Prof. Dr. Erick M. CarreiraLaboratorium für Organische Chemie

Eidgenössiche Technische Hochschule (ETH)Zürich, Switzerland

Organic Chemistry in the Quest for Anticancer

Therapeutics

Why we do, what we do

ESTIMATED NUMBER OF

“REALIAZABLE” MOLECULES

10120–10200

ESTIMATED NUMBER OF

KNOWN MOLECULES

0.000000000000000000000000000000000000000000000000000000000000000000000000000000000000000000000000000000000001%

108

How do we find our way?

At a rate of 10 billion molecules/year,the synthesis project would take between 10110 and 10190 years !!!

Continuing with the thought experiment...

In the search for new pharmaceuticals, how do chemists deal with such odds?

High through-put screening (Combinatorial Chemistry)

Bio-informatics (Genomics)

Biostructural Chemistry

Synthetic Chemistry

Computational Chemistry

Sets the limit to what is realizable or “makeable”

Top Twenty Drugs (Based on Sales) in 1990

RanitidineAmoxicillinAmipicillinCaptoprilEnalaprilIbuprofenNifedipineCefaclorCimetidineAtenololDiclofenacDiltiazemNaproxenCefalexinLovastatinFamotidineIohexolCefatriaxoneProxicamAlbuterol

AntiulcerAntibioticAntibioticACE-inhibitorACE-inhibitorNSAIDCalcium antagonistAntibioticAntiulcerBeta-BlockerNSAIDCalcium antagonistNSAIDAntibioticAntihypercholesterolemicAntiulcerContrast mediumAntibioticNSAIDBronchodilator

2400200018001500150014001300100010001000100010001000900750600600600600600

(in Millions USD)Therapeutic ClassDrug

Optically Pure

Optically Pure

Optically Pure

Optically Pure

Racemic mixture

Racemic mixture

Racemic mixture

Racemic mixture

When you lookinto a mirrorit is notyourself you see,but a kindof apish errorposed in fearfulsymmetry

kool uoy nehWrorrim a otniton si ti,ees uoy flesruoydnik a tubrorre hsipa folufraef ni desop.yrtemmys

Mirror by J. Updike

HSCOOH

NH2

MeMe

SHHOOC

NH2

Me Me

Penicillamin

anti-arthritisch toxisch

süss bitterH2N COOH

O NH2

NH2HOOC

ONH2Asparagin

N

O

ONH

O

O N

O

OHN

O

OThalidomid

beruhigend teratogen

The Bioactivity of Mirror-image Molecules can be Starkly Different

Die Entwicklung des Arzneimittelmarktes seit 1980

(Geschätzter Jahresumsatz 2000: 100 Mia US$, davon 43% in Europa.)

1980 1990

1994 2000

Single Enantiomer 3%

Achiral 60%

Fermentation/Extraction

20%

Racemic

17%

Single Enantiomer 9%

Achiral 50%

Fermentation/Extraction

26%

Racemic

15%

Achiral

Fermentation/Extraction

27%

Racemic

17%

Single Enantiomer

22%Racemic

17%

Single Enantiomer

34%

Achiral

21%34%

Fermentation/Extraction

28%

Source: Lehman Brothers Pharmaceutical Research and DSM Fine Chemicals

1980 1990

1994 2000

Benefits to Optically Pure Pharmaceuticals

• Dose reduction

• Simplification of dose-response relationship

• Reduction in intersubject variability

• Minimization of toxicity from enantiomer

Cancer

1. Arises from the stepwise accumulation of genetic changes which lead a cell to undergo unlimited growth that is unresponsive to homeostatic regulatory mechanisms

2. Molecular and cell biology have identified cell phenotypes that are required for malignant transformations and the specific molecular pathways responsible for faulty programming of the cell

A Subway Map of Cancer Pathways

Subway stops represent phenotypes required for malignant transformations

Molecular pathways are represented by the subway lines

Natural Products with Anticancer Activities

Antitumor:isolated from Vinca rosea Anticancer: isolated

from mandrake

Antineoplastic:Isolated from crocus

Anticancer: isolated from Pacific Yew

Natural Products with Promising Anticancer ActivitiesBreast CancerPromotes assembly of microtubules and inhibits tubulin disassembly

Sold as Paclitaxel

Ongoing investigations at Novartis, Roche, Brystol Myers Squibb, Schering

Epothilones: Promising Anticancer Leads

Epothilone-producing myxobacterium (Sorangium cellulosum)

Epothilone-disrupts microtubulin dynamics (formation and growth)-inhibit cell proliferation at mitosis

Tubulin is a heterodimeric protein which constitutes the monomeric building blocks of microtubules.

