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Plasma drug protein binding

Update: 01:/07/2006
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Biological relevance of drug binding
The binding of drug to plasma (and tissue)
proteins is a major determinant of drug
disposition (distribution)
Binding has a very important effect on
drug dynamics since only the free
(unbound) drug interacts with receptors
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Relevance of plasma and tissue
protein binding
From a biological point of view YES
From a clinical point of view NO
problem of drug interaction and
displacement has been overestimated
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Influence of drug binding on
pharmacokinetic parameters
Distribution
Tissue
u
u
Plasma D
V
f
f
V V
T

+ =
Elimination
int u organ
int u
organ
organ
Cl f Q
Cl f Q
Cl
+

=
o
o
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Clinical relevance of drug binding
The importance of plasma protein
binding displacement interaction has
been overestimated and overstated
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For extrapolation
from in vitro to in vivo
in vitro, Kd (binding) and EC
50
(functional response)
are free concentrations but EC
50
(for PK/PD) is total
concentration
CMI (free) vs effective plasma concentration (Ctot)
between species
comparison of EC
50
between animals requires to take
into account free fraction
Case for which we need to know
in vivo free concentration
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The problem of drug interaction
and displacement has been
overestimated
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The classical example:
Phenylbutazone/warfarin interaction
Interaction actually exists
Displacement actually exists
but the plasma binding displacement
is not the underlying mechanism of
interaction
PBZ stereoselectivity inhibits the
metabolism of s-warfarin
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Why plasma binding seldom
has clinical relevance
Because few drugs (so-called displacer) are
therapeutically used
Because when displacement exists, it has
no consequence on the receptor exposure
to the free concentration of the displaced
drug which generally remains unaffected
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Is there often displacement of
drug from the binding site?
No
For a substantial displacement to take
place, the displacer must occupy most of
the available binding site thereby lowering
the binding site available to the primary
drug
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No
To take place, the molar concentration of
the drug in plasma must exceed the molar
concentration of albumin (150 g/mL for a
a drug with a MW of 250)
e.g.: PBZ, phenytoin, valproic acid
This is not true for o1-glycoprotein acid
(basic drug)
Is there often displacement of
drug from the binding site?
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Why plasma protein displacement
seldom has clinical relevance
Generally only the free (unbound) drug is
metabolized and can access to the
receptor
AND
the free drug concentration is controlled
by the free drug clearance which is
independent of the plasma binding
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Plasma drug protein binding
Physiological aspects
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Plasma binding proteins
Proteins MW Concentration
g/L M
Albumin 67 000 35-50 500-700
o
1
-glycoprotein 42 000 0.4-1.0 9-23
acid
Lipoproteins 200 000 variable
to 2.4 10
6

Transcortin 53 000 0.03-0.07 0.6-1.4
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Drug binding protein concentration and
percentage of free drug in serum of healthy
dogs and dogs with inflammation
Healthy Inflammation level of significance
Total protein (g/L) 71.6 72.3 NS
Albumin (g/L) 31.3 27.6 xxx
o-acid glycoprotein (mg/L) 374 1632 xxx

Percentage free
Lidocaine 43.5 11.7 xx
Propanolol 27.8 9.3 xx
Phenytoin 18.1 17.6 NS
Digitoxin 15.5 18.9 xx
Diazepam 1.57 2.78 x
Baggot The physiological basis of vet clin pharmacol p.103
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The free fraction
&
the free concentration
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Drug plasma protein binding
Expressed in % or by fu (free fraction)
>90% = highly bound
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The free fraction : fu
fu = =
free concentration
total concentration
C
free

C
tot

Definition:
fu and Cfree are not synonymous terms
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The free, the Bound
&
the total concentration
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C K
C
B

C
D
free
free
max
bound
+

=
The bound concentration
C
bound

C
free

B
max

K
D

The bound concentration
B
max
: maximal concentration of
binding sites
proportionnal to plasma protein concentration

