plasma protein binding: friend or foe in drug discovery? · pharmacokinetics, dynamics and...

Kevin Beaumont and Tristan Maurer

Pharmacokinetics, Dynamics and Metabolism

Pfizer Worldwide Research and Development

Memorial Drive, Cambridge

Massachusetts USA

Plasma Protein Binding: Friend or Foe in Drug Discovery?

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Tota

l and

Fre

e Pl

asm

a Co

ncen

tratio

n (n

g/m

L)

Total Free

Drugs with 2 log units between total and free plasma concentration, percent bound greater than 98-99.9 %.

Plasma Protein Binding of Some Top 100 Most Prescribed Drugs

Smith, D.A. et. Al. Nat. Rev. Drug Disc. 2010, 9, 929-939.

If ppb drives dose, why are these compounds successful drugs?

Individual Drugs

•  Fluconazole

•  fu = 0.88

•  Identical free drug

concentration (SD or MD)

–  Plasma

– CSF, Saliva

– Vaginal secretions

– Breast milk, sputum

–  Prostatic seminal vesicle fluid

  Naproxen

  fu = 0.003

  Identical free drug

concentration (SD or MD)  Plasma

 Synovial fluid (deep tissue fluid)

Suggests high plasma protein binding is not a hindrance to distribution or success

Title

Pfizer Internal Use Only

Importance of selecting your battles

We have many foes in Drug Discovery Toxicity Potency Selectivity Intrinsic Clearance Drug Drug Interactions Reactive metabolites Etc

Why pick a fight with a neutral agent? Attacking ppb is like invading Switzerland! No benefit and a whole heap of trouble!

Title


Plasma Protein Binding is a Neutral Parameter

Plasma protein binding is a key parameter to measure

Plasma protein binding is not a key parameter to modulate

Important to unmask free concentration

Important in PK and dose prediction

Neutral on free concentration

Neutral on half-life*

Neutral on dose Exception is when Vd is fixed to blood volume*

Free drug concentrations and antimuscarinic activity of zamifenacin

Rat Man

Dose mg/kg Cmax (total) ng/ml Cmax (free) ng/ml

10

1

.1

100

1000

10000

Translatable Biomarker Inhibition of pupillary response to light (rat) or transient blurred vision (man)

Zamifenacin, potent and selective M3 muscarinic antagonist.

99.80% 99.98%

Beaumont et al, 1994, Brit J. Clin. Pharm. 38, 179P

10-fold difference in free fraction, 0.002 and 0.0002 respectively Highly bound in both species

Free concentration driving effect is identical

Comparison of M3 Muscarinic Receptor Antagonists

Dose depends on potency and intrinsic clearance, but not PPB

Compounds Zamifenacin / Darifenacin

In Vitro K b (nM) 2 fold

Intrinsic Clearance Similar

F u 300 fold (0.06 vs. 0.0002)

Daily Clinical Dose 2 fold

Zamifenacin Darifenacin

Dose – Steady State Equation

(1965) Wagner,F

CLCτ

Dose avg ss,Nature

•=

Potency Clearance

Absorption + CL

Objective

Important: CLint is the unbound intrinsic clearance (the theoretical maximum rate of clearance in the absence of flow and binding limitations)

(1965) Wagner,F

CLCτ

Dose avg ss,Nature

•=

Potency Clearance

Absorption + CL

Objective

For hepatically cleared drugs dosed orally

( )( )

⎟⎟⎠

⎞⎜⎜⎝

⎛−•=

•+

••=

QhCL1faF

CLintfuQhCLintfuQhCL Pang, J Pharmacokin Biopharm (1973)

Substitute and simplify

⎟⎟⎟⎟⎟

⎠

⎞

⎜⎜⎜⎜⎜

⎝

⎛⎥⎦

⎤⎢⎣

⎡

•+••

−•

•⎟⎟⎠

⎞⎜⎜⎝

⎛

•+••

•⎟⎠

⎞⎜⎝

⎛

=

QhCLint)(fuQhCLintfuQh

1fa

τCLint)(fuQhCLintfuQh

fuuCss,

Dose

( )