Microtubules are fundamental component of the cytoskeleton (the beams and conveyor belts of cells-the nanoweb of the cell). Microtubule dynamics is critical during cell division.

A Cell Biology Primer

Cell Biology Primer:

Natural Products bind to microtubule assemblies and interfere with the dynamics-arresting cell growth

Medicinal Chemistry Organic Synthesis

-what are the structural features of the molecule that lead to activity?

Chemical Genetics Small molecules as probes with which to understand cellular processes

-can analogs be generated

-improved activities-reduced side effects

How do we carve out the important structural features?

Me

TIPSO

OBz

OH

Me

N

SMe

Me

OHO

RTrocO

Me

O OPg

O

OH

Me

N

SMe

Me

OTESO

ROR2

Me

O OTBS

Me

TIPSO

O

H

Me

TIPSO

OH

O3

Me

O OTBS

Me

TIPSO

NOH

P

OEtO

EtOMe

H

BH3•NH3

TISCl/Et3N

Me

N

SMe

Me

H

OR1 OR

X

N

P

OEtO

EtOMe

O

OH Me

TIPSO

Me

O

N

Me

Me

Ph

OH

O

O OOPg

O

OTrocM e

M e

MeS

NMe

R

I

BuLi

N

SMe

Me

N O

Me

TIPSO

OTBS

N

SMe

Me

OH

Me

TIPSO

PO R

OH

MeO

NMe

MePh

OH

70%

87%

91%95%

(+)-N-Methylephedrine3-methylbutyn-3-olZn(OTf)2 , NEt3, rt

BzCl, NEt3

75%

85%

18-C -6, K2CO3

LiAlH4

94%

t-BuOClEtMgBr, i-P rOH

TBSOTfLiCl, DBU2-Me thylthiazole a ldehyde

S mI2 (75%)Et3 B, Na BH4 (90%)

S OC l2, Et3NBu4NF (75%)

TESC l (79%)AcOH/H2OTP AP(75%)

LDA (86%)TrocCl (91%)

OsO4, P b(OAc)4 (86%)HF•pyrNa OCl2 (76%)

2,4,6-trichlorobenzoylchloride (74%)Zn, NH4Cl, MeOH (84%)3:1 (pyr:[HF•pyr])

Total Synthesis of Epothilone

Jeffrey Bode, E.M. Carreira J. Am. Chem. Soc. 2001, 123, 3611

A complementary approach to the synthesis problem involves bioengineering

Biosynthesis is accomplished by the concatenation of synthetic enzymatic modules

Biosynthesis of Epothilones

Can an analogous modular chemical strategy be developed?

Construction of Aldol Fragments

Me Me

O

Me

OR O

Me

H3 6

Me Me

O

Me

OH

Me

OR

3 6

Me Me

N

Me

O

Me

OR

3 6Me Me

OR N

H

OH

Me Me753

+

+

The Classic Aldol Approach

The Dipolar Cycloaddition Approach

O

O OOH

OHMe

Me

R1

MeS

NMe

O

Retrosynthetic Analysis

O

O

Me

Me

O

HO

OHO

R

Me

N

SMe

Me

Me1

6

15

9

O

O

Me

Me

N

O

OHO

R

Me

N

SMe

Me

Me

O

O

Me

Me

OHO

R

Me

N

SMe

Me

MeC

NO

HOC

OPgON

O

OH

Me

S

NMe

Me

OR

HOC

OPgON

O

OH

Me

S

NMe

Me

OR

N

SMe

Me

NO

OHHO R Me

Analogous Disconnections on Fragments

OHN

SMe

Me HO R Me

OHOH

The Synthesis of Polyketides is Reduced to Modular Nitrile Oxide Cycloadditions

A Parallel Analysis Can be Applied to Others-

Me

N

SMe

Me

OO

MeOH

Me

O OH O

O

Me

OH

Me

Me

OH

O

O

Me

HO

Me Me

OH

Me

O

Me

O

NH2

O

HO

O

OH

O

Me Me

O

O

Me

O

OH

O

OOOMe

MeMe

N

OMe

O

O

O

Epothilone B tubulin polymerization promoter

Discodermolidetubulin polymerization promoter

Laulimalidetubulin polymerization promoter

Rhizoxintubulin polymerization inhibitor

Nitrile Oxide Cycloadditions: Aldol Equivalents

CN

O

R1

R2

R2

N

R1

O+

Regio- and stereoselectivity can be difficult to control

CN

OMe

OMgBr CH2Cl2

Me

OH

N O

+

99:1 syn:anti

The Kanemasa Observation:

S. Kanemasa J. Am. Chem. Soc. 1994, 116, 2324

Are such cycloadditions compatible with more highly functionalized edducts?

