Drug-Receptor Interaction IX

Time Course of Plasma Concentration
and Effect
After the administration of a drug, its
concentration in plasma rises, reaches a
peak, and then declines gradually to the
starting level, due to the processes of
distribution and elimination.
Plasma concentration at a given point in
time depends on the dose administered.
Many drugs exhibit a linear relationship
between plasma concentration and
dose within the therapeutic range
(dose-linear kinetic ; note different
scales on ordinate). However, the
same does not apply to drugs whose
elimination processes are already sufficiently
activated at therapeutic plasma
levels so as to preclude further proportional
increases in the rate of elimination
when the concentration is increased
further. Under these conditions,
a smaller proportion of the dose administered
is eliminated per unit of time.
The time course of the effect and of
the concentration in plasma are not
identical, because the concentrationeffect
relationships obeys a hyperbolic
function. This means
that the time course of the effect exhibits
dose dependence also in the presence
of dose-linear kinetics .
In the lower dose range (example
1), the plasma level passes through a
concentration range (0 ! 0.9) in which
the concentration effect relationship is
quasi-linear. The respective time courses
of plasma concentration and effect
are very similar.
However, if a high dose (100) is applied,
there is an extended period of time during
which the plasma level will remain
in a concentration range (between 90
and 20) in which a change in concentration
does not cause a change in the size
of the effect. Thus, at high doses (100),
the time-effect curve exhibits a kind of
plateau. The effect declines only when
the plasma level has returned (below
20) into the range where a change in
plasma level causes a change in the intensity
of the effect.
The dose dependence of the time
course of the drug effect is exploited
when the duration of the effect is to be
prolonged by administration of a dose
in excess of that required for the effect.
This is done in the case of penicillin G
, when a dosing interval of 8 h is
being recommended, although the drug
is eliminated with a half-life of 30 min.
This procedure is, of course, feasible only
if supramaximal dosing is not associated
with toxic effects.
Futhermore it follows that a nearly
constant effect can be achieved, although
the plasma level may fluctuate
greatly during the interval between
doses.
The hyperbolic relationship be
tween plasma concentration and effect
explains why the time course of the effect,
unlike that of the plasma concentration,
cannot be described in terms of
a simple exponential function. A halflife
can be given for the processes of
drug absorption and elimination, hence
for the change in plasma levels, but generally
not for the onset or decline of
the effect.

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Drug-Receptor Interaction VIII

Mode of Operation of G-Protein-
Coupled Receptors
Signal transduction at G-protein-coupled
receptors uses essentially the same
basic mechanisms . Agonist binding
to the receptor leads to a change in receptor
protein conformation. This
change propagates to the G-protein: the
!-subunit exchanges GDP for GTP, then
dissociates from the two other subunits,
associates with an effector protein, and
alters its functional state. The !-subunit
slowly hydrolyzes bound GTP to GDP.
G!-GDP has no affinity for the effector
protein and reassociates with the " and
subunits . G-proteins can undergo
lateral diffusion in the membrane; they
are not assigned to individual receptor
proteins. However, a relation exists
between receptor types and G-protein
types . Furthermore, the !-subunits
of individual G-proteins are distinct in
terms of their affinity for different effector
proteins, as well as the kind of influence
exerted on the effector protein. G-
GTP of the GS-protein stimulates adenylate
cyclase, whereas G-GTP of the Giprotein
is inhibitory. The G-proteincoupled
receptor family includes muscarinic
cholinoceptors, adrenoceptors
for norepinephrine and epinephrine, receptors
for dopamine, histamine, serotonin,
glutamate, GABA, morphine, prostaglandins,
leukotrienes, and many other
mediators and hormones.
Major effector proteins for G-protein-
coupled receptors include adenylate
cyclase (ATP ! intracellular messenger
cAMP), phospholipase C (phosphatidylinositol
intracellular messengers
inositol trisphosphate and diacylglycerol),
as well as ion channel
proteins. Numerous cell functions are
regulated by cellular cAMP concentration,
because cAMP enhances activity of
protein kinase A, which catalyzes the
transfer of phosphate groups onto functional
proteins. Elevation of cAMP levels
inter alia leads to relaxation of smooth
muscle tonus and enhanced contractility
of cardiac muscle, as well as increased
glycogenolysis and lipolysis .
Phosphorylation of cardiac calcium-
channel proteins increases the
probability of channel opening during
membrane depolarization. It should be
noted that cAMP is inactivated by phosphodiesterase.
Inhibitors of this enzyme
elevate intracellular cAMP concentration
and elicit effects resembling those
of epinephrine.
The receptor protein itself may
undergo phosphorylation, with a resultant
loss of its ability to activate the associated
G-protein. This is one of the
mechanisms that contributes to a decrease
in sensitivity of a cell during prolonged
receptor stimulation by an agonist
(desensitization).
Activation of phospholipase C leads
to cleavage of the membrane phospholipid
phosphatidylinositol-4,5 bisphosphate
into inositol trisphosphate (IP3)
and diacylglycerol (DAG). IP3 promotes
release of Ca2+ from storage organelles,
whereby contraction of smooth muscle
cells, breakdown of glycogen, or exocytosis
may be initiated. Diacylglycerol
stimulates protein kinase C, which
phosphorylates certain serine- or threonine-
containing enzymes.
The subunit of some G-proteins
may induce opening of a channel protein.
In this manner, K+ channels can be
activated (e.g., ACh effect on sinus node,
; opioid action on neural impulse
transmission).

