Advanced Medicinal
Chemistry
Lecture 2:
Finding a Lead
Dr Jeff Stonehouse
AstraZeneca R&D Charnwood
The Drug Discovery Process
Target
Identification
3 months to
2 years!
HTS
3-4 months
Active-to-Hit
(AtH)
3 months
Hit-to-Lead
(HtL)
6-9 months
New Lead
Optimisation
Projects (LO)
2 years
Candidate
Drug (CD)
Lead Compounds from a Variety of Sources
H
N
R
1. Chance Discovery
S
O
penicillins
N
O
O
OH
O
O
NH
O
O OH
O
2. Natural Products
HO
O
OH
taxol
O
H
O
O
O
O
O
3. Clinical Observation
O O
S
N
N
N
HN
N
N
O
4. Natural Ligands
5. Existing Drugs
6. High Throughput Screening (HTS)
Viagra
Natural Ligands
OH
HO
H
N
R=H adrenaline
R
R=Me noradrenaline
HO
Salbutamol
GlaxoSmithKline
Formoterol
AstraZeneca
H
HN
O
OH
OH
H
N
HO
H
N
HO
HO
O
Catechol
bioisostere
(toxicity)
Increased size
(selectivity and duration)
Catechol
bioisostere
(toxicity)
Increased size
(selectivity and duration)
Existing Drugs
Also known as the “Me-Too” or “Me-Better” Approach
Pfizer
Issues: short duration
O
O O
S
N
N
HN
N
N
N
Viagra
Multiple side effects and
incompatibility with other drugs
O
BEWARE: Patent Issues!!
Eli Lilly
O
N
Bayer
N
H
O
O O
S
N
N
HN
N
N
N
O
Levitra
O
Fewer side effects and
incompatibility with other drugs
Cialis
N
O
O
36h duration (“the weekend pill”)
High Throughput Screening (HTS)
“An industrialised process which brings together validated,
tractable targets and chemical diversity to rapidly identify
novel lead compounds for early phase drug discovery”
50-70% of new drug projects originate from a HTS
How?
•
validated, tractable targets
•
•
industrialised process
•
•
target selection for HTS
HTS assay technologies
and automation
chemical diversity
•
sample selection for HTS
Establishing a HTS
validated/
tractable
targets
HT Screen
Development
target
ID
O
O
human & pathogen
genomes
Cl
N
chemical
space
compound
collection
OH
compound
selection
Microtitre Plates – the HTS test tube
For 200K data points:
96
300-100ml
9mm pitch
125 x 1536
well plates
384
100-25ml
4.5mm pitch
1536
384LV
25-5ml
4.5mm pitch
10-1ml
2.25mm pitch
500 x 384 well plates
9mm
2000 x 96 well plates
Charnwood HTS Technologies; 1995-2001
30%
1%
1%
4%
2%
16%
19%
3%
24%
SPA
FLIPR
Filter
Fluorescence
Reporter
Yeast
TR-FRET
Alphascreen
FP
•Screening can utilise numerous
technologies e.g radioactivity,
fluorescence, luminescence
•None are universally applicable, each
with advantages and disadvantages
High throughput radioligand binding assays
Scintillation Proximity Assay – the first true homogeneous HTS screening technology
Molecule too far
away to activate
bead
Nothing bound
bead not activated,
I125
Bound molecule
Molecule binds
bead activated
I125
light produced
no light
Antibody/receptor
I125
Molecule cannot bind
Suitable for I125, 3H, 33P
I125
SPA (Scintillation Proximity Assay)
• First true homogeneous HTS technology
• Allows throughput of ~30K compounds/day in
384 format
• Easy to automate, no significant volume of
aqueous waste
BUT:
•Radioactive (safety headaches)
•Long read times (>30min/plate)
•Susceptible to quench artefacts
•Not applicable to all targets
FLIPR – a high throughput fluorimeter
Fluorescent Imaging Plate Reader
Real-time simultaneous imaging of 96- & 384-well plates
Used for HTS Ca2+ flux assays and ion channel screening
FLIPR – how it works