K
D
: free drug concentration corresponding to half maximal binding
inversely proportional to drug affinity for the protein
B
max
/2
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C
tot
is a function of C
free

C
tot
= C
free
+ C
bind


C
tot
= C
free
+
Bmax x C
free

Kd + C
free

Dependent variable
Parameters
Independent variable
controlled by Cl
free

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Relationship between fu, the Free
and the bound concentrations
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The free fraction fu
Physiological factors controlling fu

fu = =


C
free

C
tot

C
free

C
free
+ C
bind

free
free
free
free
C Kd
C B
C
C
fu
+

+
=
max
cfre ctot
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C K B
C K

f
D max
free
D
free
u
+ +
+
=
Linear binding : C
free
<< K
D

D max
D
u
K B
K

f
+
=
The unbound fraction : fu
C C
C
f
bound free
free
u
+
=
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Total concentration:
a convenient but illicit rearrangement
which can be misleading when
discussing drug interaction
C
free
= fu x C
total

indirectly
estimated
known from in
vitro assay
measured by
analytical technique
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Total concentration

C
tot
=


! When conceptualizing dependency and
functionality, this equation should not be
rearranged
C
free

fu
!
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Total concentration:
Why the free displaced drug concentration
is not controlled by plasma binding
The fundamental relationship

Total concentration =
free concentration
fu
(free fraction)
parameters
dependent variable
Where fu is altered, C
tot
is modified, not C
free

independent variable
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What is the consequence of fu |

C
tot
+ = or C
free
| = fu | x C
total
Total concentration:
a convenient but illicit rearrangement
which can be misleading when
discussing drug interaction
C
free

fu |
NO
YES
Displacement (fu) modifies C
tot
, not C
free

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Drug interaction and
protein binding
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Ctot
Cfree
fu =
AND
fu
Cfree
Ctot =
fu
Cfree
Ctot =
Definition
Physiological relationship
Bmax
Kd
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Drug interaction and protein binding
C
tot
= C
free
+
Bmax x C
free

Kd + C
free

possible interaction
Interaction will modify C
tot
but not C
free

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Conditions in which the plasma
concentration of the 2 major plasma
proteins to which drug binds are altered
Conditions Change in concentration
Albumin hepatic cirrhosis
burns
nephritic syndrome
pregnancy

o-glycoprotein myocardial infarcts
surgery
trauma
rheumatoid arthritis
Rowland p.152
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Competitive interaction
C
tot
= C
free
+



C
tot
is decreased
Bmax x C
free

Kd (1 + A/K
i
) + C
free

Displacement
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Competitive interaction
Case of restrictively eliminated drug
Cl
free
= Clint = constant Cl
tot
= fu x Clint
perfusion rate: K
0

redistribution
C
tot
C
free
Cl
free
= cst
Cl
tot
Administration of the 2nd ligand, displacement fu |
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Competitive interaction for
restrictively eliminated drugs
when interaction occurs,
C
tot
is altered not C
free

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in vitro vs. in vivo situation
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In vitro - closed system In vivo - open system
Effect
C
tot

C
free

Competitive interaction
1.0
0.5
0.2
fu = 0.2
fu = 0.4
if fu | then C
tot
+
Time
C
ss, free

fu
1
C
ss, tot

=
tot ss, u
C f
=
constant
Cl
rate perfusion
C
ss, free

int
= =
constant
=
Drug with low extraction ratio
C
tot

C
free

Competitive interaction
1.0
0.5
0.2
fu = 0.2
fu = 0.4
if fu | then C
free
|
Time
C
ss, free

C
ss, tot

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fu vs Cfree: in vitro situation
fu = 0.5
Cfree = 3/V
Ctot = 6/V
fu = 0.83
Cfree = 5/V
Ctot = 6/V
1 2
6
5
4
3
1 2
6
5
4
3
displacer
displacee
V= volume of
the baker
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1
2
6
5
4
3
Infusion=A

