Transcript No Slide Title

DEFINITION
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The automated synthesis of a large number of compounds in a
short time period using a defined reaction route and a large
variety of reactants
Normally carried out on small scale using solid phase synthesis
and automated synthetic machines
Parallel synthesis
• Single product formed in each reaction vessel
• Useful for SAR and drug optimisation
Synthesis of mixtures
• Mixtures of compounds formed in each reaction vessel
• Useful for finding lead compounds
SOLID PHASE TECHNIQUES
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Beads must be able to swell in the solvent used, and remain
stable
Most reactions occur in the bead interior
Swelling
Resin bead
Starting material,
reagents and solvent
Linkers
Merrifield resin for peptide synthesis (chloromethyl group)
= resin bead
Cl
HO2C
+
NHBoc
O
R H
NHBoc
Linker
R
HO2C
O
NH2
R
O
O
O
coupling
NH
R
H
H
NHBoc
R2
H
HF
O
O
aa1aa2aa3
H
NHBoc
R2 H
O
Deprotection
O
aan
NH2
Release from
solid support
OH
HO2C
aa1aa2aa3
Peptide
aan
NH2
Wang resin
OH
Linking functional group
O
Linker
Wang Resin
OH
Bead Linker
Wang resin
Carboxylic
acid
+
OH
HO2C
O
NH(Fmoc)
C
peptide
synthesis
NH2
R
R
H
O
TFA
cleavage
C
deprotection
R
O
O
NH(Fmoc)
piperidine
H
O
O
C
aa1aa2aa3
aan
H
Carboxylic
acid
HO2C
aa1aa2aa3
aan
NH2
Fmoc =
OH
O
O
NH2
Parallel Synthesis
Houghton’s Tea Bag Procedure
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Each tea bag contains beads and is labelled
Separate reactions are carried out on each tea bag
Combine tea bags for common reactions or work up
procedures
A single product is synthesised within each teabag
Different products are formed in different teabags
Economy of effort - e.g. combining tea bags for workups
Cheap and possible for any lab
Manual procedure and is not suitable for producing large
quantities of different products
Parallel Synthesis
Automated parallel synthesis
AUTOMATED SYNTHETIC MACHINES
Parallel Synthesis
Automated parallel synthesis of all 27 tripeptides from 3 amino acids
ETC
Parallel Synthesis
Automated parallel synthesis of all 27 tripeptides from 3 amino acids
27 TRIPEPTIDES
27 VIALS
4. Mixed Combinatorial Synthesis
The Mix and Split Method
Synthesis of all possible tripeptides using 3 amino acids
4. Mixed Combinatorial Synthesis
The Mix and Split Method
4. Mixed Combinatorial Synthesis
The Mix and Split Method
4. Mixed Combinatorial Synthesis
The Mix and Split Method
MIX
4. Mixed Combinatorial Synthesis
The Mix and Split Method
SPLIT
4. Mixed Combinatorial Synthesis
The Mix and Split Method
4. Mixed Combinatorial Synthesis
The Mix and Split Method
4. Mixed Combinatorial Synthesis
The Mix and Split Method
4. Mixed Combinatorial Synthesis
The Mix and Split Method
MIX
4. Mixed Combinatorial Synthesis
The Mix and Split Method
SPLIT
4. Mixed Combinatorial Synthesis
The Mix and Split Method
4. Mixed Combinatorial Synthesis
The Mix and Split Method
4. Mixed Combinatorial Synthesis
The Mix and Split Method
4. Mixed Combinatorial Synthesis
The Mix and Split Method
No. of
Tripeptides
9
9
9
4. Mixed Combinatorial Synthesis
The Mix and Split Method
No. of
Tripeptides
9
9
27 Tripeptides
3 Vials
9
4. Mixed Combinatorial Synthesis
The Mix and Split Method
TEST MIXTURES FOR ACTIVITY
4. Mixed Combinatorial Synthesis
The Mix and Split Method
Synthesise each tripeptide and test
5. Identification of structures from mixed combinatorial
synthesis
5.1 Recursive Deconvolution
• Method of identifying the active component in a mixture
• Quicker than separately synthesising all possible components
• Need to retain samples before each mix and split stage
Example
Consider all 27 tripeptides synthesised by the mix and split strategy
