DEFINITION The automated synthesis of a large number of compounds in a short time period using a defined reaction route and a large variety of reactantsThe

DEFINITIONDEFINITION

• The automated synthesis of a large number of compounds in a The automated synthesis of a large number of compounds in a short time period using a defined reaction route and a large short time period using a defined reaction route and a large variety of reactantsvariety of reactants

• Normally carried out on small scale using solid phase synthesis Normally carried out on small scale using solid phase synthesis and automated synthetic machinesand automated synthetic machines

Parallel synthesisParallel synthesis • Single product formed in each reaction vesselSingle product formed in each reaction vessel• Useful for SAR and drug optimisationUseful for SAR and drug optimisation

Synthesis of mixturesSynthesis of mixtures• Mixtures of compounds formed in each reaction vesselMixtures of compounds formed in each reaction vessel• Useful for finding lead compoundsUseful for finding lead compounds

Starting material,reagents and solvent

Swelling

Linkers

SOLID PHASE TECHNIQUESSOLID PHASE TECHNIQUES

• Beads must be able to swell in the solvent used, and remain Beads must be able to swell in the solvent used, and remain stablestable

• Most reactions occur in the bead interiorMost reactions occur in the bead interior

Resin bead

Deprotection

O

aa1aa2aa3

O

aan NH2

Merrifield resin for peptide synthesis (chloromethyl group)Merrifield resin for peptide synthesis (chloromethyl group)

O

O

R

NHBoc

H

O

O

R

NH2

H

HO2C NHBoc

R2H

coupling

O

O

R

NH

H

O

NHBoc

R2H

= resin bead

Cl HO2C NHBoc

R H+

Linker

HF

OH

aa1aa2aa3 aanHO2C NH2

Peptide

Release from solid support

Linking functional groupO

OH

Wang Resin

Wang resinWang resin

Linker

OH

Bead Linker

peptidesynthesis

O Caa1aa2aa3

O

aan NH2

TFAcleavage

OH

aa1aa2aa3 aan NH2HO2C

Fmoc =

O

O

Wang resinWang resin

OH

Carboxylic Carboxylic acidacid

Carboxylic Carboxylic acidacid

HO2C NH(Fmoc)

R H

+ O C NH(Fmoc)

R H

O

O C NH2

R H

O

piperidine

deprotection

• Each tea bag contains beads and is labelledEach tea bag contains beads and is labelled

• Separate reactions are carried out on each tea bagSeparate reactions are carried out on each tea bag

• Combine tea bags for common reactions or work up Combine tea bags for common reactions or work up proceduresprocedures

• A single product is synthesised within each teabagA single product is synthesised within each teabag

• Different products are formed in different teabagsDifferent products are formed in different teabags

• Economy of effort - e.g. combining tea bags for workupsEconomy of effort - e.g. combining tea bags for workups

• Cheap and possible for any labCheap and possible for any lab

• Manual procedure and is not suitable for producing large Manual procedure and is not suitable for producing large quantities of different productsquantities of different products

Parallel SynthesisParallel SynthesisHoughton’s Tea Bag ProcedureHoughton’s Tea Bag Procedure

22

AUTOMATED SYNTHETIC MACHINES

Parallel SynthesisParallel SynthesisAutomated parallel synthesisAutomated parallel synthesis

ETC

Parallel SynthesisParallel SynthesisAutomated parallel synthesis of all 27 tripeptides from 3 amino acidsAutomated parallel synthesis of all 27 tripeptides from 3 amino acids

27 TRIPEPTIDES

27 VIALS

Parallel SynthesisParallel SynthesisAutomated parallel synthesis of all 27 tripeptides from 3 amino acidsAutomated parallel synthesis of all 27 tripeptides from 3 amino acids

Synthesis of all possible tripeptides using 3 amino acidsSynthesis of all possible tripeptides using 3 amino acids