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Drug-Receptor Interaction VII

The insulin receptor protein represents
a ligand-operated enzyme , a
catalytic receptor. When insulin binds
to the extracellular attachment site, a
tyrosine kinase activity is “switched on”
at the intracellular portion. Protein
phosphorylation leads to altered cell
function via the assembly of other signal
proteins. Receptors for growth hormones
also belong to the catalytic receptor
class.
Protein synthesis-regulating receptors
for steroids, thyroid hormone,
and retinoic acid are found in the
cytosol and in the cell nucleus, respectively.
Binding of hormone exposes a normally
hidden domain of the receptor
protein, thereby permitting the latter to
bind to a particular nucleotide sequence
on a gene and to regulate its transcription.
Transcription is usually initiated or
enhanced, rarely blocked.

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Drug-Receptor Interaction VI

Receptor Types
Receptors are macromolecules that bind
mediator substances and transduce this
binding into an effect, i.e., a change in
cell function. Receptors differ in terms
of their structure and the manner in
which they translate occupancy by a ligand
into a cellular response (signal
transduction).
G-protein-coupled receptors
consist of an amino acid chain that
weaves in and out of the membrane in
serpentine fashion. The extramembranal
loop regions of the molecule may
possess sugar residues at different Nglycosylation
sites. The seven !-helical
membrane-spanning domains probably
form a circle around a central pocket
that carries the attachment sites for the
mediator substance. Binding of the mediator
molecule or of a structurally related
agonist molecule induces a change
in the conformation of the receptor protein,
enabling the latter to interact with
a G-protein (= guanyl nucleotide-binding
protein). G-proteins lie at the inner
leaf of the plasmalemma and consist of
three subunits .
There are various G-proteins that differ
mainly with regard to their !-unit. Association
with the receptor activates the
G-protein, leading in turn to activation
of another protein (enzyme, ion channel).
A large number of mediator substances
act via G-protein-coupled receptors.
An example of a ligand-gated ion
channel is the nicotinic cholinoceptor
of the motor endplate. The receptor
complex consists of five subunits, each
of which contains four transmembrane
domains. Simultaneous binding of two
acetylcholine (ACh) molecules to the
two !-subunits results in opening of the
ion channel, with entry of Na+ (and exit
of some K+), membrane depolarization,
and triggering of an action potential .
The ganglionic N-cholinoceptors
apparently consist only of ! and " subunits
. Some of the receptors for
the transmitter #-aminobutyric acid
(GABA) belong to this receptor family:
the GABAA subtype is linked to a chloride
channel (and also to a benzodiazepine-
binding site). Glutamate
and glycine both act via ligandgated
ion channels.

