This document discusses opportunities for developing novel compounds targeting medically relevant protein families using non-mainstream scaffolds. It notes 400 novel scaffolds in a European compound library that exhibit structural complexity, stereochemistry, scaffold diversity, and challenging chemistry. The document discusses opportunities for targeting G protein-coupled receptors, kinases, and protein-protein interactions using spirocyclic, anellated, and DNA-encoded scaffold libraries. It emphasizes the potential of these approaches for developing compounds with improved target residence time and selectivity profiles through disrupting a protein's hydrophobic spine formation.

1. Target Family-Centric Privileged Structures:
unexploited opportunities
for medicinal chemistry
Gerhard Müller
Senior Vice President
Medicinal Chemistry
Mercachem bv
gerhard.mueller@mercachem.com

2. • Bayer Pharma AG
• AstraZeneca AB
• H. Lundbeck A/S
• Janssen Pharma NV
• Merck KGaA
• Sanofi
• UCB Pharma SA
300.000 compound library
to be complemented with
200.000 novel compounds
400 novel scaffolds
 non-mainstream
 structural complexity
 stereochemistry
 scaffold diversity
 challenging chemistry
€196 million pan-European drug discovery platform
WP9: Program Recruitment
(crowd-) sourcing of proposals from e.g. academia
WP10: Review & Selection
Library Selection Committee
(Prof. Adam Nelson)
WP11: Experimental Validation
Validation Management Team
(Mercachem)
WP12: Library Generation
5 SMEs involved, each SME:
8000 compounds annually based on
16 scaffolds annually
European Lead Factory – 200.000 novel „non-main stream“ compounds
attempts to surf the chemical space
2
as of May 1st:
680 scaffolds reviewed
> 390 scaffolds approved
~ 140 scaffolds validated
~50.000 compounds made

3. “What is clear is that certain “privileged structures“
are capable of providing useful ligands for more than
one receptor …“
“… judicious modification of such structures could be a viable
alternative in the search for new receptor agonists and
antagonists”
B. E. Evans et al. J. Med. Chem., 31, 2235 (1988)
A. A. Patchett et al. Annu. Rep. Med. Chem. 35, 289 (2000)
G. Müller, Drug Discovery Today 8, 681-691 (2003)
one ligand for more than one target system – very generic – no teaching!
navigating with privileged structures
N
N
N
H
O
S
F
tifluadom
k-opioid agonist
Nature, 298, 759 (1982)
CCK-A antagonist
Neurosci. Lett., 72, 211 (1982)
3
N
H
N
O
N
OH
N
N
O
Merck:
ORL1 antagonist
Roche:
ORL1 agonists
N
N
H
N
O
N N
NH
delete
bond
Dan Rich, 1990’s
Prevent hydrophobic collapse

4. Spirocyclic compounds:
 conformationally constrained  -TDS 
 frequently occur in natural products  protein binding 
 characteristic three-dimensionality  shape 
 structural novelty  IP 
 Fsp3-rich carbon skeleton
 increasingly utilized in drug discovery
 numerous therapeutical areas, often CNS
 majority of spiro chem space unexplored!
1857 compounds
active at 200 targets
75 GPCRs (!)
quantitative analysis of molecular spiro topologies
spiro systems
4
CGRP-1
V1a, V1b, V2
sstr1, sstr2, sstr3, sstr4, sstr5
A2a, A3
Nociceptin
Oxytocin
Orexin-1
µ-opioid, k-opioid, d-opioid
s-opioid
H3, H4
C5a
GPCR-35
M1, M2, M3, M4, M5
Calcitonin-1
D1, D2, D3, D4, D5
a1a, a1b, a1d, a2a, a2b, a2d
b1 adrenergic
CB1, CB2
B1, B2
5-HT1a, 5-HT1b, 5-HT1d, 5-HT2a,
5-HT2b, 5-HT2c, 5-HT1a, 5-HT5a,
5-HT6, 5-HT7
MC1, MC3, MC4, MC5
mGluR5
MCHR-1, MCHR-2
Vanilloid
Neuromedin B
CGRP-1
NPY-2, NPY-5
CCR-1, CCR-2, CCR-3, CCR-5
NK1, NK2, NK3
Glucagon
Melatonin-1B
Prof. J. Bajorath