4. Mixed Combinatorial Synthesis4. Mixed Combinatorial SynthesisThe Mix and Split MethodThe Mix and Split Method



MIXMIX


SPLITSPLIT





MIXMIX


SPLITSPLIT





No. of Tripeptides

9 9 9


No. of Tripeptides

9 9 9

27 Tripeptides 3 Vials


TEST MIXTURES FOR ACTIVITYTEST MIXTURES FOR ACTIVITY


Synthesise each tripeptide and testSynthesise each tripeptide and test


5.1 Recursive Deconvolution5.1 Recursive Deconvolution• Method of identifying the active component in a mixtureMethod of identifying the active component in a mixture• Quicker than separately synthesising all possible componentsQuicker than separately synthesising all possible components• Need to retain samples before each mix and split stageNeed to retain samples before each mix and split stage

Example Example Consider all 27 tripeptides synthesised by the mix and split strategy Consider all 27 tripeptides synthesised by the mix and split strategy from glycine, alanine and valinefrom glycine, alanine and valine

5. Identification of structures from mixed 5. Identification of structures from mixed combinatorial synthesiscombinatorial synthesis

Gly

Ala

Val

Gly

Ala

Val

Gly

AlaVal Gly

AlaVal Gly

AlaVal

Gly Ala Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Mix and Split

All possible dipeptides in three vesselsAll possible dipeptides in three vesselsRetain a sample from each vesselRetain a sample from each vessel

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly Ala Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Gly

Gly

Gly

Gly

Gly

Gly

Gly

Gly

Ala

Ala

Ala

Ala

Ala

Ala

Ala

Ala

Ala

Val

Val

Val

Val

Val

Val

Val

Val

Val

Mix andSplit

All possible tripeptides in three vesselsAll possible tripeptides in three vessels


Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Ala

Val

Gly

Gly

Gly

Gly

Ala

Val

Ala

Ala

Ala

Gly

Ala

Val

Val

Val

Val

Gly

Gly

Gly

Gly

Gly

Gly

Gly

Gly

Gly

Ala

Ala

Ala

Ala

Ala

Ala

Ala

Ala

Ala

Val

Val

Val

Val

Val

Val

Val

Val

Val

MixtureInactive

MixtureInactive

MixtureActive

• 9 Possible tripeptides in active mixture9 Possible tripeptides in active mixture• All end in valineAll end in valine• Add valine to the three retained dipeptide mixturesAdd valine to the three retained dipeptide mixtures

5.1 Recursive Deconvolution5.1 Recursive Deconvolution

Gly

Ala

Val

Gly

Ala

Val

Gly

Ala

Val

Gly

Gly

Gly

Ala

Ala

Ala

Val

Val

Val

Val Val Val

Gly

Ala

Val

Gly

Ala

Val

Gly

Ala

Val

Gly

Gly

Gly

Ala

Ala

Ala

Val

Val

Val

Val

Val

Val

Val

Val

Val

Val

Val

Val

Active

• Active component narrowed down to one of three possible Active component narrowed down to one of three possible tripeptidestripeptides

• Synthesise each tripeptide and testSynthesise each tripeptide and test

5. Identification of structures from mixed 5. Identification of structures from mixed combinatorial synthesiscombinatorial synthesis5.1 Recursive Deconvolution5.1 Recursive Deconvolution

NH

O NH2

HN

O

O

HN

MeOS

SOMe

O

H2N

HN

H

H

Lysine

Tryptophan

NH2

NH2

5.2 Tagging5.2 Tagging

SCAL = Safety CAtch LinkerSCAL = Safety CAtch Linker


NH2

NH2

RCHBrCO2H

Step 1

NH2

NH

O

R

Br

amino acid(aa 1)

Tag 1

HN

NH

O

R

Br

aa1

NH2

R'NH2

Step 2

HN

NH

O

R

NHR'

aa1

NH2

amino acid(aa 2)

Tag 2

HN

NH

O

R

NHR'

aa1

aa2

NH2

R"COCl

Step 3

HN

NH

O

R

NR'COR"

aa1

aa2

NH2

amino acid(aa 3)

