The role of pharmacokinetics in
drug discovery:
Where are we now? How did we
gett here?
h
? Where
Wh
are we going?
i ?
P t Webborn
Peter
W bb
RSC February 2013
PK studies in Drug Discovery
Why conduct PK studies in animals?
“The primary purpose of pre-clinical pharmacokinetic
studies is to validate the tools that will be used to
predict human kinetics”
2
The role of pharmacokinetics in drug discovery
Overview
1. The past - The 4 key steps that got us here
2. PK studies / data / tactics in a modern Drug
Discovery programme
3. The future – When free plasma concentrations
don’tt tell us everything
don
3
How did we get here?
Four key Developments
1.
2.
3
3.
4.
4
A Bioanalytical breakthrough
A Pharmacokinetic breakthrough
An Experimental breakthrough
A Conceptual change
Bioanalytical breakthrough
LC-MS : The Thermospray interface
Thermospray Source design
design,
Blackley et al (1978)
Historically
Gas chromatography (+derivatisation)
Thin layer chromatography
uv HPLC
LC MS (triple quadrupole)
LC-MS
Key for analysis in drug discovery - Sensitive, Selective, Generic, Fast
5
Pharmacokinetic breakthrough
The Introduction of “clearance concepts”
The ‘well stirred’ liver model
CL 
6
Q. fub .CLint
Q  fub .CLint
Pharmacokinetic breakthrough
Clearance – Six cornerstones of understanding
“The Clearance is the volume of blood cleared of drug per unit time”
1. Clearance ((not half-life)) is the best measure of the efficiencyy of an elimination p
process
2. Clearance is the scaling factor between the iv dose you give and the AUC you get!
3 Clearance relates the rate of elimination (ng/min) to the substrate concentration (ng/ml)
3.
V = CL x S
4. Determined from CL = Dose/AUCiv (units of flow)
5. Is influenced by plasma protein binding and by blood flow
6. “Intrinsic clearance” or CLint - relates free drug concentration to the rate of
elimination…….
elimination
7
Experimental breakthrough
Prediction of clearance from in vitro data
Also: Rane A,, Wilkinson GR and Shand DG,, Prediction of hepatic
p
extraction ratio from in vitro measurement of intrinsic
clearance. J Pharrnacol Exp Ther 200: 420- 424, 1977.
Also– No need to monitor appearance of metabolites, can be derived from
loss of parent compound - and - From V/S not Vmax / Km
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Conceptual breakthrough
PK properties are predictable and amenable to optimisation
Amounts and routes of
elimination. Types of
metabolism.
“The compound”
Dose and half-life
N
Name
and
d structure
t
t
Rates and affinities
The Chemical series
SAR
Concentration
and clearance
Physicochemical
properties
Predictions don’t always have to be right
“If you know what to expect – you are more likely to spot the unexpected”
9
Conceptual breakthrough - Example
SAR - Ionisation and Volume of Distribution (Vss)
60
40
Acid
20
Base
10
8
6
4
Neutral
2
1
0.8
0.6
0.4
0.2
01
0.1
0.08
-4
-3
-2
-1
0
logD
1
2
3
4
5
•
Vss tends to be acid < neutral <
base
•
Little influence of log D7.4
•
Why do we see this?
Understand drug behaviour – anticipating risks/issues
Examples: Acidic drugs
p
Poor Absorption
risk
Renal CL
Low permeability
Uptake
Transporter
substrate?
Interspecies
differences
Interspecies
differences
Interpretation of
PK data
Biliary CL
Acids
High Albumin
affinity
Glucuronidation
Enterohepatic
recirculation
Difficult to
measure very
high ppb
Interspecies
differences
Low Vss
Risk of short t½
Acyl
Glucuronide
reactivity risk
Interspecies
differences
Gut metabolism
Interspecies
differences
Standard
microsome
assays no use
Intrinsic
clearance must
be low
o
Interspecies
differences
PK studies in Drug Discovery
Where are we now?
Why conduct PK studies in animals?
- How to get most value from studies?
- How to use optimally to progress projects?
What have we learnt about experimental systems?
- Accuracy, reproducibility
- Optimal protocols
- Cassette dosing
Specific issues
- Plasma protein binding
- Formulation choice
- Hepatic
H
ti uptake
t k transporters
t
t
12
PK studies in Drug Discovery
Why conduct PK studies in animals?
“The primary purpose of pre-clinical pharmacokinetic
studies is to validate the tools that will be used to
predict human kinetics”
13
PK studies in Drug Discovery
Why conduct PK studies in animals?
Why do you want to know the......
Clearance
Half-life
Bioavailability
Volume of distribution*
..........of a compound in animals?