A=MT
-1

K
12
Cfree
K
21
Cfree
Plasma Extracellular fluid Intracellular fluid
Elimination = K
10
x Cfree (3) = A equated by infusion
TOTAL CONCENTRATION = 6/V
FREE CONCENTRATION = 3/V
fu vs Cfree
in vivo situation: initial steady state
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1
2
6
5
4
3
Infusion=A

A=MT
-1

K
12
xCfree: increase
transitively
K
21
x Cfree
Plasma Extracellular fluid Intracellular fluid
K
10
x Cfree (5) > A
displacer
Increase transitorily
TOTAL CONCENTRATION = 6/V
FREE CONCENTRATION = 5/V
fu vs Cfree: in vivo situation:
just after administration of displacer
Just
displaced
free drug
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1
2
6
3
Infusion=A

A=MT
-1

K
12
x Cfree
K
21
x Cfree
Plasma Extracellular fluid Intracellular fluid
Elimination = K
10
x Cfree (3) = A
displacer
TOTAL CONCENTRATION = 4/V
FREE CONCENTRATION = 3/V
fu vs Cfree:
in vivo situation: final steady state
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The three main exceptions to the
general rule for which drug interaction
has no clinical meaning
1. Rapid bolus IV injection
2. Parenteral administration of displaced drug
with a high extraction ratio
3. Therapeutic drug monitoring and drug
displacement from the plasma binding site
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Case for which drug interaction
at the binding site is relevant
1. Rapid IV injection
If the displacing agent is given rapidly (IV
bolus), the C
free
could increase
dramatically due to rapid displacement of
the displaced drug before the
compensatory mechanism (redistribution)
takes place
Sulfamide and bilirubin kernicterus
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3. Therapeutic drug monitoring and drug
displacement from plasma binding
Therapeutic drug monitoring is performed for
drugs with a narrow concentration range
between therapeutic and toxic effect
Monitoring is carried out on total plasma
concentrations
Case for which drug interaction at
the plasma binding site is relevant
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Case for which drug interaction at
the plasma binding site is relevant
3. Therapeutic drug monitoring and drug
displacement from plasma binding
An example:
Phenytoin alone: C
tot
= 20 g/mL
Phenytoin + Valproic acid: C
tot
= 15 g/mL
no dosage adjustment is necessary because
C
tot
decreased but not C
free
due to fu increase

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Algorithm for determining clinical significance
of potential binding displacement interaction
Is drug of interest >90%
protein bound?
Yes
Does the drug have a
narrow therapeutic index ?
Yes
What is the hepatic extraction
ratio of the drug ?
High
Is the drug given IV?
Clinically significant interaction likely.
Perform a clinical study to quantify effects
Clinically significant
interaction not likely
Would a transient increase
in free drug concentration
be clinically relevant ?
no
no
low
no
no
Yes
Yes
Roslan 1994, B.J.Clin Pharmacol. 37, 125
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the overall clinical importance of plasma protein
binding displacement interactions continues to be
overstated
Despite the theoretical and experimental data to the
contrary, the concept that plasma protein binding
displacement is a common cause of clinically significant
interactions may still be widely taught in some medical
schools, often appears in textbooks and is accepted by
many in the medical community and by drug
regulators.
Protein Binding Interactions
Sansom LN & Evans AM. Drug Safety 1995;12:227-233.
Rolan PE. Br J Clin Pharmacol 1994;37:125-128.
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Protein Binding Interactions
Drugs for which pure plasma protein binding
displacement interactions will lead to sustained
changes in Css
u

Extensively bound to plasma proteins
Nonrestrictively cleared
Administered by non-oral route
alfentanil, buprenorphine, lidocaine, verapamil
Very few orally administered drugs exhibiting
properties of extensive plasma protein binding,
high hepatic first-pass extraction and narrow
therapeutic index