⎟⎟⎠

⎞⎜⎜⎝

⎛⎥⎦

⎤⎢⎣

⎡

•+•

−•

•⎟⎟⎠

⎞⎜⎜⎝

⎛

•+•

•

=

CLint)(fuQhCLintfu1fa

τCLint)(fuQh

CLintQhuCss,Dose

( )

CLint)(fuQhCLintfu-CLint)(fuQhfa

τCLint)(fuQh

CLintQhuCss,Dose

•+••+

•

•⎟⎟⎠

⎞⎜⎜⎝

⎛

•+•

•

=

⎟⎠

⎞⎜⎝

⎛ ••=

faτCLintuCss,Dose

Cancel fu and Qh terms

simplify

( )CLintfu-CLint)(fuQhfa

CLint)(fuQhτCLint)(fuQh

CLintQhuCss,Dose••+•

•+••⎟⎟

⎠

⎞⎜⎜⎝

⎛

•+

••=

Consolidate numerator and denominator

Substitute terms to reveal protein binding influence

Simplify denominator

“the chemists most important pharmacokinetic equation”

Determinants of Dose

No requirement for fu

“Plasma protein binding alone cannot be regarded as either a positive or negative aspect of a compound” - van de Waterbeemd, J Med Chem (2001)

“The only determinants of unbound steady state exposure are intrinsic clearance and dose” - Harold Boxenbaum

(1965) Wagner,F

CLCτ

Dose avg ss,Nature

•=

Potency Clearance

Absorption + CL

Objective

For hepatically cleared drugs dosed orally

( )( )

⎟⎟⎠

⎞⎜⎜⎝

⎛−•=

•+

••=

QhCL1faF

CLintfuQhCLintfuQhCL Pang, J Pharmacokin Biopharm (1973)

Substitute and simplify

⎟⎟⎟⎟⎟

⎠

⎞

⎜⎜⎜⎜⎜

⎝

⎛⎥⎦

⎤⎢⎣

⎡

•+••

−•

•⎟⎟⎠

⎞⎜⎜⎝

⎛

•+••

•⎟⎠

⎞⎜⎝

⎛

=

QhCLint)(fuQhCLintfuQh

1fa

τCLint)(fuQhCLintfuQh

fuuCss,

Dose

( )

⎟⎟⎠

⎞⎜⎜⎝

⎛⎥⎦

⎤⎢⎣

⎡

•+•

−•

•⎟⎟⎠

⎞⎜⎜⎝

⎛

•+•

•

=

CLint)(fuQhCLintfu1fa

τCLint)(fuQh

CLintQhuCss,Dose

( )

CLint)(fuQhCLintfu-CLint)(fuQhfa

τCLint)(fuQh

CLintQhuCss,Dose

•+••+

•

•⎟⎟⎠

⎞⎜⎜⎝

⎛

•+•

•

=

⎟⎠

⎞⎜⎝

⎛ ••=

faτCLintuCss,Dose

Cancel fu and Qh terms

simplify

( )CLintfu-CLint)(fuQhfa

CLint)(fuQhτCLint)(fuQh

CLintQhuCss,Dose••+•

•+••⎟⎟

⎠

⎞⎜⎜⎝

⎛

•+

••=

Consolidate numerator and denominator

Substitute terms to reveal protein binding influence

Simplify denominator

“the chemists most important pharmacokinetic equation”

Determinants of Dose

No requirement for fu

“Plasma protein binding alone cannot be regarded as either a positive or negative aspect of a compound” - van de Waterbeemd, J Med Chem (2001)

“The only determinants of unbound steady state exposure are intrinsic clearance and dose” - Harold Boxenbaum

