Plasma drug protein binding
Update: 01:/07/2006
binding- 1
binding- 2
binding- 3
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
binding- 4
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
binding- 5
Influence of drug binding on
pharmacokinetic parameters
• Distribution
• Elimination
 fu 
VD  VPlasma     VTissue
 f uT 
o
Cl organ 
Q organ  f u  Cl int
o
Q organ  f u  Cl int
binding- 6
Clinical relevance of drug binding
• The importance of plasma protein
binding displacement interaction has
been overestimated and overstated
binding- 7
Case for which we need to know
in vivo free concentration
For extrapolation
• from in vitro to in vivo
– in vitro, Kd (binding) and EC50 (functional response)
are free concentrations but EC50 (for PK/PD) is total
concentration
– CMI (free) vs effective plasma concentration (Ctot)
• between species
– comparison of EC50 between animals requires to take
into account free fraction
binding- 8
The problem of drug interaction
and displacement has been
overestimated
binding- 9
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
binding- 10
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
binding- 11
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
binding- 12
Is there often displacement of
drug from the binding site?
• No
• To take place, the molar concentration of
the drug in plasma must exceed the molar
concentration of albumin (150 µg/mL for a
a drug with a MW of 250)
– e.g.: PBZ, phenytoin, valproic acid
• This is not true for a1-glycoprotein acid
(basic drug)
binding- 13
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
binding- 14
Plasma drug protein binding
Physiological aspects
binding- 15
Plasma binding proteins
Proteins
MW
Albumin
67 000
42 000
a-glycoprotein
acid
Lipoproteins
Transcortin
200 000
to 2.4 106
53 000
Concentration
g/L
µM
35-50
500-700
0.4-1.0
9-23
variable
0.03-0.07
0.6-1.4
binding- 16
Drug binding protein concentration and
percentage of free drug in serum of healthy
dogs and dogs with inflammation
Healthy
Total protein (g/L)
71.6
Albumin (g/L)
31.3
a-acid glycoprotein (mg/L) 374
Inflammation
72.3
27.6
1632
level of significance
NS
xxx
xxx
11.7
9.3
17.6
18.9
2.78
xx
xx
NS
xx
x
Percentage free
Lidocaine
Propanolol
Phenytoin
Digitoxin
Diazepam
43.5
27.8
18.1
15.5
1.57
Baggot The physiological basis of vet clin pharmacol p.103
binding- 17
The free fraction
&
the free concentration
binding- 28
Drug plasma protein binding
• Expressed in % or by fu (free fraction)
• >90% = highly bound
binding- 29
The free fraction : fu
Definition:
free concentration
• fu = total concentration =
Cfree
Ctot
fu and Cfree are not synonymous terms
binding- 30
The free, the Bound
&
the total concentration
binding- 32
The bound concentration
• The bound concentration
Bmax  Cfree
Cbound 
K D  Cfree
Cbound
Bmax
Bmax/2
Bmax : maximal concentration of
KD
binding sites
– proportionnal to plasma protein concentration
Cfree
KD : free drug concentration corresponding to half maximal binding
– inversely proportional to drug affinity for the protein
binding- 33
Ctot is a function of Cfree
Ctot = Cfree + Cbind
Ctot = Cfree +
Dependent variable
Bmax x Cfree
Kd + Cfree
Parameters
Independent variable
controlled by Clfree
binding- 34
Relationship between fu, the Free
and the bound concentrations
binding- 35
The free fraction fu
• Physiological factors controlling fu
• fu =
Cfree
Ctot
=
Cfree
ctot
cfre
Cfree + Cbind
fu 
Cfree
B max Cfree
Cfree 
Kd  Cfree
binding- 36
The unbound fraction : fu
Cfree
fu 
Cfree  Cbound
K D  Cfree
fu 
Bmax  K D  Cfree
• Linear binding : Cfree << KD
KD
fu 
Bmax  K D
binding- 38
Total concentration:
a convenient but illicit rearrangement
which can be misleading when
discussing drug interaction
Cfree = fu x Ctotal
indirectly
estimated
known from in
vitro assay
measured by
analytical technique
binding- 39
Total concentration
Ctot =
Cfree
fu
!! When conceptualizing dependency and
functionality, this equation should not be
rearranged
binding- 40
Total concentration:
Why the free displaced drug concentration
is not controlled by plasma binding
The fundamental relationship
Total concentration =
dependent variable
independent variable
free concentration
fu
(free fraction)
parameters
Where fu is altered, Ctot is modified, not Cfree
binding- 41
Total concentration:
a convenient but illicit rearrangement
which can be misleading when
discussing drug interaction
• What is the consequence of fu 
• Ctot  =
YES
Cfree
fu 
or Cfree  = fu  x Ctotal
NO
Displacement (fu) modifies Ctot, not Cfree
binding- 42
Drug interaction and
protein binding
binding- 46
Cfree
fu 
Ctot
AND
Cfree
Ctot 
fu
Ctot 
Cfree
fu
Bmax
Kd
Definition
Physiological relationship
binding- 48
Drug interaction and protein binding
• Ctot =
Cfree +
Bmax x Cfree
Kd + Cfree
possible interaction
Interaction will modify Ctot but not Cfree
binding- 49
Conditions in which the plasma
concentration of the 2 major plasma
proteins to which drug binds are altered
Albumin
a-glycoprotein
Rowland p.152
Conditions
Change in concentration
hepatic cirrhosis