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Drug-Receptor Interaction V

Enantioselectivity of affinity. If a
receptor has sites for three of the substituents
(symbolized in B by a cone, a
sphere, and a cube) on the asymmetric
carbon to attach to, only one of the
enantiomers will have optimal fit. Its affinity
will then be higher. Thus, dexetimide
displays an affinity at the muscarinic
ACh receptors almost 10000 times
that of levetimide; and at !-
adrenoceptors, S(-)-propranolol has an
affinity 100 times that of the R(+)-form.
Enantioselectivity of intrinsic activity.
The mode of attachment at the
receptor also determines whether an effect
is elicited and whether or not a substance
has intrinsic activity, i.e., acts as
an agonist or antagonist. For instance,
(-) dobutamine is an agonist at "-adrenoceptors
whereas the (+)-enantiomer is
an antagonist.
Inverse enantioselectivity at another
receptor. An enantiomer may
possess an unfavorable configuration at
one receptor that may, however, be optimal
for interaction with another receptor.
In the case of dobutamine, the
(+)-enantiomer has affinity at !-adrenoceptors
10 times higher than that of the
(-)-enantiomer, both having agonist activity.
However, the "-adrenoceptor
stimulant action is due to the (-)-form
(see above).
As described for receptor interactions,
enantioselectivity may also be
manifested in drug interactions with
enzymes and transport proteins. Enantiomers
may display different affinities
and reaction velocities.
Conclusion: The enantiomers of a
racemate can differ sufficiently in their
pharmacodynamic and pharmacokinetic
properties to constitute two distinct
drugs.

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Drug-Receptor Interaction IV

Enantioselectivity of Drug Action
Many drugs are racemates, including !-
blockers, nonsteroidal anti-inflammatory
agents, and anticholinergics (e.g.,
benzetimide A). A racemate consists of
a molecule and its corresponding mirror
image which, like the left and right
hand, cannot be superimposed. Such
chiral (“handed”) pairs of molecules are
referred to as enantiomers. Usually,
chirality is due to a carbon atom
linked to four different substituents
(“asymmetric center”). Enantiomerism is
a special case of stereoisomerism. Nonchiral
stereoisomers are called diastereomers
(e.g., quinidine/quinine).
Bond lengths in enantiomers, but
not in diastereomers, are the same.
Therefore, enantiomers possess similar
physicochemical properties (e.g., solubility,
melting point) and both forms are
usually obtained in equal amounts by
chemical synthesis. As a result of enzymatic
activity, however, only one of the
enantiomers is usually found in nature.
In solution, enantiomers rotate the
wave plane of linearly polarized light
in opposite directions; hence they are
refered to as “dextro”- or “levo-rotatory”,
designated by the prefixes d or (+) and l
or (-), respectively. The direction of rotation
gives no clue concerning the spatial
structure of enantiomers. The absolute
configuration, as determined by
certain rules, is described by the prefixes
S and R. In some compounds, designation
as the D- and L-form is possible
by reference to the structure of D- and
L-glyceraldehyde.
For drugs to exert biological actions,
contact with reaction partners in
the body is required. When the reaction
favors one of the enantiomers, enantioselectivity
is observed.

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Drug-Receptor Interaction III

Agonist stabilizes spontaneously
occurring active conformation. The
receptor can spontaneously “flip” into
the active conformation. However, the
statistical probability of this event is
usually so small that the cells do not reveal
signs of spontaneous receptor activation.
Selective binding of the agonist
requires the receptor to be in the active
conformation, thus promoting its existence.
The “antagonist” displays affinity
only for the inactive state and stabilizes
the latter. When the system shows minimal
spontaneous activity, application
of an antagonist will not produce a measurable
effect. When the system has
high spontaneous activity, the antagonist
may cause an effect that is the opposite
of that of the agonist: inverse agonist.
A “true” antagonist lacking intrinsic
activity (“neutral antagonist”) displays
equal affinity for both the active and inactive
states of the receptor and does
not alter basal activity of the cell.
According to this model, a partial agonist
shows lower selectivity for the active
state and, to some extent, also binds
to the receptor in its inactive state.
Other Forms of Antagonism
Allosteric antagonism. The antagonist
is bound outside the receptor agonist
binding site proper and induces a decrease
in affinity of the agonist. It is also
possible that the allosteric deformation
of the receptor increases affinity for an
agonist, resulting in an allosteric synergism.
Functional antagonism. Two agonists
affect the same parameter (e.g.,
bronchial diameter) via different receptors
in the opposite direction (epinephrine
dilation; histamine ! constriction).