Determinants of Dose – Oral drugs, hepatically cleared

Cliu = 1000 mL/min/kg, Q = 50 mL/min/kg, Complete absorption, Fixed dose Clearance

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0 0.2 0.4 0.6 0.8 1

ClH = Q(fu.Cliu)/(Q+fu.Cli

u)

fu

AUC

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10000

0 0.2 0.4 0.6 0.8 1

AUC total = dose/Cl

fu

Bioavailability

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100

0 0.2 0.4 0.6 0.8 1

F = 1 - (ClH/Q)

fu

AUCu

0 0.4 0.8 1.2 1.6

2

0 0.2 0.4 0.6 0.8 1

AUC u= dose/ Cl u

fu

Thought Experiment 1 –Modulate Fraction Unbound, Maintain Dose and Intrinsic Clearance (unbound)

Math courtesy of Don Walker

It is tough to modulate fraction unbound - IPRL

In vivo: Ctot decreases with increase in fu but Cfree does not change

Isolated perfused rat liver model: modulate ppb with amount of BSA in perfusate

Liu, X. et. Al. Curr. Top. Med Chem. 2011, 11, 450-466.

Total Total

Free Free

15

Nagase analbuminemia rats (NAR): mutant strain from wild type Spraque-Dawley rats (less albumin)

Decrease in fu does not change the Concu ONLY the Conctot

Liu, X. et. Al. Curr. Top. Med Chem. 2011, 11, 450-466.

Total Free

3X

It is tough to modulate fraction unbound – NAR Rats

16

Dose is the most important parameter to control

FτCLC

Dose avg ss, ••=

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1 10 100 1000 10000

Unbound clearance (ml/min/kg)

Aver

age

unbo

und

conc

entr

atio

n

(nM

)

Clu 10ml/min/kg

Cav 520nM

Clu 1000ml/min/kg

Cav 5nM

Average plasma concentration at a dose of 200mg

Lower dose More potent Lower Clun

τ is dose interval

Css is steady state Cav (unbound) Related to potency Cl is the clearance of the compound (unbound) Related to metabolism/excretion rate

Modulating Dose - Unbound Clearance and Potency (unbound concentration)

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Pote

ncy

(nM

)

CLint (ml/min/kg)

Unbound intrinsic clearance (Clint), in vitro potency and dose size

Dose = 20mg

Dose = 200mg

Dose = 2000mg

Propranolol (60mg)

Dofetilide (10mg)

Revatropate (1mg)

Sildenafil (100mg)

Atenolol (75mg)

Carbemazepine (1000mg)

Fluoxetine (20mg) Indomethacin (100mg)

Acetaminophen (4 g)

Artemisinin(250mg)

Quinine (2g)

Modulating Dose - Dose calculation for marketed drugs

18

Why is the literature replete with approaches that target lower plasma protein binding?

A Conundrum Wrapped up in a Riddle

The Paradox:

A strategy to lower plasma protein binding by lowering log D(7.4) may appear to improve dose………

……… but only by lowering unbound intrinsic clearance!!

van de Waterbeemd, J Comp Aid Mol Des (2001)

The tactics to modulate fraction unbound are the same as those to modulate unbound intrinsic clearance

QhCLh1faF

fufu

VtVcVd

fuGFRor fuCLintQhfuCLintQh

CL

tissue

plasma

plasmaplasma

plasma

⎟⎠⎞⎜

⎝⎛ −×=

⎟⎟⎠

⎞⎜⎜⎝

⎛+=

××+

××=

Target Exposure: 5nM as Cu,ss,max, Cu,ss,min or Cu,ss,avg Primary parameters: fu: 0.01 – 1 CLint: 1 – 100,000 ml/min/kg fa=1 Tau=24 hrs ka=1.8 hr-1

futissue=0.01 Vc = 0.2 L/kg (extracellular fluid) Vt = 0.4 L/kg (intracellular fluid) Secondary parameters: Vd = 0.6 – 40 L/kg CL:0.01 – 19.996 ml/min/kg t1/2: 1 – 800 hr

Physiological Parameter Models:

Compartmental Model:

Unless we want to make life complicated!

