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Tailoring Drug Development in the Cholinergic Nervous
System through Structural Templates
Three targets:
Nicotinic Acetylcholine Receptors (nAChR)
~15 subunits in various pentameric permutations
Acetylcholinesterase (AChE)
Alternatively spliced single gene
Neuroligin and Neurexin
Synaptic adhesion molecules
in situ, Click-Chemistry Leads to Drug
Candidates
(1) in situ Azide-Alkyne
Cycloaddition on
WT AChBPs
(2) in situ Azide-Alkyne
Cycloaddition on
MT AChBPs
Triazole Regioisomer
Identification
Cu++/Ru++ catalyzed lead
synthesis in microarray
AChBP
Binding Assay (SPA), Kd
X-Ray
Structure
with AChBPs
Whole Cell
Binding Assay
with nAChRS
(SPA), Kd
Concentration
dependence
functional responses
Interaction Energy
with crystallographic
model
Binding poses,
potential interacting
residues
Receptor Subtype
Occupation
Receptor Selectivity
Agonism (EC50) or
Antagonism (KA)
H3-Radioligand
Novel Drug Discovery & Development
• Synthesis by in situ freeze-frame, click chemistry
• Physical analysis of lead complexes (binding energy,
conformational changes, X-ray crystallography)
• Catalytic synthesis at the milligram level (refinement of SAR is
done in microarrays and only leads are taken forward).
• Quantitate affinity and activity in vitro
• Assay cellular toxicity and efficacy
• Efficacy in intact animals
• Pharmacokinetic studies of disposition in the target tissues
(CNS) and plasma
Mechanism and Therapeutic Indications
Acetylcholinesterase
• Mechanism
Antidotal Action at CNS and
skeletal muscle targets
Scavenging in environment,
portals of entry & circulation
• Therapeutic Indications
Organophosphate pesticide
toxicity
Nerve agents in terrorism
Nicotinic Acetylcholine Receptor
• Mechanism
Agonist, Partial Agonist and
Antagonist Activity at nAChRs.
Allosteric Modifiers at nAChRs
• Therapeutic Indications
Disorders of nervous system
development & aging dementias
Tobacco cessation
Click Chemistry TSRI
K. Barry Sharpless*
Valery Fokin*
Rakesh Sit*
Timo Weide
Joseph Fotsing
Neil Grimster
M.G. Finn, Georgia Tech*
Kasia Kaczanowska*
Gabi Amitai (IIBR)
Acetylcholine Binding Protein
Scott Hansen
Todd Talley
Ryan Hibbs
Zoran Radic’*
Ákos Nemecz*
John Yamauchi*
Nan Arunrungvichian*
Bill Chen*
AChE and AChBP
Crystal Structures
Scott Hansen
Ryan Hibbs
Todd Talley
Akos Nemecz
Kasia Kaczanowska*
Zoran Radic’*
Jean-Pierre Changeux*
Pascale Marchot (Marseille)*
Yves Bourne (Marseille)*
Andy McCammon(UCSD)
Jon Lindstrom (Penn)
Michal Harel (Weizmann)*
David Johnson (UC, Riverside)
Steve Sine (Mayo, Rochester)
Pharmacokinetics
John Casida (UC, Berkeley)
& Disposition
Zoran Radic*
Jordi Molgo (Gif-sur-Yvette)
Jeremiah Momper*
Mike McIntosh (Univ. of Utah)
Edmund Capparelli*
Toto Olivera (Univ. of Utah)
Manjunatha Kini (NUS)
Active center gorge
Peripheral
site
Active
center
CNS
O
P O
H3C
F
CH3
CH3
+
AChE-OH
O
P O
AChE-O
CH3
CH3
CH3
Blood-Brain Barrier
CH3
O
P O
CH3
H3C
F
+
AChE-OH
CH3
O
P O
CH3
AChE-O
CH3
+
R N OH
Plasma
CH3
O
P O
CH3
R N-O
CH3
+
AChE-OH
Reactivation of native peripheral AChE
Oxime-assisted Catalytic OP hydrolysis in Plasma
Alveolar Membrane
Site of Exposure
CH3
O
P O
CH3
H3C
F
CNS
O
P O
H3C
F
CH3
CH3
+
AChE-OH
O
P O
AChE-O
CH3
CH3
CH3
CH3
O
P O
CH3
H3C
F
+
AChE-OH
CH3
O
P O
CH3
AChE-O
CH3
x
Blood-Brain Barrier
HON
Plasma
N
Reactivation of native peripheral AChE
Oxime-assisted Catalytic OP hydrolysis in Plasma
Alveolar Membrane
Site of Exposure
CH3
O
P O
CH3
H3C
F
CH 3
O
P O
H3C
F
CH3
CH3
+
O
P O
O CH 3
N
R
CH3
AChE-OH
O
P O
CH
3
AChE-O
OH
CH
R3 N
+
ACh
+
H3
E-OH
CNS
C
NRR'
NRR'H
HON
HN
HON
H+
HN
+
O
O
x
Oxime Reactivation of native AChE in CNS
Blood-Brain Barrier
Plasma
NRR'
CH3
O
P O
CH3
H3C
F
+
AChE-OH
O
P O
AChE-O
CH3
CH3
CH3
NRR'H
HON
HN
HON
H+
HN
+
O
O
Reactivation of native peripheral AChE
Oxime-assisted Catalytic OP hydrolysis in Plasma
Alveolar Membrane
Site of Exposure
CH3
O
P O
CH3
H3C
F
AChBP: a structural surrogate of the
nAChR Ligand Binding Domain
5 subunits each of which have:
I.
A large extra cellular domain
II.
4 membrane-spanning regions
III.
α β combinations make up
most neuronal receptors
Hansen, S. et. al. 2005
Brejc, K. et. al. 2001
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