PC
96/384-Tip Pipettor
Drawer Holding
5 Microplates
6 W Argon Ion Laser
Cooled CCD Camera
• Cells loaded with fluorescent
dye sensitive to Ca2+ (fluo-3)
• CCD camera images base of
microtitre plate
• Addition of receptor agonist
stimulates Ca2+ release,
resulting in fluorescence
increase
• Whole plate is read
simultaneously, allowing
kinetic analysis
• ‘Functional’ screen (i.e.whole
cell) – greater relevance than
simpler screening methods
• Throughput is 1000x greater
than cuvette-based
fluorimeter assay
Establishing a HTS
validated/
tractable
targets
HT Screen
Development
target
ID
O
O
human & pathogen
genomes
Cl
N
chemical
space
compound
collection
OH
compound
selection
The AstraZeneca Compound Collection
1994
ASTRA ARCUS
ASTRA PAIN CONTROL
1993
1999
Ca 9% compound overlap
Not a recipe for an optimal screening bank
Compound Collection Enhancement
• AZ global initiative to boost screening collection
– Target: ensure viable Hits from 75% of AZ HTS
• Five-year initial lifespan. Two concurrent themes…
Acquisition
300K from 107 available
Stringent filters
Big Medchem input
Accept IP risks
Synthesis
Nominal 500K over 5 years
Target-class focus
Aligned to Research Areas
Early Bioscience input
CCE Structure
HTS
Charnwood
GPCR
Kinase
Charnwood
Alderley
Park
HTS
AP
~60 Scientists
Med Chem
Bioscience
Comp Chem
Informatics
Central
Bioscience
Cheminformatics
Channel
Protease
HTS
Mölndal
Mölndal
Compound
Management
AP
Södertälje
HTS
US
• Chemistry deliberately embedded in Research Areas
– Not centralised
– Benefit of Project exposure
– Feeds parallel synthesis skill back into projects
CCE – Library Chemistry
Types of reactions
amide coupling
sulphonamide formation
3 most commonly used reactions-
reductive amination
Boronic acid coupling
aminopyrazoles
Amide coupling
imidazopyridines
Multicomponent reaction (3 variants so far)
imidazothiazoles
Sulphonamide arylation
Reductive amination
imidazopyrimidines
Ester hydrolysis
Acyl sulphonamide formation
Urea formation
Sulphonamide formation
aminothiazoles
aminooxadiazoles
triazolopyrimidines
Epoxide opening
aminotriazoles
Anhydride opening
aminobenzimidazoles
Condensation to form benzamidazoles
triazolopyridines
Mitsunobu
pyrazolopyrimidine
N-, O- and S-Alkylation
3-aminoquinolines
Sulfonylurea formation
benzoxazinone formation
Pyridone formation
tetrazole formation
Boc or t-butyl deprotection
cyclization to heterocycles (21 types - see list)
Nucleophilic aromatic substitutions (2 types)
triazolopyridazines
triazolopyrazines
thiazolidin-4-one
3-amino-1,2,4-triazoles
pyrimidin-2-ones
triazolo[1,5-c]quinazoline
imidazolidin-2-one
quinazolinone
1,2,4-oxadiazole
CCE – Common Combinatorial Reactions
• Amide Coupling
R
1
N
2
R
HATU, Et3N
O
H
+
HO
R
O
R
1
3
NMP
N
2
R
PF 6-
N
R
3
N
N
N
O
HATU
+
N
N
• Sulphonamide Formation
R
1
N
2
R
O
H
+
Cl
Et3N
O
S
R
O
R
1
N
2
R
3
NMP
O
S
R
O
• Reductive Amination
R
O
1
N
2
R
Na(AcO)3BH
H
+
H
R
3
AcOH, NMP
R
1
N
2
R
N
3
R
3
NMP
Mechanism
PF -
PF 6-
6
+
N
Amide
Coupling
N
O
N
N
N O
+H+
R
HO
N
3
N
+
O
O
R
O
+H+
3
-H+
-H+
N
R
O
Sulphonamide
Formation
Cl
R
1
N
2
R
1
N
2
R
R
O
-H+
3
R
H
R
N
2
R
3
H
O
S
R
1
1
N
2
R
O
S
R
+
+
3
N
H Cl
+
Na
O
Reductive
Amination
+
H
+
H
-H+
O
R
H
1
N
2
R
H
O
O
O B O
H
O
R
3
OH
R
-H2O
1
N
2
R
R
3
R
1
+
N
2
R
R
3
R
1
N
2
R
R
3