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PK studies in Drug Discovery
Assessing risk
What you really want to know how well you were able to predict in vivo kinetics
- Clearance - microsome /hepatocyte CLint - in silico
- Volume of distribution – Physicochemical properties – Vssu across species
- Bioavailability - Permeability /solubility – 1st pass metabolism
“I understand why this compound behaves the way it does”
Or
“I have no idea why this compound behaves the way it does”
“If I can predict the kinetics in rat and dog, I have a reasonable case to ask you
to believe I can predict human kinetics”
kinetics
Or
“If I can’t predict the kinetics in rat and dog, why should anyone think I can
predict human kinetics”
kinetics
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Predicting hepatic metabolic clearance
In vitro scaling factors,
fuP, Rb, fuinc
Scaled
CLint
Predicted
In vivo CLint
Predicted
in vivo clearance
• A regression approach adjusts for systematic underpredictions observed when scaling in vitro CLint directly
using the well stirred model, unbound fractions in blood
and the in vitro matrix, and physiological scaling factors.
Log(Q
Qh*CLb)/(Qh-CLb)
Raw
R
CLint
Well stirred
model (WSM)
Lab specific
correction
Riley, McGinnity and Austin (2005)
Log(CLint*SF*fu
g(
b/fuinc)
• This is commonly seen, and is not understood
• Correction factor is associated with the assay,
y, not the compound
p
PK studies in Drug Discovery
Setting criteria for an acceptable IVIVE
IVIVE.
1-sided 90% upper prediction limit
Log10 O
Observed CL
Lint in vivo (mll/min/kg)
2-sided
2
sided 80% prediction interval
Project 2
Project 3
Having an optimised, standardised
method puts the focus on the
compound, not the scaling method
Project 1
Rat reference set
Log10 Predicted CLintin vivo (ml/min/kg)
Allows a common understanding of
“scaling” and “non-scaling” compounds,
and uncertainty in predictions.
PK studies in Drug Discovery
Project example - IVIV correlations
RAT
• Compounds in general scaled well in rats if LogD was kept below 3
Clarity of message / ”rules” all can understand
Allows focus on compound – not scaling method
DOG
PK studies in Drug Discovery
Effective use of PK data - focus on prediction validation
Re-enforce positive project behaviours
• Predict - measure - learn
• Ensure correct use of in vitro data
• Build trust in in silico /in vitro systems
Supporting
S
ti
prediction
di ti continuum:
ti
IIn silico
ili
- in
i vitro
it - PK - PKPD
• Understanding relative risk/uncertainty in extrapolations
Efficient projects work in chemical series that are “predictable”
Candidate drugs that are understood, are less risky
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PK studies in Drug Discovery
A Problem...... Projects like having compounds tested
Less beneficial behaviours
• Overriding belief that more data = better informed
• Rigid screening cascades
• More “success” in testing cascade validates molecules
• Projects
P j t lik
like tto demonstrate
d
t t progress
• “It’s our best compound to date –let’s get a full data package”
Consequences
• Many measurements that fit with predictions
• Many results we learn nothing from
• Much data gathered on incrementally “better”
better compounds
• Too much data to manage –unfocussed optimisation strategies
Management of study requests
“I’ll run any test you are prepared to make a decision on”
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PK studies in Drug Discovery
A PK screening
i
strategy
t t
–“Enabling
“E bli the
th nextt decision”
d i i ”
HI
Number of
studies
Fold underprediction of
rat CL
21
LO
PK studies in Drug Discovery
The Power of Databases
What can we learn from having PK data on 1000’s of compounds?
• SAR
• Cassette dosing studies – Yielding a better understanding of Variability
(inter-animal /inter-study)
• Protocol
P
l enhancement
h
di
diagnosis
i
- N=3 v n=2
- First time point after iv bolus
- Cannulated v non-cannulated animals
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Renal Clearance Model
Key descriptors:
– Lipophilicity
– Ability of compound to carry a positive charge
PK studies in Drug Discovery
Use of Reference compounds to track assay performance
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Mean Clearance over 5 months
CL ml/min/kgg
25
20
Non-cannulated Animals
Cannulated Animals
15
10
5
0
06/05/2013
25/06/2013
14/08/2013
03/10/2013
22/11/2013
11/01/2014
Date of study
Reference compounds - Key advantage of cassette studies
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PK studies in Drug Discovery
How many animals to use? n=2 vs n=3
Impact on Vss estimates
Vss
within Count
0-20%
490
20-30%
65
30-40%
24
>40%
32
25
% of values
80
11
4
5
%C
Change iin AUC
PK studies in Drug Discovery
Impact on CL - 2min or 5 min first sample
300
30
~Clearance (ml/min/Kg)
3
N=2750
%C
Change iin AUC
Charnwood rat iv PK data
2min or 5 min first sample - Impact on CL
73% of TV within 10%
83% of JVC within 10%
90% of TV within 20%
93% of JVC within 20%
300
30
Clearance (ml/min/Kg)
3
N=2750
PK studies in Drug Discovery
Specific Issues
•Plasma protein binding
•Formulation
Formulation choice
•Transporters
28
Author | 00 Month Year
Set area descriptor | Sub level 1
Only one of these statements is mechanistically
correct..................