Tag 3

HN

NH

O

R

NR'COR"

aa1

aa2

aa3 NH2

5.2 Tagging5.2 Tagging

ExampleExample

NHX NHX NHX NHX NHX

NHXNHXNHXNHXNHXNHX

NHXNHXNHXNHXNHXNHX

6. Identification of structures from combinatorial 6. Identification of structures from combinatorial synthesissynthesis6.2 Photolithography - example6.2 Photolithography - example

NHXNHXNHXNHXNHXNHX

MASK 1

Mask

LIGHT

LIGHT

NHXNHXNHXNHX

NHXNHXNHXNHXNHXNHX

NHX

CO2H

coupling

NHX NHX

NHX NHX NHX

NHX NHXNHX NHX NH2 NH2 NH2

NHXNHXNHXNH2NH2NHX

NHXNHXNHXNHXNHXNHX

Deprotection

Y Y Y

repeat

6. Identification of structures from combinatorial 6. Identification of structures from combinatorial synthesissynthesis6.2 Photolithography - example6.2 Photolithography - example

Y

amino acids

OMe

O MeO

O2N

O

X= Nitroveratryloxycarbonyl

fluorescent tag

Target receptor

Y

7. Combinatorial synthesis7. Combinatorial synthesisHeterocyclic synthesis - 1,4-benzodiazepinesHeterocyclic synthesis - 1,4-benzodiazepines

O

Ar

NHFmocR

X

X=OH or CO2HFmoc=protecting group

NHFmoc

R'

O

F

1. Base2. R"I

Alkylation

TFA/H2O/Me2S

Cleavage

R

X

N

N

Ar

R'

OR"

Tentagelresin O

Ar

NHFmocR

Piperidine

deprotectionO

Ar

NH2

R

O

Ar

NHR

ONHFmoc

R'

Piperidine

deprotection O

Ar

NHR

ONH2

R'

AcOH

Cyclisation

R

N

N

Ar

R'

OH

R

N

N

Ar

R'

OR"

Drawback:Final product must contain X= OH or CO2H

7. Combinatorial synthesis7. Combinatorial synthesisHeterocyclic synthesis - improved synthesis of benzodiazepinesHeterocyclic synthesis - improved synthesis of benzodiazepines

• Functional group released from the resin takes part in the final Functional group released from the resin takes part in the final cyclisationcyclisation

• Does not remain as an extra, possibly redundant groupDoes not remain as an extra, possibly redundant group

NH

R3

R4

R2

NH

Imines

+

O

OR1

NH2

Amino acid

Cl(CH2)2Cl

O

OR1

N

R2

NHR4

R3

Adducts

TFA

N

N

R2

R1

OR4

R3

8. Planning a Combinatorial Synthesis8. Planning a Combinatorial Synthesis8.1 Aims8.1 Aims

• To generate a large number of compoundsTo generate a large number of compounds

• To generate a diverse range of compoundsTo generate a diverse range of compounds

• Increase chances of finding a lead compound to fit a binding Increase chances of finding a lead compound to fit a binding sitesite

• Synthesis based on producing a molecular core or scaffold with Synthesis based on producing a molecular core or scaffold with functionality attachedfunctionality attached

Centroid or scaffold

Substituent'arms' Binding groups

8. Planning a Combinatorial Syntheses8. Planning a Combinatorial Syntheses8.1 Aims8.1 Aims

Target molecules should obey Lipinski’s ‘Rule of Five’ for oral Target molecules should obey Lipinski’s ‘Rule of Five’ for oral activityactivity

• a molecular weight less than 500a molecular weight less than 500• a calculated log a calculated log PP value less than +5 value less than +5• no more than 5 H-bond donating groupsno more than 5 H-bond donating groups• no more than 10 H-bond accepting groupsno more than 10 H-bond accepting groups

8.2 Scaffolds8.2 Scaffolds

Molecular weight of scaffold should be low to allow variation of Molecular weight of scaffold should be low to allow variation of functionality, without getting products with a MWt > 500functionality, without getting products with a MWt > 500

8. Planning a Combinatorial Syntheses8. Planning a Combinatorial Syntheses

• ‘‘Spider’ scaffolds preferable for exploring conformational Spider’ scaffolds preferable for exploring conformational spacespace