If so…

Generalized, versatile strategy for polyketide synthesisConvergent fragment coupling of polyketides

A General Strategy for the Construction of Polyketides:

TBSO N

MeMe OH

Me

O

Me

Me

OH

Me

OH

Me

TBSO N

H

Me

OHTBSO N

H

Me

OH

TBSO N

MeMe OH

O

Me

TBSO N

MeMe OH

Me

O

TBSO N

MeMe OH

Me

O

anti synsyn syn

syn antianti anti

P

N

Cl

O

EtOEtO

Me

OH

MeOH

N O

MeTBSOMe

N

SMe N

SMe

O

P

NO

EtOEtO

Me

O

MeOH

P

NO

EtOEtO

Me

O

MeOTBS

+

iPrOH

EtMgBr

CH2Cl2, rt77 %

87 %

LiHMDS-78 °C → rt

TBSOTf,Et2N

iPr90 %

OTIPS OTIPS

OTIPSOTIPS

O

O OOH

OHMe

Me

MeS

NMe

O

Fragment SynthesisEpothilone A

N

SMe

H

O

EtOP Me

OEtO

Me

N O

OH

Me

Me

N O

OH

N

SMe

Me

OH

Me

Me

N O

N

SMe

O

EtOP

H

OEtO

Me

NOH

+

79%

93%

LiCl, DBUMeCN

TPAP, NMO

CH2Cl2, rt

Synthesis of Epothilone BO

O OOH

OHMe

Me

MeS

NMe

OMe

O

Me

N O

N

SMe

Me

Mg

BrBr

TIPSO

Me

MgBr

Me

N O

N

SMe HO Me Me

TIPSO

81% 10:1 dr

Epothilone B

Chelation-Controlled Addittion

O

O OOH

OHMe

Me

MeS

NMe

OMe

NOH

R1 Cl

R4

OHR3

R2

R1

N O

R4

OHR3 R2

+iPrOH

EtMgBr

CH2Cl20 °C - rt1 equiv 1.0 - 1.3 equiv

1,2-syn to hydroxyl moeity

N

Me

Pg1OOH

OH

Me

Me

N

Me

Pg1O

Me

O

Me OH

N

Me

Pg1OOH

OH

MeMe

OH

MeMe

OH

Me

Me

N

Me

Pg1O

Me

O

Me OH

N

Me

Pg1O

Me

O

Me OH

N

Me

Pg1O

Me

O

Me OH

N

Me

Pg1O

Me

O

Me OH

N

Me

Pg1O

Me

O

Me OH

N

Me

Pg1O

Me

O

Me OH

N

Me

Pg1O

Me

O

Me OH

ROH

RCNO

anti-anti-syn syn-syn-syn syn-anti-syn anti-syn-syn

syn-anti-dyn anti-syn-syn anti-anti-syn syn-syn-syn

Key Methodology: Modular Synthesis of Polypropionates

General Protocol

TBSO

H

NOH

Me

TBSO

Me

Me

N O

OHMe

1. tert-BuOCl, CH2Cl2,

2. Allylic Alcohol,iPrOH, EtMgBrCH2Cl2, 0oC - rt

-78 oC

J. W. Bode, N. Fräfel, D. Muri, E. M. Carreira Angew. Chem. 2001, 113, 2128; J. W. Bode, N. Fräfel, D. Muri, E. M. Carreira Angew. Chem. Int. Ed. Engl. 2001, 40, 2082.

TBSO

H

NOH

Me

TBSO

H

NOH

Me

TBSO

H

N

Me

TBSO

H

NOH

Me

TBSO

H

NOH

Me

Me

Me

OH

Me

Me

OH

Me

OH

Me

Me

OH

Me

Me

OH

TBSO

Me

Me

N O

OHMe

TBSO

Me

Me

N O

OHMe

TBSO

Me

Me

N O

OHMe

TBSO

Me

Me

N O

OHMe

TBSO

Me

Me

N O

OH

82 %

87 %

68 %

73 %

83 %

1

2

3

4

5

Entry NO Precursor Allylic Alcohol Cycloadduct Yield

OH

A General Strategy for the Construction of Polyketides:

TBSO

Me

Me

N O

OHMe

TBSO

Me

Me

N O

OHMe

TBSO

Me

Me

N O

OHMe

TBSO

Me

Me

N O

OHMe

TBSO

Me

Me

O OH

OTESMe

TBSO

Me

Me

O OH

OTESMe

TBSO

Me

Me

O OH

OTESMe

TBSO

Me

Me

O OH

OTESMe

1. TESCl, NEt3

2. Ra-Ni, H2, B(OH)3

Yields: 90-95%

RO

Me

Me

O OH

OR1Me

RO

Me

Me

O OH

OR1Me

RO

Me

Me

O OH

OR1Me

RO

Me

Me

O OH

OR1Me

RO

Me

Me

NOH

OHMe

RO

Me

Me

NOH

OHMe

RO

Me

Me

NOH

OH

TBSO

Me

Me

NOH

OH

Me

Me

Stereochemical Correlations

cis olefin syn aldol

trans olefin anti aldol

Me

Me

O

OH

Amphotericin: The Clinician's Drug of Choice for Fungal Infections

Me O

O OH OH

OH

OH OH

OH

O

OMycosamine

MeHO

Me

CO2H

OH

II. Future Applications of the Catalytic Chemistry to Bio-organic Chemistry

Serious Side effects; much interpatient variation

Mechanism of action largely unknown

Amphotericin: The Clinician's Drug of Choice for Fungal Infections

Me O

O OH OH

OH

OH OH

OH

O

OMycosamine

MeHO

Me

CO2H

OH

PolyeneAmphotericin B

Nystatin

FormulationLiposomalLipid ComplexColloidal DispersionLiposomal

NameAmbisomeAbelcetAmphotecAmphocilNyotran

CompanyNeXstarThe Liposome Co.Sequua in USAZeneca in UKAronex

StatusMarketed in UKMarketed in UK and USAPhase 4MarketedPhase 2/3

Me O

O OH OH

OH

OH OH

OH

O

OH

MeHO

Me

CO2H

OH

Rigid, Hydrophobic(Polyene) Spacer

Hydrophilic(Polyol) Spacer

Hairpin TurnsHairpin Turns

Structural Analysis: Amphotericin

The chemoselective synthetic manipulation of this natural product, as well as the construction of “smart” designed probes is non-trivial

“…however, we are aware that selective modification of the hydroxyl groups involves very difficult chemical synthesis task…probably only through synthesis can derivatives be prepared…”

Molecular Pharmacology 1997, 52, 560

Amphotericin B

Channel Model

Extrazellulär

Cytoplasma

AmBCholesterin

O O

OH

O

O

O

CH3

NH2

H3C

HOCH3

H3C

OH OH

OH

OH OH

OHOH

COOH

OHAmphotericin B

Me O

O OH OH

OH

OH OH

OH

O

OMycosamine

MeHO

Me

CO2H

OH

Calculations suggest that it is critical in organizing channel

Implicated in the formation of double channels

Suggested to interact with sterol

Me

OH

MeHO

O

Me

OH

HO MeMeMe

OH

Me

OH

HO

O

Erythronolide A Seco-Acid

Ongoing Applications in Polyketide Syntheses

O

O

O

OH

OH

Me

Me

Me

OH

OH

Me

Me

Me

Me

HO

Dieter Muri

Syn-Propionate aldols

TBSO

OHMe

O

Me

N

Me

OH

TBSO

Me

O

Me

Me

N

N

Me

O

Me HO

N

Me

O

MeMe

TBSO

HO Me

OH

Me

N

Me

O

Me Me

EtIN

Me

Me

O

Me

TBSO

O

+

91 % 65 %

+

Fragment Coupling O

O

O

OH

OH

Me

Me

Me

OH

OH

Me

Me

Me

Me

HO

O

N

OHO

Me

Me

Me

O

N

Me

Me

Me

Me

HO

Et

O

OHOH

Me

Me

Me

OH

OH

O

Me

Me

HO

OO

NH

N

OHO

Me

Me

Me

O

N

Me

Me

Me

Me

HO

N N

Me Me

O O

MeMe OHMe

HO

Me

OHMe

OH

O

O

OHOH

Me

Me

Me

OH

OH

Me

Me

Me

Me

HO

O

Diversity Oriented Synthesis

Arbeitsgruppe

P. Aschwanden

N. Becker

D. Cereghetti

C. Czeckelius

L. Fader

R. Faessler

A. Fettes

C. Fischer

L. Scherer

J. Waser

T. Watanabe

N. Wurtz

J. Yasuoka

P. Zarotti

W. Zhao

A. Zumbuehl

S. Jonnson

T. Knoepfel

A. Lerchner

N. Lohse

C. Marti

D. Muri

S. Reber

T. Ritter

Arnold and Mabel Beckman Foundation

Pharmacia Upjohn

Merck & Co.Pfizer Inc.Eli Lilly

David and Lucile Packard Foundation

Camille and Henry Dreyfus FoundationAlfred P. Sloan Foundation

Astra-ZenecaAT & T

Novartis

ETH-Zürich and KGF FundsF. Hoffmann-La Roche

Schweizerischer Nationalfonds