Medicinal Chemistry: Combinatorial Chemistry-Parallel Synthesis 1 ...

Medicinal Chemistry: Combinatorial Chemistry-Parallel Synthesis Agenda

1. Introduction: The Drug Discovery and Development Process 2. Lead Discovery and Lead Optimization-Drugability -Drugability: Lipinski’s rule of 5 -Drugability parameters -Shape analysis -Is there a difference between leads and drugs? the rule of 4 -Fragments: the rule of 3 -Privileged structural elements -Bioisosteres -Unwanted molecular properties

3. Combinatorial and Parallel Synthesis in Medicinal Chemistry -Historical background-objective -The role of combinatorial chemistry and parallel synthesis in drug discovery -Compound mixtures versus single compounds -Solid phase synthesis versus synthesis in solution -Parallel versus split-mixed synthesis

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Daniel Obrecht, Polyphor Ltd

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4. Combinatorial synthesis of Biopolymers -Linear, modular synthesis of biopolymers -Solid-phase synthesis of polypeptides; peptoids; oligosaccharides -Parallel synthesis vs combinatorial synthesis: split-mixed synthesis -Examples for solid-phase synthesis: Split-mixed synthesis; tagging strategies; pin synthesis; tea-bags; photolithography; radiofrequency tags; binary encoding; factor Xa inhibitors; thrombin inhibitors; inhibitors of protein-protein interactions; hot spots and o-rings; synthesis of a-helix mimetics; phage libraries

-Peptide mimetics

5.

Strategies for the Synthesis of Small Molecule Libraries -Library synthesis planning -Synthesis strategies -Classical multi-component reactions (MCR’s) -Sequential multi-component reactions (SMCR’s) -Diversity-oriented synthesis (DOS) -Collective synthesis of natural products -Fragment-based lead discovery -Dynamic Combinatorial Synthesis; -Target-guided synthesis (TGS) -Disulfide thethering; click chemistry

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5. Strategies for the Synthesis of Small Molecule Libraries (cont.) -Most important reactions used in parallel and combinatorial synthesis -Most important building blocks used in parallel and combinatorial synthesis -Parallel and/or combinatorial synthesis -Parallel work-up

6. Applications of Parallel Synthesis and Combinatorial Chemistry in Medicinal Chemistry -Case studies -Drug targets

7. Appendix (Definitions; Reviews; Literature)

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Medicinal Chemistry: Combinatorial Chemistry-Parallel Synthesis 1. Introduction: The Drug Discovery and Development Process The value added chain of pharmaceutical R & D

Drug Discovery

Drug Development

NCE

Market/Revenues

Generics

since 1980: 9-13 years patent life span: 20 years -From 1960 to 1980 the development time of a new NCE (new chemical entity) have quadrupled -Since 1980 9-13 years have been necessary for the development of a new drug -Costs have gone up from 300-450 MioSFr (1987) to 600-800MioSFr in 2000 -Main reasons: higher degree of scientific knowledge of the drug required; regulations for clinical quality assurance; change in the professional regulations of physicians; increase for administrative work for health authorities -It is of prime importance to reduce the development time and costs for the development of a new NCE in order to reduce the costs of new drugs while keeping the profitability: increase productivity of R & D J. Kuhlmann, Int. J. Clinical Pharmacol. Ther. 1997, 35, 541-552 Winter Semester 12


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Medicinal Chemistry: Combinatorial Chemistry-Parallel Synthesis 1. Introduction: The Drug Discovery and Development Process The Drug Discovery Process

Drug Discovery Process Assay development capabilities

Target identification

Screening libraries

Establishing primary screening

Screening capabilities

Parallel chemistry

Hit identification

PEM, small molecules, fragments

Medicinal chemistry

Hit exploration, hit-to-lead

Molecular modeling

Lead optimization

Preclinical and clinical development

ADMET properties

Genomics; Proteomics; Phage display, Fragment screening; X-ray crystallography

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Medicinal Chemistry: Combinatorial Chemistry-Parallel Synthesis 1. Introduction: The Drug Discovery and Development Process The Drug Discovery Process