• Allows variation of functional groups around whole molecule Allows variation of functional groups around whole molecule to increase chances of finding suitable binding interactionsto increase chances of finding suitable binding interactions

RECEPTORBINDINGSITE

Binding regions

Screen compoundlibrary

8.2 Scaffolds8.2 ScaffoldsTadpole scaffoldsTadpole scaffolds

- variation restricted to a specific region round the molecule- variation restricted to a specific region round the molecule- less chance of favourable interactions with a binding site- less chance of favourable interactions with a binding site

Privileged scaffolds Privileged scaffolds - scaffolds which are common in medicinal chemistry and - scaffolds which are common in medicinal chemistry and which are associated with a diverse range of activitieswhich are associated with a diverse range of activities- benzodiazepines, hydantoins, benzenesulphonamide etc - benzodiazepines, hydantoins, benzenesulphonamide etc


'Spider' Scaffold with 'dispersed' substituents

'Tadpole' scaffold with 'restricted' substituents

8.2 Scaffolds - examples8.2 Scaffolds - examples

BenzodiazepinesBenzodiazepines

R

X

N

N

Ar

R'

O

R"

HydantoinsHydantoins

N

N

O

O

R4

R3R1 R2

-Lactams-Lactams

N

O R5

R4R3R1

R2

PyridinesPyridines

N

R

HO2C

Me

R2

O

R3

• Good scaffoldsGood scaffolds• Spider likeSpider like• Low molecular weightLow molecular weight• Variety of synthetic routes availableVariety of synthetic routes available

DipeptidesDipeptides

O

R2

C

O

CN

N

O

R4

R1

R3

R5

R6


8.2 Scaffolds - poor examples8.2 Scaffolds - poor examples

O

OR2

OR1

OR5

R4O

R3O

GlucoseGlucose

Spider like and small molecular weight - good pointsSpider like and small molecular weight - good pointsBut multiple OH groupsBut multiple OH groupsDifficult to vary RDifficult to vary R11-R-R55 independently independently

R1CO

Me

Me

R2

SteroidSteroid

M.Wt. relatively highM.Wt. relatively highRestricts no. of functional groups to keep MWt.< 500Restricts no. of functional groups to keep MWt.< 500Relatively few positions where substituents easily Relatively few positions where substituents easily addedadded

H2N

O

N

R3

R2

R

IndoleIndole

Tadpole like scaffoldTadpole like scaffoldRestricted region of Restricted region of variabilityvariability


Example - Ligands for carbonic anhydraseExample - Ligands for carbonic anhydrase9. Dynamic combinatorial chemistry9. Dynamic combinatorial chemistry

• Reaction - reversible formation of iminesReaction - reversible formation of imines

C OH

R+ H2N R'

Aldehyde Primary amine

C NH

R

R'

Imine

• Reaction carried out in presence of carbonic anhydraseReaction carried out in presence of carbonic anhydrase• Three aldehydes and four amines present as building blocksThree aldehydes and four amines present as building blocks• Sodium cyanoborohydride added to ‘freeze’ the mixture Sodium cyanoborohydride added to ‘freeze’ the mixture

C NH

R

R'Imine

NaCNBH3HC NHH

R

R'Secondary amine

• Products quantified and identifiedProducts quantified and identified• Experiment repeated in absence of target to identify amplified Experiment repeated in absence of target to identify amplified

product(s)product(s)• Amplified product is not necessarily present in greatest Amplified product is not necessarily present in greatest

amountsamounts

Example - Ligands for carbonic anhydraseExample - Ligands for carbonic anhydrase9. Dynamic combinatorial chemistry9. Dynamic combinatorial chemistry

• Building blocks Building blocks

OCHOHO3S

CHO

HO2C

CHOS

Aldehydes

O

O

H2N

H2NNH2

O

H2NNH

O

HO2CH2N

H2NNH

OO

Amines

• Amplified productAmplified product

SNO

O

H2N

Active compound

NaBH3CN

SHNO

O

H2N

Derived secondary amine

Documents

DEFINITION The automated synthesis of a large number of compounds in a short time period using a defined reaction route and a large variety of reactantsThe