EM [ RT ]

EA = observed response to agonist at

concentration A
EM = maximal obtainable response to agonist
[RA] = concentration of receptor-agonist
complex
[RT] = total concentration of receptor
 Eq. 2 R + A RA

Law of mass action

R = free receptor
A= free agonist
RA = receptor-agonist complex
 [ R][ A] [ R ][ A]
Eq. 3 Kd  [ RA] 
[ RA] Kd
Kd = dissociation constant
• reflects affinity for receptor
• equal to drug concentration that produces half-maximal
receptor occupancy

Eq. 4 [ RT ]  [ R]  [ RA]
Dividing equation 3 by equation 4

[ R ][ A]
Eq. 5 [ RA] Kd

[ RT ] [ R ]  [ RA]

Substituting Eq. 3 for [RA]

[ R][ A] Kd
[ RA] [ R]
Eq. 6  Kd X
[ RT ] [ R]  [ R][ A] Kd
[ R]
Kd
Simplifying

[ RA] [ A]
Eq. 7 
[ RT ] Kd  [ A]

Substituting Eq. 1
EA [ A]
Eq. 8 
EM Kd  [ A]
 EA [ A]

EM Kd  [ A]
• Clark proposed that this relationship adequately described the
response of an agonist simply as a function of the fractional
receptor occupancy.
• This theory predicts that any compound occupying a receptor
would produce a response proportional to the fraction of receptors
occupied. A maximal response would require 100% receptor
occupancy.
• No consideration is made for partial agonists (occupy receptor,
produce less than maximal response) or competitive antagonists
(occupy receptor, produce no response).
• Equation for rectangular hyperbola - Michaelis-Menten kinetics
• ED50 = Kd
 Receptor Theory

E.J. Ariëns - 1954

• Proposed that biological response should be divided
into two separate and independent parameters:
1. Affinity - binding of drug to receptor
2. Intrinsic activity - ability of drug to induce an effect after binding
• Agonist - affinity for receptor, intrinsic activity = 1
• Competitive antagonist - affinity for receptor, intrinsic
activity = 0
• Partial agonist - affinity for receptor, intrinsic activity >0,
<1
• Assumptions
1. Response to a full agonist is proportional to the
receptor occupancy with maximal response being
obtained only when all receptors occupied
2. Binding of drug to receptor governed by the law of
mass action; one drug molecule bound to one
receptor
3. Quantity of drug bound to receptor small relative to
total amount of drug available
 EA  [ RA]
Eq. 9 
EM [ RT ]

EA  [ A]
Eq. 10 
EM Kd  [ A]

ED50 = Kd
 Receptor Theory

R.P. Stephenson - 1956

• Expanded concepts of Clark and Ariëns
1. Response to a drug was some unknown positive function of
receptor occupancy - response not necessarily linearly
proportional to percentage of receptors occupied
2. Maximal response could be produced by an agonist when
occupying only a small proportion of receptors
3. Different drugs may have varying capacities to initiate a
response (i.e., efficacies) and therefore need to occupy
different proportions of receptors when producing equal
responses
• Efficacy replaces Ariëns’ intrinsic activity
 EA
 f (S )
EM

e[ RA] e[ A]
S 
[ RT ] Kd  [ A]

EA  e[ A] 
 f 
EM  Kd  [ A] 
• Spare receptors
– may be more receptors present than the
minimum number required to produce a
maximal response
– 100% receptor occupancy not always
required to produce a maximal response
– ED50 < Kd
 Receptor Theory

R.F. Furchgott - 1966

• Argued that efficacy was not strictly a drug parameter,
but depends on characteristics of tissue in question
(i.e., receptor numbers, receptor reserve, coupling
between occupancy and response, etc.)
• Proposed that efficacy was the product of intrinsic
efficacy (ε) and the concentration of active receptor in
a given tissue
• Intrinsic efficacy would be a property of a drug alone
that relates to the ability of a drug molecule to produce
a stimulus when acting on a given receptor
EA   [ RT ][ A] 
 f 
EM  Kd  [ A] 
 V. Interactions of Chemicals
• Additivity - combined effect of two chemicals is
equal to the sum of the effect of each agent
given alone (e.g., 2+3=5)
• Synergism - Combined effect of two chemicals
is much greater than the sum of the effect of
each agent given alone (e.g., 2+2=20)
• Potentiation - One chemical does not have a
toxic effect, but when added to another chemical
it makes the latter much more toxic (e.g.,
0+2=10)
 Antagonism
• Functional antagonism - two chemicals
counterbalance each other by producing
opposite effects on the same physiologic
function, but through different mechanisms
• Chemical antagonism or inactivation -
chemical reaction between two compounds to
produce a less toxic product
• Dispositional antagonism - one chemical
alters the disposition (i.e., absorption,
distribution, metabolism or excretion) of another
chemical
 Antagonism
• Receptor antagonism - one chemical inhibits
the effect of another chemical by binding to the
same receptor
– Competitive antagonist
• reversibly binds to receptor at active site
• mutually exclusive binding - surmountable
• decreases apparent affinity of agonist, not maximal effect
– Noncompetitive antagonist
• irreversibly binds to receptor at active site (or may bind at
another site)
• not surmountable
• decreases maximal effect of agonist, does not alter affinity
 Receptors not bound by
irreversible antagonist;
available to bind to agonist
{ Spare receptors
Receptors bound by irreversible
antagonist;
not available to bind to agonist

Number of receptors
required for 100%
response

A B C D E