5. J. Mol. Biol., 425, 662-677 (2013)
Convert GPCRs to soluble protein – biochemistry, biophysics, etc
G7 Therapeutics
5
Long –term apo-state stability:
No stabilizing ligand required
– no pre-empted activation state
Important for accessibility of fragments
Unbiased on functional signature
(ago, ant, inv.ago, etc)
Detergent solubility:
Isolated receptor protein in detergent micelles
Vapor-diffusion crystallization works out
Protein amenable to biochemistry and biophysics
SPR, NMR, TINS, FBLG, kinetics, thermodynamics, etc.
CHESS:
Cellular High-Throughput Encapsulation Solubilising and Screening
Class A GPCRs
 Neurotensin 1 receptor (NTS1)
 κ-opioid (KOR)
 Tachykinin receptor 1 (NK1)
 Oxytocin
 Adrenergic receptors
 α1A
 α1B
Class B GPCR
 Parathyroid hormone receptor 1
(PTH1)
Fig. S2. Quality of the σA-weighted 2FO-FCelectron density map contoured at 1.2 σ. Stere
Directed evolution – error-prone PCR

6. O
N
O
O
Annelated ring systems – novelty analysis ongoing
Fsp3-rich carbon skeletons
annelation-constrained macrocycles
• sp3-rich
• stereochemical complexity
• 3D shape
• non-main stream
6
annelated
3796 compounds
395 BM scaffolds
401 unique targets
[5:5] cyclopentan-pyrolidine [6:5] piperidino-pyrrolidine
[5:5] pyrrolidino-diazolidine-dione g-exo-homo Proline
[6:7] reverse turn mimic
[6:5] reverse turn mimic

7. P. Furet et al. J. Comp.-Aid. Mol Des. 1995, 9, 465-472
P. Traxler, Exp.Opin. Ther. Patents 1998, 8, 1599-1625
N
NN
H
N
H
O
N
N
N
stabilizing inactive kinase conformation
NH
N
NH
O
O
H
O
deep
pocket
Gleevec : c-Abl
type II
NH
N
NH
O
O
H
O
back pocket
N
Cl
Cl
O
N
NN
H
S
competitive inhibition of active state
PD173955 : c-Abl
1m52.pdb
PD173955 : c-Abl
1iep.pdb
Gleevec : c-Abl
DFG-in (I) vs. DFG-out (II)
activation loop undergoes significant structural rearrangement
type I
DFG-in
DFG-out
7

8. DFGin
DFGout
many „DFG-in-between“ ?
misleading terminology
A.P. Kornev, N. M. Haste, S. S. Taylor, L. F. Ten Eyck, Proc. Natl. Acad. Sci., 103, 17783 (2006)
A.P. Kornev, S. S. Taylor, L. F. Ten Eyck, Proc. Natl. Acad. Sci., 105, 14377 (2008)
A.P. Kornev, S. S. Taylor, Biochim. Biophys. Acta, 1804, 440-444 (2010)
H.S. Meharena, et al, PLOS Biology, 11 (10), 1-11 (2013) 8

9. hydrophobic spine
comparison of intact and disrupted spines
O
N
N
H
O
N
H
N
H
O
Cl
CF3
intact R-spine disrupted R-spine (CDK8)
9
active kinase
type I inhibitors
fast kinetics
inactive kinase
non-type I inhibitors
slow kinetics (t½ 8 h)

10. Retro-Design Approach: B2F (back-to-front)
 Sets out with scaffolds rather than leads
 Disrupt hydrophobic spine
 Long residence time on target; slow koff
 Option for exploration of novel IP space
 Option for increased selectivity
NH
N
NH
O
O
H
O
“Retro Design“ approach
targeting conformational states by deep pocket-directed scaffolds
10