t
• “Because of the high plasma protein binding,
free plasma concentrations will be very low”
• “Because of the high plasma protein binding,
total plasma concentrations will be very high”
high
PK studies in Drug Discovery
Protein Binding - Don’t let it trip you up!
In vitro systems
In vivo systems
• A closed system
• An open system
• Free
F
levels
l
l d
driven
i
b
by
binding
• Free
F
levels
l
l d
driven
i
b
by
elimination rate
Smith et al (2010) Nature Drug Discovery Dec;9(12):929-39
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PK studies in Drug Discovery
Choice of formulation
““The formulation
f
should be appropriate for
f the
conclusion that will be drawn from the study”
e.g.
If you want to draw conclusions about likely human
p
a clinically
y relevant
bioavailabilityy /absorption,
formulation should be used.
If you are assessing exposure prior to an efficacy study
study,
ensure the formulation is tolerated for the duration of
study + does not affect the PD endpoint
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PK studies in Drug Discovery
Hepatic Uptake transporters
Uptake and efflux transporters have made understanding drug
clearance more complicated.....
..some
some simple concepts are no longer valid
“Additi it off clearance”
“Additivity
l
” - only
l applies
li to
t parallel
ll l processes
Transporter modelling requires “barriers”, serial processes and
concentration g
gradients across membranes
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“Uptake is the rate-limiting step in the overall hepatic
elimination of p
pravastatin at steady-state
y
in rats”
Yamazaki, M., Akiyama, S., Nishigaki, R., Sugiyama, Y. 1996 Pharmaceutical Research 13 (10), 1559
What is this really saying?
Consider this example: in a 3 step reaction:
For the formation of D (excretion into bile) - The rate determining step is
always
y the slowest step
p in the p
process.
For loss of A, the rate determining step is always k1 (plasma clearance)
In this scenario:
The rate of conversion of A to B depends on k1, k2 and k4.
For poorly permeable compounds uptake is the rate determining step
in the plasma clearance of active uptake substrates (because the
back-rate is insignificant)
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PK studies in Drug Discovery
The future
More Chemical diversity
-
Oligonucleotides
Extremely polar molecules - Intravenous antibiotics
Drug – Antibody conjugates
Nanotechnology delivery systems
I t
Instrumentation
t ti
- New interfaces – More sensitive
- Higher throughput - No chromatography
More reliance on predictions
- Cost
- Trust
Moving beyond plasma
- Mass Spectrometry Imaging (MSI)
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PK studies in Drug Discovery
When (free) plasma concentrations can’t tell the
whole story
• Free drug hypothesis
• It is the unbound drug that is in equilibrium with the target
• At equilibrium the free drug concentration in plasma and tissues are the
same
•Problem areas
•
•
•
•
•
•
Poorly perfused tissues
Hypoxic regions
Substrates for drug transporters
Active / toxic metabolites
Low target off-rates
Local administration (eg lung, skin)
•Mass spectrometry Imaging – Key points
•
•
•
•
•
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Drug / metabolite / biomarkers / metabonomics in tissues
Not “free
free drug”
drug
Not all compounds/ not all studies – a targeted approach
Resolution is not at the cellular level ~100µm (10µm)
Several rapidly evolving technologies (eg MALDI, DESI, LESA, SIMS)
PK studies in Drug Discovery
Mass Spectrometry Imaging
A unique insight into PK and PKPD – MALDI MSI
If y
you can g
get the
same answer
through tissue
homogenisation
– Don’t do MSI
Ch ll
Challenges
– Sensitivity
S
iti it and
d spatial
ti l resolution
l ti
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PK studies in Drug Discovery
Drug localisation as a driver of toxicity
Polymyxin nephrotoxicity – Aiding compound design
PMB1
PMB
AZ1
•The technique can meet speed/volume requirements of discovery
Programmes
•Need to translate results to man
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Summary
•PK is a well established component of drug discovery
•There is now a big opportunity to exploit PK databases
•PK resource management remains a challenge
• The future will be PKPD and translation
•MSI
MSI iis a rapidly
idl d
developing
l i ttechnique
h i
with
ith reall potential
t ti l tto
solve both efficacy and toxicity related problems
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