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Medicinal Chemistry: Combinatorial Chemistry-Parallel Synthesis 1. Introduction: The Drug Discovery and Development Process Attrition rates in the discovery and preclinical phases

Lead Optimisation

5-12 m

Clinical Candidate Selection (CCS) from alternatives (Candidate Profiling)

12-36 months

3-6 m

A ttr itio n in D is c o v e r y

>30% 30%

1 2 .0 0 1 0 .0 0 8 .0 0

>50% 30%

6 .0 0 4 .0 0

Of 10 projects starting in Lead Identification 5 -In addition additional parameters determining favorable oral bioavailability are [2]: -not more than 5 (10) fully rotatable bonds -polar surface area 30 Ki = 85nM (strongest binder)

M. Hochgürtel et al. J. Med. Chem. 2003, 46, 356-8

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: Dynamic Combinatorial Synthesis: disulfide thethering SH

+ IL-2

disulfide exchange

R1 S S R2

S S

R

binding stabilizes disulfide

S S R

IL-2

IL-2

C S

best R series:

S

O

N

A, B, C: H, CO2H, CO2Me or MeO

B A

improve design of a known inhibitor with tethering "hit"

N

Me N

N

R

Cl Cl

O C

A

Me N

R

Cl Cl

B improved inhibitors IC50 = 0.2 mM

existing inhibitor IC50 = 3 mM

J. A. Wells et al. Proc. Natl. Acad. Sci. USA 2000, 97, 9367-72; A. C. Brainsted et al. J. Am. Chem. Soc. 2003, 125, 3714-15 Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: Click chemistry

Click Chemistry: Diverse chemical function from a few good reactions H. C. Kolb, K. B. Sharpless, Angew. Chem. Int. Ed. 2001, 40, 2004

Development of a set of powerful reactions for the rapid synthesis of useful new compounds and combinatorial libraries through heteroatom links (C-X-C); an approach called Click Chemistry. Reactions that have a high thermodynamic driving force, usually greater than 20 kcal/mol -Cycloadditions ([1,3]-dipolar additions; Diels-Alder reactions) -Nucleophilic Substitution reactions on strained heterocyclic electrophiles -Carbonyl Chemistry of the non-Aldol-type: synthesis of ureas, thioureas, aromatic heterocycles, oxime ethers -Addition reactions to C-C carbon multiple bonds: epoxidations, aziridinations, dihydroxylations

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: Click chemistry Nature X

+

R1-N=N=N-

n

:Nuc

Petroleum

O 1

R

R2 2

R

N

X: O, NR

N N

XH Nuc R1

R3XNH2 R1

O N

N N R3

N

N

H R

4

R1

XR3 R2

H. C. Kolb, K. B. Sharpless, Drug Discovery Today 2003, 8, 1128-37

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: Click chemistry

[1,3]-Dipolar additions of acetylenes and azides

-

OH

+

N N N

+

N N N-

Cu (turnings) (ca 1g)

HO (10.0mMol)

+ Ph

H2O/tBuO(2:1) (50ml) RT, 24h CuSO4(cat.) (10Mol%)

Ph

N

N N

OH N N HO

N

Ph

3.7g (95%) white solid

(20.0mMol) V. V. Rostovtsev et al. Angew. Chem. Int. Ed. 2002, 41, 2596

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: application of Click chemistry O O

O O

O-

O

N O

O

N

O-

HO

N H N

NH2

+ O R

N=N=Nn

O

N N N

N H

4

R

O O

O-

O-

N n

O

O HO

N N N

O O

O O

O-

N

O

N O

O

N

O-

HO

N H N

NH2

OH

O

N

O O

O

H

H2O/tBuO(2:1) (50ml) RT, 24h CuSO4(cat.) (10Mol%)

+

N H

Cu (turnings) (ca 1g)

OH

N H N

NH2

Ki: 62nM; inhibition of fucosyl transferase cancer metastasis; lymphocyte trafficking

