Modern Tools for Drug Discovery NIMBUS Biotechnology Modern Tools for Drug Discovery

Modern Tools for Drug Discovery

NIMBUS Biotechnology


www.nimbus-biotechnology.com


Innovative tools in “ready to go plates”


NIMBUS Biotechnology


Bioavailability - an important issue

Absorption

Distribution

Metabolism

Excretion

Pharmacokinetic

Bioavailability


Absorption on a cellular level

getting into the blood system

Cross section of the gut wall

TRANSPORT:• passive paracellular• passive transcellular• active transcellular• efflux

blood stream


Distribution

bloodstream

Plasma/Tissuepartitioning

tissue e.g. brain

Balance of interactions: lipophilicity vs. serum binding


Absorption on a cellular level

getting into the blood system

Cross section of the gut wall

TRANSPORT:• passive paracellular• passive transcellular• active transcellular• efflux

blood stream


Kwater/lipid

Klipid/water

Klipid/water

Passive Transportin search of a good descriptor

lipid bilayers mimicking cellular systems

Passive TransportIn search of a good descriptor


Passive Transportin search of a good descriptor

“only absorption” additional information “hidden”

Klipid/water

Passive TransportIn search of a good descriptor


Lipids and Proteins on Solid Supports Innovative tools in “ready to go plates”

Prediction of absorption processes

Unspecific binding to HSAPrediction of distribution


• Determination of LIPOPHILICITY = Membrane Affinity (logMA)

• Non-covalently attached SINGLE LIPID BILAYER

• Different LIPID COMPOSITIONS Mimicking natural membranes

• High LONG TERM STABILITY 9 months

• DMSO ASSAY CONCENTRATION up to 5 %

• FAST SEPARATION due to the solid support

TRANSIL®


Validation log MA (charged and uncharged compounds) Comparison to liposome partitioning

Good correlation with liposome approach

1,0

1,5

2,0

2,5

3,0

3,5

4,0

1,0 1,5 2,0 2,5 3,0 3,5 4,0

log MA liposomes (ultracentrifugation)

log

MA

384

TR

AN

SIL

pla

te

384 well A

384 well B

384 well C

1 : 1 correlation


Validationfraction absorbed

logMA correlates with fraction absorbed

-8 -4 0 4 -2 0 2 4


High reproducibility from lot to lot

Validation IIILot to Lot and Reproducibility

0

0.5

1

1.5

2

2.5

3

3.5

4

4.5

5

0 10 20 30 40 50 60 70 80

n

log

MA

High reproducibility from measurement to measurement


Validation IV

0 1 2 3 4

0

1

2

3

4

logMA_NIMBUS 1:1 correlationlo

gMA

NIM

BU

S in

-hou

se d

ata

logMA Bayer Health Care

Good lab to lab reproducibility

Lab to Lab

* Introduced by Seiffert at NIMBUS Meeting 2005

0,0

0,5

1,0

1,5

2,0

2,5

3,0

3,5

4,0

0,0 1,0 2,0 3,0 4,0

logMA Schwarz Pharma*

logM

A N

IMB

US


TRANSIL-HSAProperties of the material

• random orientation• immobilized on a soft and inert surface• binding sites freely accessible• stable in presence of organic modifiers (e.g. 5% DMSO)


good correlation with classical equilibrium dialysis

TRANSIL-HSA Validation TRANSIL®-HSA vs. equilibrium dialysis

0 20 40 60 80 1000

10

20

30

40

50

60

70

80

90

100 TRANSIL assay 1:1 correlation

f b TR

AN

SIL

ass

ay

fb [%] equilibrium dialysis


TRANSIL- HSALot-to-lot reproducibility

good lot-to-lot reproducibility

0,0 0,1 0,2 0,3 0,4 0,5 0,6 0,7 0,80,95 0,96 0,97 0,98 0,99 1,000,00,10,20,30,40,50,60,70,8

0,95

0,96

0,97

0,98

0,99

1,00

lotA1 lotA2 lotA3 lotA4 lotA5 lotA6

1:1 correlation

f b l

ots

A1

- A

6

fb starting lot


TRANSIL-AssaysPrinciples


TRANSIL® Assays Available Formats

Type Compounds per 96-well plate

Compounds per 384-well plate

High-Throughput23 93

High-Precision 10 45

1E-7 1E-6 1E-5 1E-4 1E-3101520253035404550556065707580859095

100105

high affinity medium affinity low affinity

free

com

poun

d [%

]

protein or lipid concentration

1E-7 1E-6 1E-5 1E-4 1E-3101520253035404550556065707580859095

100105

high affinity medium affinity low affinity

free

com

poun

d [%

]

protein or lipid concentration

High-Throughput

High-Precision


???

Lipophilicity/Membrane Affinity ADME-

OptimisedCompounds

HSA Binding

logMA

Log Kd,HSA

PK-MapTM


PK-MapTM

...via Physiology-based ADME Models

PHYSIOLOGICAL PROPERTIES

organ volume and composition, blood flow rates, pH, effective accessible surface area, gastric emptying and intestinal transit time, feeding status,...

Physico-chemical-/in vitro properties

• membrane affinity• HSA binding• solubility• molecular weight• ...

Pharmacokinetic-/ADME-in vivo properties

• fraction dose absorbed• organ/plasma PC• free serum concentration• volume of distribution• ...


Validation of the Absorption Model

Human Fabs calculated only

from MA and Mw compared to published in vivo data

Willmann et al., J. Med. Chem. 2004,47, pp 4022-4032

Excellent agreement

All outliers known as substrates for active transporters.

Compound set (126 marketed drugs)


Validation of the Distribution Model

organ/plasmapartition coefficients

Korg/water


Combination of TRANSIL

and PK-MAP™*

as the most reliable toolfor ADME Prediction

PK-MAP™*

Lipophilicity/Membrane Affinity ADME-

OptimisedCompounds

HSA Binding

logMA

Additional Input

Solubility, MW

Log Kd,HSA

Conclusions


TRANSIL and TRANSIL-HSA Benefits and Summary

Determination of lipophilicity and serum binding in real High Throughput

Processing time is less than one minute per drug in the 384 well format

First 384 well assay for both parameters

Small amount of compound needed

Easy handling: Compound addition, separation, quantification

Correlation with established approaches and to ADME parameters

High throughput assays for high quality data

Documents

Modern Tools for Drug Discovery NIMBUS Biotechnology Modern Tools for Drug Discovery