An Introduction to Medicinal Chemistry 3/e COMBINATORIAL CHEMISTRY Patrick An Introduction to Medicinal Chemistry 3/e Chapter 14 COMBINATORIAL CHEMISTRY Part 1: Sections 14.1 – 14.4

Contents Part 1: Sections 14.1 – 14.4 1. Definition 2. Solid Phase Techniques 2.1. Advantages 2.2. Requirements 2.3. Examples of Solid Supports (2 slides) 2.4. Anchor or linker 2.4.1. Merrifield resin for peptide synthesis (chloromethyl group) 2.4.2. Wang resin (2 slides) 2.4.3. Rink resin (2 slides) 2.4.4. Dihydropyran resin (2 slides) 3. Parallel Synthesis 3.1. Houghton’s Tea Bag Procedure 3.2. Automated parallel synthesis (2 slides) 3.3. Automated parallel synthesis of all 27 tripeptides from 3 amino acids (2 slides) 4. Mixed Combinatorial Synthesis (21 slides) [41 slides]

1. 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 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

2. SOLID PHASE TECHNIQUES Reactants are bound to a polymeric surface and modified whilst still attached. Final product is released at the end of the synthesis 2.1 Advantages Specific reactants can be bound to specific beads Beads can be mixed and reacted in the same reaction vessel Products formed are distinctive for each bead and physically distinct Excess reagents can be used to drive reactions to completion Excess reagents and by products are easily removed Reaction intermediates are attached to bead and do not need to be isolated and purified Individual beads can be separated to isolate individual products Polymeric support can be regenerated and re-used after cleaving the product Automation is possible

2. SOLID PHASE TECHNIQUES 2.2 Requirements A resin bead or a functionalised surface to act as a solid support An anchor or linker A bond linking the substrate to the linker. The bond must be stable to the reaction conditions used in the synthesis A means of cleaving the product from the linker at the end Protecting groups for functional groups not involved in the synthesis

2. SOLID PHASE TECHNIQUES 2.3 Examples of Solid Supports Partially cross-linked polystyrene beads hydrophobic in nature causes problems in peptide synthesis due to peptide folding Sheppard’s polyamide resin - more polar Tentagel resin - similar environment to ether or THF Beads, pins and functionalised glass surfaces

2. SOLID PHASE TECHNIQUES 2.3 Beads must be able to swell in the solvent used, and remain stable Most reactions occur in the bead interior Starting material, reagents and solvent Swelling Linkers

2. SOLID PHASE TECHNIQUES 2.4 Anchor or linker A molecular moiety which is covalently attached to the solid support, and which contains a reactive functional group Allows attachment of the first reactant The link must be stable to the reaction conditions in the synthesis but easily cleaved to release the final compound Different linkers are available depending on the functional group to be attached and the desired functional group on the product Resins are named to define the linker e.g. Merrifield Wang Rink

2.4.1 Merrifield resin for peptide synthesis (chloromethyl group) Linker Peptide Release from solid support

2.4.2 Wang resin Linker Linking functional group Bead Linker

2.4.2 Wang resin piperidine deprotection Carboxylic acid Carboxylic

2.4.3 Rink resin Linking functional group Linker

2.4.3 Rink resin Carboxylic acid Primary amide cleavage

2.4.4 Dihydropyran resin Linker Linking functional group

2.4.4 Dihydropyran resin Alcohol Alcohol

3. Parallel Synthesis Aims To use a standard synthetic route to produce a range of analogues, with a different analogue in each reaction vessel, tube or well The identity of each structure is known Useful for producing a range of analogues for SAR or drug optimisation

Each tea bag contains beads and is labelled 3. Parallel Synthesis 3.1 Houghton’s Tea Bag Procedure 22 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

AUTOMATED SYNTHETIC MACHINES 3. Parallel Synthesis 3.2 Automated parallel synthesis AUTOMATED SYNTHETIC MACHINES

Use beads or pins for solid phase support 3. Parallel Synthesis 3.2 Automated parallel synthesis Wells Automated synthesisers are available with 42, 96 or 144 reaction vessels or wells Use beads or pins for solid phase support Reactions and work ups are carried out automatically Same synthetic route used for each vessel, but different reagents Different product obtained per vessel

3. Parallel Synthesis ETC 3.3 Automated parallel synthesis of all 27 tripeptides from 3 amino acids ETC

3. Parallel Synthesis 27 TRIPEPTIDES 27 VIALS 3.3 Automated parallel synthesis of all 27 tripeptides from 3 amino acids 27 TRIPEPTIDES 27 VIALS

4. Mixed Combinatorial Synthesis Aims To use a standard synthetic route to produce a large variety of different analogues where each reaction vessel or tube contains a mixture of products The identities of the structures in each vessel are not known with certainty Useful for finding a lead compound Capable of synthesising large numbers of compounds quickly Each mixture is tested for activity as the mixture Inactive mixtures are stored in combinatorial libraries Active mixtures are studied further to identify active component

4. Mixed Combinatorial Synthesis The Mix and Split Method Example - Synthesis of all possible dipeptides using 5 amino acids Standard methods would involve 25 separate syntheses Combinatorial procedure involves five separate syntheses using a mix and split strategy

Split Gly Ala Phe Val Ser

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 9 27 Tripeptides 3 Vials

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

4. Mixed Combinatorial Synthesis The Mix and Split Method 20 AMINO ACIDS HEXAPEPTIDES etc. 34 MILLION PRODUCTS (1,889,568 hexapeptides / vial)