OH

Lee et al. J. Am. Chem. Soc. 2003, 125, 9588-89

Dramatic rate acceleration of the azide-alkyne cycloaddition by sequestering the two components inside the host structure (enzyme or receptor) Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: application of Click chemistry

-Emerging resistance in clinical isolates of bacteria render existing antibiotics such as Neomycin and Ciprofloxaxin inactive -Enzymes such as aminoglycoside 3‘-phsphotransferases inactivate 3‘ position in aminoglycoside antibiotics by phosphorylation -Combination of two antibiotics has emerged as a valuable strategy to overcome rapid resistance mechanisms

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: application of Click chemistry

-biological activities (MICs) depended significantly on the variable spacer groups X and Y -best combinations were X= -(CH2)2and Y= -CH2OCH2-MIC (minimal inhibitory concentration): E.coli (R477-100): 3mg/ml E.coli (ATCC 25922): 3mg/ml E.coli (AG100A): 0.38mg/ml B. subtilis (ATCC 6633): 0.75mg/ml

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: application of Click chemistry Azide 1

Alkynes

O

O

OMe O

x O S N O

O N3 OH

N H

O

OH

COOMe N

N H

OH

O N H

COOMe

OMe N

OMe

HIV-protease (SF-2), buffer, 23°, 24h OMe

O S N O

O OH

N N N

Ki = 1.7 nM

O HN HO

M. Whiting et al. Angew. Chem. Int. Ed. 2006, 45, 1435-39; K. B. Sharpless, R. Manetsch, Exp. Opin.Drug. Disc. 2006, 1(6), 525-38

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Synthesis strategies: application of Click chemistry Summary of fragment-based approaches: -fragment libraries are smaller: few hundreds to thousands -screening effort smaller; however, weak binders have to be detectable -leads derived from fragments are often smaller; allows more extensive optimization -fragments can be assembled in a thermodynamically or kinetically controlled fashion: dynamic combinatorial synthesis -fragments can be assembled using click chemistry -finding the appropriate linkers to assemble fragments is a big challenge

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 2. Lead Discovery and Lead Optimization-Drugability Most important building blocks (toolbox) used in parallel and combinatorial synthesis Systematic enumeration of of key heteroaromatic reagent classes from commercially available sources which have been used in medicinal chemistry programs

R. Ward et al. J. Med. . Chem. 2011, 54, 4670-4677; S. D. Roughley et al. J. Med. Chem. 2011, 54, 3451-3479 Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Most important reactions used in parallel and combinatorial synthesis

Couplage Suzuki:

Formation dàmides et dùrées: O

O OH

BB

NH2 BB

HV

NH(R)R

BB

H N BB

H N

Ar

X

BB

BB

Réduction au diborane: RHV

O

O BB

Amination réductrice:

N H

RHV

NHRHV

BB

Alkylation du groupe thiol:

O BB

H

BB

NH(R)RHV

R1-SH

+

R2

Br O

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base

R2

R1S O

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Chemical Biology: Combinatorial Chemistry-Parallel Synthesis 2.5. Parallel reactions Most important reactions used in parallel and combinatorial synthesis Substitution nucleophillique:

BB

Alkylation de NH activés:

N

N

BB

N

2

N H

Réaction de Mitsunobu:

N O HV R

O

Réaction de Mannich:

R2

2

R1

OH

R

1

2

R

R

2

R CHO

3

O

R

R1 N

O

R

OH

R

1

2

N R1

R

N H

3

N H

N R2

R

N

3

NH R

O 1

1

R

R2

NH(R)RHV

N

X

R

R1

O

R1

NH2 R2

R1

OH

R1

R2

R1

N3 R2

OH R2

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R1

O R

2

R3


183

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Questions

1. Please name five efficient reactions that can be used for final parallel derivatization? 2. Please name potential advantages of fragment-based lead discovery over screening large combinatorial libaries? 3. What is the rule of 3?