11. UPR as one major cause for neurodegeneration
AD disease mechanisms: UPR
ER
UPR (unfolded protein response):
• cellular stress pathway
• protein misfolding in ER = stress
• stress sensors:
• PERK, ATF6, IRE1
• „sense“ mis-folded proteins
• expression of Chaperons – assist in protein folding
11
pPERK
Phospho-Tau (AT8)
Hippocampus anatomy
PERK

12. N
N N
NH2
N
O
N
CH3
F
GSK2656157 (IC50: 0.8 nM)
development candidate
WO2011119663A1
J.Med.Chem. 2012, 55, 7193-7207
Med.Chem.Lett. 2013, 4 (10), pp 964–968
IC50: 2.72 nM
Novel, IP-free scaffold
12
4x7n4g313qd2
R-Sp2 and R-Sp3 inhibition mode = f(inhibitor)
hydrophobic spine topology

13. CDK8 in oncology
marked modulation of selectivity profile within deep pocket - controversial
Steady increase in selectivity throughout the consecutive compound generations
Initial generation Novel generation Latest generation
13
See poster 25

14. 14
IC50(M)
similar potency;
huge difference in residence time
similar residence time;
difference in potency
Residence time (h)
similar potency;
huge difference in residence time
Escape Trajectory: retrograde induced-fit mechanims of dissociation
IC50 / koff Correlation (?)
With appreciation to R. Buijsman, NTRC, Oss, NL
Ponatinib
d
on
off
K
k
k

P + L
PL
ΔG‡
on
ΔGd
ΔG‡
off
RT/G-
on
‡
on
ek D

RT/G-
off
‡
off
ek D

/RTG-Δ
d eK d

Bindingcoordinate

15. 15
compound synthesis in DNA-cavity
 Holliday Junction
50 Mio compound library
linear and branched compounds
spiked with PPI motifs
• turns (beta and gamma)
• strands (exteded conformations)
• Trp, Arg, Tyr
Protein-Protein Interaction-targeted Library
DNA-encoded Library Technology

16. p38a as a prototypic kinase amenable to type II inhibition
Tri-peptide mimetics for kinases ?
16
IC50>2µM
% remaining activity @ 2µM compound concentration
0
25
50
75
100

17. Hydrophobic R-spine disrupted
17
Vpc00628
Hinge binding via single hydrogen bond – DFG out
Stephan Knapp, SGC Oxford, UK

18. Structurally Related Inhibitors
18

19. rapid interrogation of SAR by point mutations in compound structure
Advantage of modular chemistry
N
H
O
N
N
NH2
O
N
H
NH2
O
required ?
R-groups
heterocycles
ring size
acceptor !
ring size
heterocycles
R-groups
required ?
NH-Me, Cl, F
Me, CF3
stabilize kinked cofo!
stereochemistry
small substituents
spacing
heterocycles
R-groups
ring size
scan pocket better
change physchem
stabilize cofo
donor required !
spacing, b-AA
NH-R, explore pocket
Weeks 1 2 3 4 5 6
step 1: amide coupling
step 1: purification
Step 2: Pg removal
Step 2: purification
Step 3: amide coupling
Step 4: purification
Unfold near-to-complete SAR in 6 weeks
of chemistry
Iterative analogue libraries
19

20. summary
• pharmaceutically relevant targets cluster into densely
populated families
• high Fsp3 compounds pursuing an indirect approach
chemical similarity correlates with biological similarity
• target family-centric rationales allow for imprinting
family-wide commonalities into new scaffolds
• functional attributes can be pre-engineered
you might want to look for slow-off compounds
• despite the run on epigenetic targets, the days of kinases
are not yet over
• DNA-encoded library approaches uncover truly novel
chemotypes for target families
20
consider binding kinetics
optimize residence time for multiple reasons
Bob Copeland (CSO Epizyme Inc.)
10th Swiss med chem course, Leysin, 2012
„any chemist reporting IC50‘s should
be boiled in oil!“

21. www.mercachem.com
Arjen Brussard
Tim Moser
Christoph Schächtele
Michael Kubbutat
Nils Jakob Vesten Hansen
Tara Heitner Hansen
Kiyoshi Takayama
Tomoko Shimizu
Jürgen Bajorath
Dagmar Stumpfe
Carlo Bertozzi
Lutz Kummer