from glycine, alanine and valine
Gly
Gly
Ala
Ala
Val
Val
Mix and Split
Gly
Val
Gly
Ala
Ala
Gly
Gly
Val
Gly
Val
Val
Val
Gly
Ala
Ala
Gly
Gly
Ala
Ala
Gly
Gly
Ala
Val
Ala
Ala
Val
Ala
Val
Val
Val
All possible dipeptides in three vessels
Retain a sample from each vessel
Gly
Gly
Ala
Gly
Val
Gly
Gly
Ala
Ala
Ala
Val
Ala
Gly
Val
Ala
Val
Val
Val
Gly
Gly
Ala
Val
Gly
Gly
Ala
Ala
Gly
Val
Mix and
Split
Val
Gly
Ala
Val
Gly
Gly
Ala
Ala
Val
Gly
Val
Ala
Val
Val
Gly
Gly
Gly
Gly
Gly
Ala
Gly
Gly
Gly
Gly
Gly
Ala
Gly
Gly
Val
Ala
Ala
Val
Gly
Gly
Val
Ala
Val
Val
Gly
Gly
Gly
Ala
Ala
Ala
Ala
Val
Gly
Val
Ala
Val
Ala
Val
Ala
Val
Gly
Ala
Val
Val
Val
Val
Ala
Gly
Gly
Gly
Ala
Val
Ala
Val
Gly
Ala
Val
Val
Gly
Gly
Gly
Ala
Ala
Val
Val
Ala
Val
Gly
Ala
Gly
Gly
Ala
Gly
Val
Ala
Gly
Gly
Ala
Ala
Ala
Ala
Ala
Ala
Gly
Gly
Ala
Ala
Ala
Val
Ala
Val
Ala
Gly
Ala
Val
Gly
Ala
Val
Ala
Val
Val
All possible tripeptides in three vessels
Val
Gly
Val
Val
Ala
Val
Val
Ala
Val
Val
Val
Val
Val
5. Identification of structures from mixed combinatorial
synthesis
5.1 Recursive Deconvolution
Gly
Gly
Ala
Val
Gly
Gly
Ala
Ala
Val
Gly
Val
Ala
Val
Val
Mixture
Inactive
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Gly
Gly
Gly
Gly
Gly
Ala
Ala
Gly
Gly
Gly
Gly
Ala
Gly
Gly
Val
Ala
Val
Gly
Gly
Ala
Val
Gly
Gly
Val
Ala
Val
Val
Mixture
Inactive
Ala
Gly
Gly
Ala
Ala
Ala
Ala
Ala
Ala
Val
Gly
Gly
Gly
Ala
Ala
Ala
Val
Ala
Val
Ala
Gly
Ala
Val
Gly
Ala
Val
Ala
Val
Val
Val
Val
Ala
Val
Val
Ala
Val
Val
Val
Val
Mixture
Active
9 Possible tripeptides in active mixture
All end in valine
Add valine to the three retained dipeptide mixtures
Val
5. Identification of structures from mixed combinatorial
synthesis
5.1 Recursive Deconvolution
Gly
Gly
Gly
Ala
Val
Gly
Ala
Gly
Val
Val
Gly
Ala
Val
Gly
Val
Gly
Val
Ala
Ala
Val
Val
Gly
Val
Ala
Ala
Val
Val
Val
Val
Gly
Ala
Val
Ala
Ala
Val
Gly
Gly
Ala
Ala
Val
Val
Val
Val
Val
Val
Val
Val
Val
Active
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Active component narrowed down to one of three possible
tripeptides
Synthesise each tripeptide and test
5. Identification of structures from mixed combinatorial
synthesis
5.2 Tagging
SCAL = Safety CAtch Linker
H
N
H2N
O
NH2
HN
MeOS
NH
H
O
H
Tryptophan
O
SOMe
HN
O
Lysine
NH2
NH2
5.2 Tagging
Example
NH2
NH2
NH 2
NH2
RCHBrCO2H
NH
amino acid(aa 1)
R
Step 1
Tag 1
O
R
HN
amino acid(aa 2)
O
Br
Step 2
NH2
aa2
aa1
R"COCl
NH
NHR'
O
NH2
aa2
Tag 3
R
R
HN
HN
aa1
NH
R
O
NR'COR"
Step 3
aa3
amino acid(aa 3)
NH
NH2
Tag 2
O
R'NH2
aa2
aa1
NH
aa1
Br
NH2
HN
HN
aa1
NH
R
O
NR'COR"
NHR'
6. Identification of structures from combinatorial synthesis
LIGHT
6.2 Photolithography - example
LIGHT
NHX
NHX NHX
NHX
NHX
NHX
NHX NHX NHX
NHX
NHX
MASK 1
Mask
NHX NHX
NHX NHX NHX
NHX
NHX NHX
NHX NHX NHX
NHX
NHX
NHX NHX
NHX NHX
NHX
NHX
NHX
NH2
NH2
Deprotection
NHX NH2
NH2
NHX NHX
NHX
NH2
CO2H
coupling
NHX NHX
NHX NHX NHX
NHX
NHX
NHX
NHX NHX
NHX NHX
NHX NHX NHX
NHX
NHX
NHX
6. Identification of structures from combinatorial synthesis
6.2 Photolithography - example
Y
Y
Y
repeat
O
O
OMe
Y
O2N
amino acids
fluorescent tag
OMe
X= Nitroveratryloxycarbonyl
Target receptor
Y
7. Combinatorial synthesis
Heterocyclic synthesis - 1,4-benzodiazepines
R
R
NHFmoc
Te ntagel
resi n
O
R
NHFmoc
NH2
Piperidine
O
O
deprotecti on
X
Ar
Ar
O
R'
X=OH orCO2H
Fmoc=protecting group
R'
R'
O
O
NHFmoc
R
NH
NH
Piperidine
R
R
N
R'
N
N
1. Base
2. R"I
Alkylation
Ar
R"
O
R'
Cycli sation
O
Ar
R"
O
N
AcOH
deprotection
Ar
Ar
H
NH2
R
O
R
NHFmoc
F
Ar
O
N
TFA/H2O/Me2S
R'
Cleavage
N
X
Ar
Drawback:
Final product must contain X= OH or CO2H
7. Combinatorial synthesis
Heterocyclic synthesis - improved synthesis of benzodiazepines
R4
O
NH
NH
1
R
O
2
R
+
1
R
O
NH2
R3
Im i n e s
Am i no aci d
O
4

R
Cl(CH2)2Cl