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up procedures

Extractions : principle Liquid-liquid extractions Solid-phase extractions

Solid-supported scavengers Ion-exchange resins Fluorous phase extractions

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up procedures: principle

1. Two phase extractions: manuel extraction

Upper phase: contains product (EtOAc or fluorous phase): separated manually Lower phase: contains impurities (aqueous phase)

2. Two phase extractions: robotic system (style Tecan)

Upper phase: contains impurities (aqueous phase): separated by robot Lower phase: contains product (CHCl3 or CH2Cl2): dried and evaporated

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up strategies: liquid-liquid extractions

1. Two phase extractions: solubilize impurities in the aqueous phase O

S

+

NHR1

H2N

Br

1. MeOH, 60° R3

R

2

2

1

Me2N(CH)nNH2

H N

NH2 S

HOOC

R3

R2

N 2. 4, 60°

R1HN

3. aq.NaOH CH2Cl2

3

H N

excess 2

S R2

N

HOOC

4

S

5

R3

Products of type 5 are soluble in the basic aqueous phase A. Chuchulowski, T. Masquelin, D. Obrecht, J. Stadlwieser, J.-M. Villalgordo, Chimia 1996, 50, 530

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up strategies: solid-phase extractions

Solid phase extractions/filtrations

Solid phase: one or several solid pahses are filled into a polypropylene syringe or cartridge

Solid phases: SiO2; Al2O3; `ìon exchange resins (basic, acidic and mixed bed)``; Kieselguhr; MgSO4; polymère functionalisé: -NH2, -SH, -PPh2, COOH, CHO, CH2OH, isothiourée, N3...; The organic phases are passed through these cartridges in order to get rid of impurities which are adsorbed onto the solid phase. They can be applied manually or by a robotic system (Tecan)

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up strategies: solid-supported scavengers

R2-N=C=O

R1-NHCONHR2 2

3

R -COCl

R1-NH2

R4-SO2Cl

1

R1-NHCOR3 3 1

R -NHSO2R4 4

NH2

excess

NHCONHR2 NHCOR3

NHSO2R4

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up strategies: solid-supported scavengers Parallel synthesis in solution using polymer-bound reagents

P1

R CbzHN

i O 1)

O

R

COOH

P2 GP

i

N H

R'

P1'

O R

cysteine trap

Cl R CbzHN

N

N2

H N+

O

O DMF, -5°

(5-10% methylester)

CbzHN

Br O

N

or R

1 2

R R NH CbzHN or R3SH DCM, 18h

DMF, 1.5h, 25°

-10°, 1h 85-90%

O

Br-

2) CH2N2/DCM

NR1R2

CbzHN

R

SR3 O

80-85% HN

HN N

CbzHN O

N H

Ph

CbzHN

S

S

O

N. Y. Yadav-Bhatnagar, N. Desjonquères, J. Mauger, J. Comb. Chem. 2002, 4, 49-55 Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up strategies: solid-supported scavengers; intermediate catch Parallel synthesis in solution using polymer-bound reagents

COOH R

("intermediate catch" or "resin capture"

3

N3

+

O 1

R NH2

R3

O

O

3

NHR5

R

5

NHR

-

CHO

R1 N

O

O

N O +N N

+

R2

DCM, 0°

R1 N

N O P Ph Ph

PPh3

wash

R2

R2

toluene, 60°

+ 4

O

R N=C

N

R3

R1

O NHR5

N

R2 A. Chucholowski, D. Heinrich, B. Mathis, C. Müller, Generation of benzodiazepin and benzodiazocin libraries through resin capture of Ugi-4CC, conference: 214th ACS national meeting, Las Vegas, 1997

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up strategies: fluorous phases

FP: fluorous phase; C6F13CH2CH2- or C10F21CH2CH2-

FP

+

FP

liquid phase reactions

Substrate

FP + excess reagents Products

Substrate

liquid-liquid extraction

1. cleavage Products 2. extraction

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FP Products

192

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up strategies: fluorous phases