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Drug-Receptor Interaction II

Agonists – Antagonists
An agonist has affinity (binding avidity)
for its receptor and alters the receptor
protein in such a manner as to generate
a stimulus that elicits a change in cell
function: “intrinsic activity“. The biological
effect of the agonist, i.e., the
change in cell function, depends on the
efficiency of signal transduction steps
initiated by the activated receptor.
Some agonists attain a maximal
effect even when they occupy only a
small fraction of receptors
. Other ligands , possessing
equal affinity for the receptor but lower
activating capacity (lower intrinsic activity),
are unable to produce a full maximal
response even when all receptors
are occupied: lower efficacy. Ligand B is
a partial agonist. The potency of an agonist
can be expressed in terms of the
concentration (EC50) at which the effect
reaches one-half of its respective maximum.
Antagonists attenuate the effect
of agonists, that is, their action is
“anti-agonistic”.
Competitive antagonists possess
affinity for receptors, but binding to the
receptor does not lead to a change in
cell function (zero intrinsic activity).
When an agonist and a competitive
antagonist are present simultaneously,
affinity and concentration of the two rivals
will determine the relative amount
of each that is bound. Thus, although the
antagonist is present, increasing the
concentration of the agonist can restore
the full effect . However, in the presence
of the antagonist, the concentration-
response curve of the agonist is
shifted to higher concentrations (“rightward
shift”).
Molecular Models of Agonist/Antagonist
Action
Agonist induces active conformation.
The agonist binds to the inactive receptor
and thereby causes a change from
the resting conformation to the active
state. The antagonist binds to the inactive
receptor without causing a conformational
change.

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Drug-Receptor Interaction I

Types of Binding Forces
Unless a drug comes into contact with
intrinsic structures of the body, it cannot
affect body function.
Covalent bond. Two atoms enter a
covalent bond if each donates an electron
to a shared electron pair (cloud).
This state is depicted in structural formulas
by a dash. The covalent bond is
“firm”, that is, not reversible or only
poorly so. Few drugs are covalently
bound to biological structures. The
bond, and possibly the effect, persist for
a long time after intake of a drug has
been discontinued, making therapy difficult
to control. Examples include alkylating
cytostatics or organophosphates
. Conjugation reactions
occurring in biotransformation also
represent a covalent linkage (e.g., to
glucuronic acid).
Noncovalent bond. There is no formation
of a shared electron pair. The
bond is reversible and typical of most
drug-receptor interactions. Since a drug
usually attaches to its site of action by
multiple contacts, several of the types of
bonds described below may participate.
Electrostatic attraction. A positive
and negative charge attract each
other.
Ionic interaction: An ion is a particle
charged either positively (cation) or
negatively (anion), i.e., the atom lacks or
has surplus electrons, respectively. Attraction
between ions of opposite
charge is inversely proportional to the
square of the distance between them; it
is the initial force drawing a charged
drug to its binding site. Ionic bonds have
a relatively high stability.
Dipole-ion interaction: When bond
electrons are asymmetrically distributed
over both atomic nuclei, one atom
will bear a negative, and its partner
a positive partial charge. The molecule
thus presents a positive and a negative
pole, i.e., has polarity or a dipole. A
partial charge can interact electrostatically
with an ion of opposite charge.
Dipole-dipole interaction is the electrostatic
attraction between opposite
partial charges. When a hydrogen atom
bearing a partial positive charge bridges
two atoms bearing a partial negative
charge, a hydrogen bond is created.
A van der Waals’ bond is
formed between apolar molecular
groups that have come into close proximity.
Spontaneous transient distortion
of electron clouds (momentary faint dipole)
may induce an opposite dipole
in the neighboring molecule. The van
der Waals’ bond, therefore, is a form of
electrostatic attraction, albeit of very
low strength (inversely proportional to
the seventh power of the distance).
Hydrophobic interaction. The
attraction between the dipoles of water
is strong enough to hinder intercalation
of any apolar (uncharged) molecules. By
tending towards each other, H2O molecules
squeeze apolar particles from
their midst. Accordingly, in the organism,
apolar particles have an increased
probability of staying in nonaqueous,
apolar surroundings, such as fatty acid
chains of cell membranes or apolar regions
of a receptor.

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