NH
N
R2
R3
Adducts
TFA
R3
R4
N
O
R1
N
R2
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Functional group released from the resin takes part in the final
cyclisation
Does not remain as an extra, possibly redundant group
8. Planning a Combinatorial Synthesis
8.1 Aims
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To generate a large number of compounds
To generate a diverse range of compounds
Increase chances of finding a lead compound to fit a binding
site
Synthesis based on producing a molecular core or scaffold with
functionality attached
Centroid
or scaffold
Substituent
'arms'
Binding groups
8. Planning a Combinatorial Syntheses
8.1 Aims
Target molecules should obey Lipinski’s ‘Rule of Five’ for oral
activity
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a molecular weight less than 500
a calculated log P value less than +5
no more than 5 H-bond donating groups
no more than 10 H-bond accepting groups
8. Planning a Combinatorial Syntheses
8.2 Scaffolds
• ‘Spider’ scaffolds preferable for exploring conformational
space
• Allows variation of functional groups around whole molecule
to increase chances
of finding suitable binding interactions
Binding
regions
Screen compound
library
RECEPTO R
BINDING
SITE
Molecular weight of scaffold should be low to allow variation of
functionality, without getting products with a MWt > 500
8. Planning a Combinatorial Syntheses
8.2 Scaffolds
Tadpole scaffolds
- variation restricted to a specific region round the molecule
- less chance of favourable interactions with a binding site
'Spider' Scaffold with
'dispersed' substituents
'Tadpole' scaffold with
'restricted' substituents
Privileged scaffolds
- scaffolds which are common in medicinal chemistry and
which are associated with a diverse range of activities
- benzodiazepines, hydantoins, benzenesulphonamide etc
8. Planning a Combinatorial Syntheses
8.2 Scaffolds - examples
R"
R
O
O
N
R'
N
R1
Ar
Benzodiazepines
C
R2
R3
R4
HO2C
R2
N
N
R3
Hydantoins
O
R5
N
O
R2
O
R5
b-Lactams
R3
R
Pyridines
R4
R3
R1
R1
O
O
N
X
Me
R4
N
C
N
O
R2
Dipeptides
R6
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Good scaffolds
Spider like
Low molecular weight
Variety of synthetic routes available
8. Planning a Combinatorial Syntheses
8.2 Scaffolds - poor examples
OR5
O
R4O
OR1
R3O
OR2
Spider like and small molecular weight - good points
But multiple OH groups
Difficult to vary R1-R5 independently
Glucose
Me
Me
R1CO
R2
Steroid
O
R3
H 2N
R2
N
R
Indole
M.Wt. relatively high
Restricts no. of functional groups to keep MWt.< 500
Relatively few positions where substituents easily added
Tadpole like scaffold
Restricted region of variability
9. Dynamic combinatorial chemistry
Example - Ligands for carbonic anhydrase
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Reaction - reversible formation of imines
R
R
+
C O
H
Al de h yde
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•
H2N R'
Pri mary ami n e
C N
H
R'
Imine
•
H
R'
Imi n e
Reaction carried out in presence of carbonic anhydrase
Three aldehydes and four amines present as building blocks
Sodium cyanoborohydride added to ‘freeze’ the mixture
R
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C N
NaCNBH3
R
HC NH
H
R'
Se condary amine
Products quantified and identified
Experiment repeated in absence of target to identify amplified
product(s)
Amplified product is not necessarily present in greatest amounts
9. Dynamic combinatorial chemistry
Example - Ligands for carbonic anhydrase
•
Building blocks
O
HO3S
CHO
O
•
H2N
HO2C
O
H 2N S
O
NH2
CHO
NH
H2N
O
O
O
CHO
NH
H 2N
H2N
Am i n e s
HO2C
Al de hyde s
Amplified product
O
N
NaBH3CN
O
H2N S
H2N S
O
O
Acti ve com pou nd
HN
De rive d s e con dary am in e