Multicomponent reactions: fluorous phase extraction

R1NH2 COOH (Rf)3Si

+

i, ii R2CHO (Rf)3Si

N R1

R2

O

R2

O

NHR3

iii, ii

O

N R1

NHR3 O

R3N=C Rf: C10F21CH2CH2i: TFE, 90°, 48h; ii: liquid-liquid extraction; iii: Bu4NF, THF, rt

A. Studer, S. Hadida, R. Ferritto, S.-Y. Kim, P. Jeger, P. Wipf, D. Curran, Science 1997, 275, 823

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 5. Strategies for the Synthesis of Small Molecule Libraries Parallel work-up strategies: fluorous phases Fluorous phase extraction: cleavage by cyclization X O O

N

O N

N

+

O

N Me

OTf-

2

1

ii

N H

(Rf)3Si

(Rf)3Si

(Rf)3Si

O

O

i

COOH

3

Rf: C6F13 CH2CH2-

iii

X O

O O

iv

N H

(Rf)3Si

O 4

X

NH

O

N N H

O

O

i: MeOTf, CH2Cl2, 1,1,1-(trifluoromethyl)benzene(BTF); ii: anthranilic acid, DMAP, BTF, CH2Cl2; iii: TBTU, furfuryl amine, THF;iv: Et3 N D. Schwinn, W. Bannwarth, Helv. Chim. Acta 2002, 85, 255

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies What are the prime biological targets?

-Kinases: 22%; market: 2 drugs -GPCR: 15%; ¨ : 30% -Ion channels: 5%; ¨ : 7% -Ser proteases: 4%; ¨ : 1 drug -Phosphatases: 4%; -Zn proteases: 2%; ¨ : ACE inhibitors -Nuclear receptors: 2%; ¨ : 4% -others* : 44%; *Many targets involving large surface protein-protein interactions -despite the fact that kinases, CPCR‘s and ion channels constitute only about 42% of all targets of therapeutic interest, the pharmaceutical industry is devoting about 90% of their resources to those targets; it is believed that these targets can be adressed with small molecules. -The number of biologicals (antibodies, fusion proteins, peptides) reaching the market is increasing. These molecules target mainly large surface protein-protein interactions Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Targets hit by current drugs Drugs, their targets and the nature and number of drug targets P. Imming et al. Nature Rev. Drug Disc. 2006, 5, 821-34 1. Number of drug targets : 1997 : Drews et al. Nature Biotechnol. 1997, 15, 1318-19 -Marketed drugs hit 482 targets ; human genome suggests 100'000 proteins 2002: J. Burgess et al. -after sequencing of human genome:

~8000 targets ~5000 hit by known drugs: 2400 by antibodies; 800 by proteins

2002: A. Hopkins et al. Nature Rev. Drug Disc. 2002, 1, 727 -on the basis of ligand binding studies: 399 targets, which belong to 130 target families ~3000 targets amenable to small molecules bottom line: 300-500 targets hit by current drugs; 3’000-8’000 drugable targets Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Kinase inhibitors -Recent reviews: A. J. Bridges, Chem. Rev. 2001, 101, 2541-2571; G. Scapin, Drug Disc.Today 2002, 77, 601-611; S. Orchard, Curr. Opin. Drug Disc. & Dev. 2002, 5, 713-717; D. Fabbro, C. Garcia-Echeverria, Curr. Opin. Drug Disc. & Dev. 2002, 5, 701-712; S. K. Hanks, The FASEB J. 1995, 9, 576-596 (sequences of kinases); M. E. M. Noble, J. A. Endicott, L. N. Johnson Science 2004, 303, 1800-5; J. Zhang; P. L. Yang; N. S. Gray, Nat. Rev. Drug Discov. 2009, 9, 28-39 (Targeting cancer with small molecule kinase inhibitors); -Three families of kinases:

-Serine-threonine kinases (S/TKs) -Tyrosine kinases (TKs) -Dual function kinases (DFKs)

-Roughly 2000 kinases known in the human genome -Kinases phosphorylate serine, threonine O and tyrosine and are ATP dependent OH TKs)

P O O O-

R

*

OH

ATP

ATP

phospatases Winter Semester 12

TKs)

