Pharmacoinformatic Approaches to

target polyspecific Proteins

Safety Sciences on the Molecular Level

Gerhard F. Ecker
Emerging Field Pharmacoinformatics
Department of Medicinal Chemistry, University of Vienna
Althanstrasse 14, A-1090 Wien, Austria
gerhard.f.ecker@univie.ac.at; http://homepage.univie.ac.at/gerhard.f.ecker

G. Ecker
 Pharmacoinformatics

Identification Preclinical Clinical Post-

Target Target
& obtention of marketing
identification validation assessment trials
compounds monitoring

Computational Virtual Structure-based ADMET Medical

genomics screening drug design prediction Informatics

Computational Biosimulation
systems biology

Based on the molecular structure

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Drug Development - Failures

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 Paradigm Shifts
• One drug – one target
• One drug – multiple targets (kinases)

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 Paradigm Shifts
• Many drugs – one target (hERG channel)

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 Paradigm Shifts
• Many drugs – many targets (ABC-transporter)

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Chemogenomics

H. Kubinyi, Vienna 2006

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 Mapping the Chemical Space onto
the Biological Space
• Calculate pair wise similarity of all compounds
active on a given target
• Compare the average values with the
“background” of the whole drug like chemical
space
• Requires approx. 20 mio x 20 mio similarity
calculations

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 The Systems Biology View
• ABC-transporter and Cytochromes are regulated
by nuclear receptors
• ABC-transporter and CYPs show overlapping
substrate specificity
• Selectivity has to be addressed on a systems level
rather than on individual targets

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A Network of Targets

Ekins S, unpublished
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 The ABC of Drug Transport
• membrane-bound efflux pumps
• energy driven (ATP)
• ATP Binding Cassette (ABC-transporter)
• 48 ABC transporter in humans
– P-glycoprotein (P-gp, ABCB1)
– Multidrug Resistance Related Protein (MRP, ABCC1)
– Breast Cancer Resistance Protein (BCRP, MXR,
ABCG2)
• a lot of analogous transport proteins in bacteria, fungi,
protozoes and plants
very often multispecific in ligand recognition
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 P-Glycoprotein
• 170 kD
• 2 transmembrane domains
• 2 ATP-binding sites
• Xenotoxin transporter
• hydrophobic vacuum cleaner
• intestine, liver, kidney
• blood-brain barrier
• tumors

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Substrates of P-Glycoprotein I
Vinca Alkaloides Epipodophyllotoxines
Vincristine Etoposid
Vinblastine Teniposid

Anthracyclines Taxanes
Doxorubicine
Daunorubicine Actinomycin D

C.R. Leveille-Webster, I.M. Arias, J. Membrane Biol. 143, 89-102 (1995)

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 Inhibitors of P-Glykoprotein
• Calcium Antagonists • Cyclic Peptides
– Verapamil – Cyclosporin A
– Niguldipin – Valspodar
• Protein Kinase C Inhib. – FK 506
– Calphostin C – Rapamycin
– Staurosporin • Miscellanous
• Calmodulin-Antagonists – Dipyridamol
– Fluphenazin – Quinidin
– Trifluoperazin – Amiodaron
– trans-Flupenthixol – Reserpin

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Physiological Function of P-Glycoprotein
• protection from toxins
– blood brain barrier
– testis
– placenta
• excretion of toxins
– liver
– gastrointestinal tract
– kidney
Levin, J. Med. Chem. 23, 682 (1980)

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Bioavailability and Transporter

Kim, Mol Pharm, 3, 26 (2006) G. Ecker

 Multispecificity - Challenges
• Structure and function on the molecular level
– kinetic studies
– photoaffinity labeling
– site specific mutagenesis
– protein homology modeling
• Design of inhibitors
– promiscuity
• ADMET profiling
– substrate yes/no?