burns

nephritic syndrome

pregnancy

myocardial infarcts
surgery
trauma
rheumatoid arthritis




binding- 52
Competitive interaction
Ctot =
Cfree +
Bmax x Cfree
Kd (1 + A/Ki) + Cfree
Displacement
Ctot is decreased
binding- 55
Competitive interaction
• Case of restrictively eliminated drug
Clfree = Clint = constant
perfusion rate: K0
Cltot = fu x Clint
Ctot
Cltot
Cfree
redistribution
Clfree = cst
Administration of the 2nd ligand, displacement  fu
binding- 60
Competitive interaction for
restrictively eliminated drugs
when interaction occurs,
Ctot is altered not Cfree
binding- 62
in vitro vs. in vivo situation
binding- 67
In vitro - closed system
In vivo - open system
Drug with low extraction ratio
Css, tot = constant
Css, free
Css, free = f u Css,tot
Css, tot = 1 C
ss, free
fu
perfusion
rate
= constant
=
Clint
if fu  then Ctot
if fu  then Cfree
Effect
1.0
0.5
Ctot
1.0
Cfree
0.5
fu = 0.4
fu = 0.2
Ctot
fu = 0.2
fu = 0.4
0.2
0.2
Cfree
Time
Time
Competitive interaction
Competitive interaction
binding- 68
fu vs Cfree: in vitro situation
4
4
2
1
5
5
3
3
6
fu = 0.5
Cfree = 3/V
Ctot = 6/V
2
1
6
displacer
displacee
V= volume of
the baker
fu = 0.83 
Cfree = 5/V 
Ctot = 6/V 
binding- 69
fu vs Cfree
in vivo situation: initial steady state
Extracellular fluid
Plasma
4
Infusion=A
A=MT-1
1
2
5
6
Intracellular fluid
K12 Cfree
3
K21 Cfree
Elimination = K10 x Cfree (3) = A equated by infusion
TOTAL CONCENTRATION = 6/V
FREE CONCENTRATION = 3/V
binding- 70
fu vs Cfree: in vivo situation:
just after administration of displacer
displacer
1
4
Infusion=A
A=MT-1
Extracellular fluid
Plasma
2
Intracellular fluid
K12xCfree: increase
transitively
5
6
3
K21 x Cfree
Just
displaced
free drug
Increase transitorily
K10 x Cfree (5) > A
TOTAL CONCENTRATION = 6/V 
FREE CONCENTRATION = 5/V 
binding- 71
fu vs Cfree:
in vivo situation: final steady state
displacer
Extracellular fluid
Plasma
K12 x Cfree
1
2
Infusion=A
A=MT-1
Intracellular fluid
6
3
K21 x Cfree
Elimination = K10 x Cfree (3) = A
TOTAL CONCENTRATION = 4/V 
FREE CONCENTRATION = 3/V 
binding- 72
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
binding- 73
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 Cfree could increase
dramatically due to rapid displacement of
the displaced drug before the
compensatory mechanism (redistribution)
takes place
 Sulfamide and bilirubin  kernicterus
binding- 74
Case for which drug interaction at
the plasma binding site is relevant
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
binding- 76
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:
Ctot = 20 µg/mL
– Phenytoin + Valproic acid: Ctot = 15 µg/mL
– no dosage adjustment is necessary because
Ctot decreased but not Cfree due to fu increase
binding- 77
Algorithm for determining clinical significance
of potential binding displacement interaction
Roslan 1994, B.J.Clin Pharmacol. 37, 125
Is drug of interest >90%
protein bound?
Yes
Does the drug have a
narrow therapeutic index ?
no
Clinically significant
interaction not likely
no
Yes
no
low Would a transient increase
in free drug concentration
be clinically relevant ?
What is the hepatic extraction
ratio of the drug ?
High
Is the drug given IV?
Yes
Yes
no
Clinically significant interaction likely.
Perform a clinical study to quantify effects
binding- 78
Protein Binding Interactions
“…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.”
Sansom LN & Evans AM. Drug Safety 1995;12:227-233.
Rolan PE. Br J Clin Pharmacol 1994;37:125-128.
binding- 79
Protein Binding Interactions
• Drugs for which pure plasma protein binding
displacement interactions will lead to sustained
changes in Cssu
– 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
binding- 80