R

O-

* O P OO

phospatases Daniel Obrecht, Polyphor Ltd

197

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Kinase inhibitors on the market

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies GPCR’s: introduction 50% of all drugs target G-Protein-Coupled Receptors (sales in 2001: ~50billion USD) G-protein: guanin nucleotide-binding protein -240 receptors with known ligands from which only ~30 are currently investigated by pharma companies -An additional 160 receptors with unknown ligands (orphan receptors) are known Family 1: rhodopsin-like or adrenergic-like GPCR‘s constitute the largest family; contain a short N-terminus and amino acid residues in the trans-membrane domain are highly conserved Family 2: glucagon receptor-like or secretin receptor-like GPCR‘s Family 3: metabotropic glutamate receptors Drug design strategies for targeting G-protein-coupled-receptors: Th. Klabunde, G. Hessler, ChemBioChem 2002,3, 928-44. 3D-structure of bovine rhodopsin: Science, 2000, 289, 739-45; Biochemistry, 2001, 40, 7761-72. Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies GPCR’s: introduction G-proteinprotein-coupled receptors

Extracellular

-NH2

-S-S-

e1

TM1

TM2

TM3

TM4

e3

e2

TM5

TM6

TM7

i2 i1 i3

Cytoplasmic

COOH-

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200

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies GPCR’s: some best-selling drugs H N

S Cl

N

N

H2N

N N O

COOH

N

OEt

Claritin (Schering-Plough, H1 antagonist allergies, 3.1billion USD, 2001)

N N N HN

HO HO OH

N H

N

COOH

O

Serevent (Glaxo, b1 agonist asthma, 0.91 billion USD, 2001)

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Neurontin (Pfizer, GABA B-agonist neurogenic pain, 2.35 billion USD, 2001)

Zyprexa (Ely Lilly, D2/D1/5-HT2 allergies, 2.35 billion USD, 2001)

Diovan (Novartis, AT1 antaginist hypertension, 0.8 billion USD, 2001)


201

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Extracelluular ligand gated channels

Nicotinoid AChR GABA Glycine 5-HT3

AChR: Neuromuscular Disorder 300000 patients US Hydroxytryptamin type

Migraine, depression Not considered here

14 subtypes Voltage gated channels

Extracellular ligand gated channels Winter Semester 12

Na+ Ca ++ K+

>15 subtypes 35 subtypes > 100 subtypes 50% not yet charactarized

NMDA AMPA KAINATE

Na: Migraine, back pain, 34 mio. patients US Ca: Hypertension patients US, prostate cancer K: MS, spinal cord injury 250000 patients US NMDA: Brain ischimia, CNStrauma, epilepsy, huntington disease 2.3 million patients


202

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 1: Inhibitors of influenza endonuclease

Inhibitors of influenza endonuclease: collaboration between Roche and Polyphor Ltd

R1

R1 F

O BocHN

Winter Semester 12

NO2

OH

OPiv

BocHN

N OPiv


O RHN

N OH

203


Disease and target

- Influenza infects an estimated 120 Mio. people in US, Europe and Japan in a typical year -The influenza endonuclease is an attractive target for several reasons:

i: It is a key component of the viral transcription mechanism, which has no cellular counterparts and should therefore provide a good potential for discovering selective, non-toxic drugs ii: In contrast the neuraminidase inhibitors that do not prevent the formation of new virus particles, but interfere with virus release from host cells and are therefore virustatic, endonuclease inhibitiors, due to the block of viral transcription, are expected to have a virucidal effect.