Hepatic / PO PO + Hepatic CL

1

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1000

10000

100000

0.01 0.1 1

fu

Dose

(mg)

CLint = 1

CLint = 100000

Math courtesy of Tristan Maurer

Dose to Achieve Target Cav

FτCLavgCss,Dose ••

=

Overall, dose is independent of fu

As fu Css,av but is offset by Cl and F

Hepatic / IV IV + Hepatic CL

1

10

100

1000

0.01 0.1 1

fu

Dos

e (m

g)

CLint = 1

CLint = 100000

1τCLavgCss,Dose ••

=

As fu Css,av but is only partially offset

by Cl Effect is only seen at high

values of intrinsic clearance

Bottom Line: Easier to modulate dose with Clint (and potency) rather than fu

Hepatic / PO PO + Hepatic CL

0.1

1

10

100

1000

10000

100000

0.01 0.1 1

fu

Dos

e (m

g)

CLint = 1

CLint = 100000


Dose to Achieve Target Cmax

IV + Hepatic CL

0.1

1

10

100

0.01 0.1 1

fu

Dose

(mg)

CLint = 1

CLint = 100000

Hepatic / IV

maxelel tkk- e)e-(1VdF

CmaxDose ×× ×××= τ

As fu Vd and Cl but Cmax and F Increased dose requirements

only observed at high Clint due to nonlinear effects on Cl and F

As fu Vd and Cl but Cmax Decreased dose requirements

with fu only observed at high Clint


Dose to Achieve Target Cmin

PO + Hepatic CL

0.1

10

1000

100000

1E+07

1E+09

1E+11

1E+13

1E+15

1E+17

1E+19

1E+21

0.01 0.1 1

fu

Dose

(mg)

CLint = 1

CLint = 100000

Hepatic / PO


( )( )τkelτka

a

τ-kael

eekF)e-(1kkVdCminDose

el

×−×−

×

−××

•−••=

Increasing fu, affects parameters the same way leading to a decrease in dose, but in a way that is nonlinearly related to CLint with a impact only at higher values of Clint and very high doses

IV + Hepatic CL

0.1

1

10

100

1000

10000

100000

1000000

0.01 0.1 1

fu

Dos

e (m

g)

CLint = 1

CLint = 100000

Hepatic / IV

With loss of F, the effect is less pronounced, but still evident at high Clint values


Title


Plasma Protein Binding is a Neutral Parameter

Plasma protein binding is a key parameter to measure

Plasma protein binding is not a key parameter to modulate

Important to unmask free concentration

Important in PK and dose prediction

Neutral on free concentration

Neutral on half-life*

Neutral on dose Exception is when Vd is fixed to blood volume*


Unless you want to make it complicated!

Summary and Recommendation

The next time your Discovery colleagues want to lower ppb…….

…….ask them to consider: Dose route (iv or po) In Vitro potency Target (Cav, Cmax or Cmin) Clearance route (hepatic metabolic or renal) Clintu

Keep calm and carry on!

Acknowledgements

Dennis Smith Li Di

Don Walker Tristan Maurer

Back Ups


Dose to Achieve Target Cav

Dose Css,avg CLτ F

•= Hepatic PO Hepatic IV Renal PO


Dose to Achieve Target Cmax

maxelel tkk- e)e-(1VdF

CmaxDose ×× ×××= τ

Hepatic PO Hepatic IV Renal PO


Dose to Achieve Target Cmin

Hepatic PO Hepatic IV Renal PO ( )( )τkelτka

a

τ-kael

eekF)e-(1kkVdCminDose

el

×−×−

×

−××

•−••=

plasma protein binding: friend or foe in drug discovery? · pharmacokinetics, dynamics and...

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