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 Activity of Benzofurane-Analogs
OH
O NR

R1
log(1/EC50)

O OH
NR

R1
log(1/EC50) = 0.86 logP - 1.16 IBf - 3.33 (Q2cv = 0.97)

logP ber. J. Csöllei, J. Med. Chem. 39, 4767-4774 (1996)

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 P-Glycoprotein - the Target

Optimal distance: 3.5 -CH2-

π−π-interaction
OH high partial logP
O N
steric interactions
O
H-bond acceptor

H-bond acceptor

• Hansch-Analysis
• Free-Wilson Analysis π−π-interaction

• Topliss-Approach
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 Pharmacophoric Features
15 diverse compounds
multiconformational-DB
Mapping of the features:
hydrophobic
aromatic
H-bond acceptor
positive ionizable

Chemical function based pharmacophoric features - CATALYST

Langer, Arch. Pharm. 337, 317
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Screen of the World Drug Index
F NO2
HO
HO
OH O O
N N
O N O O

F N
Terfenadine H
AHR-16303B Benidipine
O

OH
H
N
O N HO
O N
N
OH N HO O
N
N
N

CP-215548 CGP-54103
CP-117227

N
N OH

N
O HN
N N CF3 O
HO

S Cl N
O
Asocainol Homofenazine Mepacrine

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 Virtual Screen of the SPECS
Compound Library
• increase network size to 250 x 250
• projection of the propafenons together with the
SPECS database (134.767 compounds)
• analyse co-localisations

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 Kohonen Maps - Hits
O O
N
N N
O N
N N
N N
N N
O N
N
S S
O
Cl
AG-690/11972772 AG-690/12887361 AJ-131/15197008 AJ-292/13162028

N O NH2
O O
N
O N
N S N N
N S O
N S
S
N

AJ-292/15089034 AN-989/14669159 AO-364/14480185

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Benzophenones as tool for characterisation of the
“Propafenon-Site” on P-glycoprotein

z high activation wavelength (>340nm)

z short half life (10-13 sec) in presence of OH
C-H groups O NR
z repeated activation cycles possible
O
z photoactivation takes place on a
pharmacophoric sub-structure

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Consensus-labeling Regions

Ecker, Mol. Pharmacol. 61, 637 (2002)

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Extension to P-glycoprotein

Pleban, Mol Pharmacol 67, 365 (2005)

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 Selectivity Profiling B1/C1/G2
• ABCB1, ABCC1 and ABCG2 have different
physiological functions and tissue distribution
• in part overlapping substrate specificity
• Polyspecific inhibitors might have advantages in
tumor therapy
• Selective inhibitors might overcome side effects
due to physiological functions
• Problem of promiscuity!

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Selectivity Profiling B1/G2

ABCB1 ABCG2

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 Selectivity Profiling
ABCB1 vs. ABCG2
2
• ABCG2 (MXR, BCRP) is
responsible for resistance to 1

methotrexate in breast cancer

log potency ABCG2 (R482R)

• propafenones inhibit both pumps 0

• selectivity up to 1000 fold -1

• main difference: H-bond acceptor

strenght in the vicinity of the -2

nitrogen-atom
-3
-3 -2 -1 0 1 2
log potency ABCB1

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Selectivity Profiling B1/G2

ABCB1 ABCG2

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 Computational
Computational Sciences CPAMMS Life Sciences

LEWI-GRID PiP-Vie
Docking Emerging Field Compunds
MD-Simulations Properties
Binding free energy
Pharmacoinformatics Reverse Screening

MUW
Joint Publications/Projects Barcelona
Faculty of Informatics C.R.Noe: BBB,NMDA,Polyamines
T.Erker: COX Perugia
W.Holzer: P-gp Erlangen
W.Jäger: CYP
S.Hering: hERG channel G. Ecker
 EUROPIN – A European
Pharmacoinformatics Initiative
EUROPIN brings together six institutions from five European
Countries to develop a joint PhD program on Pharmacoinformatics in
order to promote training and education of top level scientists in this
field. It will provide a platform for training through research focusing
on methods and applications used in computational drug design and
development.
University of Vienna (coordinator)
Pompeu Fabra University, Barcelona
Gdansk University of Technology
Martin-Luther-University Halle-Wittenberg
University of Parma
University of Perugia

G. Ecker
 EUROPIN - Objective
Core objectives are
• to promote creativity, competitiveness, employability
and entrepreneurial spirit of young scientists.
These will be achieved
• via strong emphasis on cooperation and mobility of
students and teaching staff,
• by use of intensive programmes and workshops with
strong participation of experts from Pharmaceutical
Companies,
• and by extensive use of web-based virtual training
activities.

G. Ecker
 Thank you!
Dominik Kaiser Wilfried Gansterer
Barbara Zdrazil Peter Chiba
Karin Pleban Michael Gottesman
Michael Demel Roberto Pellicciari
Rita Schwaha Ferran Sanz
Khac-Minh Thai Johnny Gasteiger
Lars Richter
Andrea Schiesaro
Daniela Digles
Yogesh Aher

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