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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 1: Inhibitors of influenza endonuclease N-Hydroxy-imides

Ketobutanoates O

OH

N

OH

N O

Merck: IC50: 21.3mM

N O OH

Flutimide: fungal methabolite

OH OMe

OH

O Roche: IC50 95mM

Merck: IC50: 5.5mM F

O

O N

Merck: IC50: >500mM O

O N

N

O

O

H N

O N

OH OH O

Roche: IC50 5mM

O

OH

Roche: IC50 >1000mM

N O OH

O

Merck: IC50: 0.9mM

N

OH

O O

OH

H N

OH

N Ph

O

Roche: IC50 15mM OH

O

O

Roche: IC50 >500mM

NH2

O

Cl

Roche: IC50 800mM

Roche: IC50: 0.43mM

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N


205

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 1: Inhibitors of influenza endonuclease N-Hydroxy-imides

Ketobutanoates O

OH OH O

N O

Merck: IC50: 21.3mM

N OH

O

Flutimide: fungal methabolite Merck: IC50: 5.5mM

N-Hydroxy-tetramic acids R1

R1 OH

R2

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N OH

O

O R2


OH N OH

206

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 1: Inhibitors of influenza endonuclease N-Hydroxy-2-indolinones R1

R1 OH

R2

N OH

O R2

O

OH N OH

R1 OH O N OH

RHN 1

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-N-hydroxy-2-indolinone derivatives 1 were not described in the literature R1 OH

b-keto amide moiety

O high variation site

N OH

RHN

hydroxamic acid moiety

1

-Molecules of type 1 bear two potentially reactive and labile functional groups for which suitable protective groups have to be found -Molecules of type 1 are acidic and polar and thus problems of isolation and purification were anticipated; especially for a parallel approach

Winter Semester 12


208

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 1: Inhibitors of influenza endonuclease R1 OH O R3

O N OH

N H 1A

R1

R1

OH

OH O N OH

RHN

R1

O R4HN

O N OH

N H

H2N

OCOR2

O N OPG

2

1B

1

key precursors for parallel synthesis

R1 OH O O 5 S R N H

PG: protective group

O N OH 1C

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209


R1

O

O H2N

COOH

OCOR2

N OPG

N OPiv

BocHN 2

3

BocHN

NO2 4

key precursors for parallel synthesis

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210

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies

Case study 1: Inhibitors of influenza endonuclease

F H2N

NO2

F

i BocHN

NO2 7

6

CH(COOMe)2

ii BocHN

NO2 5

i: Boc2O, THF, 80°; ii: CH2(COOMe)2, NaH, DMSO; iii: aq. NaOH, MeOH, reflux

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211


problemes: -partial reduction of nitro group -cyclization -isolation of hydroxamic acid

COOH iv, v BocHN

NO2

O BocHN

4

3

N OCOtBu

iv: Pt/C(5%),H2, DMSO, EtOH; then AcOH; v: tBuCOCl, DIPEA, CH2Cl2 Winter Semester 12


212


R1

R1COX (2.5 equiv.) O N OPiv

BocHN

3

i

O

O N OPiv

O

R

R1

O

BocHN

1

O ii BocHN

O N OPiv

9a (R1= Me)

10a (R1= Me)

9b (R1= Et)

10b (R1= Et)

9c (R1= Ph) i: 9a: X=OCOCH3; 9b: X=Cl; 9c: X=Cl, DMAP, DIPEA, CH2Cl2, THF; 0°-r.t.; ii: tBuCOCl, tetrabutylammonium cyanide or NaCN, pyridine, CH2Cl2

Winter Semester 12


213

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 1: Inhibitors of influenza endonuclease O

O

Me

Me

O

O

BocHN 10a

i

O N OPiv

H2N 2a

O

O

O

O i

O BocHN

O N OPiv

N OPiv 10b

H2N 2b

O

O N OPiv

O Ph

O

Ph O

i

O BocHN 9c

N OPiv

H2N

O N 2c OPiv

i: 4N HCl/dioxane or CF3COOH/CH2Cl2; then extraction with aq. sat. NaHCO3 solution and CH2Cl2

Winter Semester 12


214

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 1: Inhibitors of influenza endonuclease R1 O

i 3

R

N H

11a

O N OPiv

R1 R1

OCOR2

O ii

H2N

4

R HN

O N OPiv

N H

11b

2a (R1= Me; R2= tBu) 2b (R1= Et; R2= tBu) 2c (R1= R2= Ph)

O R

5

S

O N H

OCOR2

O N OPiv

R1 iii

OCOR2

OCOR2

O N 11c OPiv

i: R3COCl, pyridine (DIEA), DMAP, CH2Cl2; ii: R4N=C=O, CHCl3 (ethanol-free), 70°; iii: R5SO2OCl, pyridine (DIEA), DMAP, CH2Cl2 Winter Semester 12


215

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 1: Inhibitors of influenza endonuclease - out of the library of 131 compounds 26 had an IC50< 50mM

Me

Ph

OH O

OH

O N H

S

N OH

O

S

O N H

N OH

O

IC50= 48mM

IC50= 9mM; EC50= 21mM Ph OH O HOOC

N H

O N H

N OH

IC50 = 3mM; EC50= 6mM

compounds are antiviral in cell cultures Winter Semester 12


216

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies

Case study 1: Inhibitors of influenza endonuclease

-Based on a pharmacophor hypothesis, novel 1-hydroxy-indolin-2-ones were proposed as inhibitors of influenza endonuclease -A parallel synthesis was developed which allowed to synthesize a library 131 compounds in significant quantities (6-71 mg) and high purities (75 - 99%) within 4 months

-From 131 compounds tested 26 had an IC50< 50mM

-From 26 active compounds 10 showed a good antiviral activity in cell cultures

Winter Semester 12


217

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 2: Structure-based Design Inhibitors of Metalloproteinases: TACE; MMP1

-Ligand-based design capitalizes on the presence of existing structural similarities between a set of compounds and the active ligand (also pharmacophore-based drug design) -Using a solid-phase, parallel synthesis approach optimization could be achieved efficiently -TNF-a converting enzyme (TACE) represents an attractive target for reducing circulating levels of the proinflammatory protein tumor necrosis factor alpha (TNF-a). -TACE belongs to the Zn-metalloproteinases. The hydroxamic acid moiety chelates to the Znatom located in the active site Winter Semester 12


218

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 2: Structure-based Design Inhibitors of Metalloproteinases: TACE; MMP1

M. Abou-Gharbia, J. Med. Chem. 2009, 52, 2-9 Winter Semester 12


219

Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 3: Parallel synthesis of analogues of antibiotic GE2270 A Parallel synthesis starting from a natural product-derived building block O

O N S

CONH2

N

OH O

N

S

N

N N N

N

S

S

N

S HN

O

O HN

MeHN

N O

S

NH

NH

N S MeO

N

OH O

O

N

S N

S

S

HN

O

O HN

MeHN

N O

S

OH O NH

NH

N S

O

MeO

GE2270 A

1 inhibitor of elongation factor EF-TU

active against many gram positive pathogens MIC 0.06-1.0 mg/ml; low solubility in aqueous solvents J. W. Jacobs et al. (Versicor), 40th annual ICAAC conference, Toronto, Canada, september 17-20th, 2000, Poster 2193 and 2194

Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 3: Parallel synthesis of analogues of antibiotic GE2270 A Parallel synthesis starting from a natural product-derived building block NO2

F OH

O

O S

S

N N N

HN

O

O HN

MeHN

O

S

S 2

OH O

N R

4 NO2

N S MeO

O

O

O

NH

NH

N

F

NO2

N

S

S

O

O

N R

S

O

F O

N

F

F

O S

O S

N R

3

O

N H N R

5

1

J. W. Jacobs et al. (Versicor), 40th annual ICAAC conference, Toronto, Canada, september 17-20th, 2000, Poster 2193 and 2194 Winter Semester 12


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Medicinal Chemistry : Combinatorial Chemistry-Parallel Synthesis 6. Case studies Case study 2: Parallel synthesis of analogues of antibiotic GE2270 A Parallel synthesis starting from a natural product-derived building block

F O

F

S

O

F O

F

R R NH

S

N

2

R =H R1:

6

R

2

R1:

R2

N R1

1 2

N R

O

O O S R

O

R1R2NH S

N 3

COOH

0.5

COOH

0.91

OH

R2=Me

NO2

O

2.0

0.5

O

S

MeO

12