The Mode of Action and Possible Target

of Artemisinin

Mike Van Linn

Chemistry 496

23 April 2004

Outline

Introduction Malaria Artemisinin

Rationale for Research Modes of Action

Iron-Oxo route Epoxidation reactions Alkylation reactions

The Target of Artemisinin

Introduction

Malaria Four species of

Plasmodium Infects 200 million

people annually 1 million lethal Resistance to current

drugs

Introduction

Artemisinin Natural product

extracted from sweet wormwood, Artemisia annua

Used by Chinese for over 2000 years

A. absinthium used to make absinthe

Rationale for Research

Anti-malarial Activity of Artemisinin Artemisinin Derivatives

Used currently for life threatening cases

Rationale for Research

Anti-malarial Activity of Artemisinin Artemisinin Derivatives

• Used currently for life threatening cases Drug Resistance of Plasmodium

• Malaria spreading• Synthesis of new drugs

Possible Modes of Action

Iron-Oxo Route Epoxidation Reactions Alkylation Reactions

Iron-Oxo Route

Donation of Oxygen from Peroxide Bridge to Iron Generate Fe(IV)=O

No Support from Raman Resonance Data Signal/Noise < 2 Should be ~10 or 20

Epoxidation Reactions

+ ARTEMISININ NO EPOXIDE FORMATION

O

MnIITPP or FeCl2

+

ORMnIITPP or FeCl2 EPOXIDE FORMATION

NH

NN

HN

Ph

PhPh

Ph

MnII

Fe

ClCl

Na+ -OCl

Robert, et al

Cazelles, et al

Cazelles, et al

1,5 H Shift Possible???

Critical Distance Calculated to be 2.1Å

Exceeded in Stable Conformation

Boat-like Conformation (High energy state) Houk

Comparing Route 1 and 2

Route 1 Dominant to Route 2 90/10 ratio from isolated products

Artemisinin + MnIITPP 1,5 H shift?

Route 1 Biologically Active Route 2 Inactive Stereochemistry Effecting Alkylation

Cazelles, et al

Robert, et al

Mode of Action

Route 1 Dominant Alkyl radical formation

from reduction of peroxide bridge

Derivatives Used Observe correlation of

alkylating ability to drug activity

Alkylate MnIITPP Pharm. active

The Target

Alkylation of Heme within Infected Erythrocytes (RBC’s) Free heme in food vacuole of erythrocyte Cleavage of peroxide bond Alkylation of heme or specific parasite proteins

can occur Too General…

The Target, More Specifically

Sarco/Endoplasmic Reticulum Ca2+-ATPase (SERCA) Enzyme PfATP6 gene sequence Testing the hypothesis

Heme Not Required? Free heme blocked with Ro 40-4388 protease

inhibitor Localized in the Food Vacuole?

Fluorescent labeled artemisinin

Conclusions

Malaria Remains as a Problem Resistant strains

Anti-malarial Activity of Artemisinin Mode of Action is Now Understood

Alkylation via route 1 A Specific Target Found

PfATP6 gene sequence of the SERCA enzyme

Fe2+ is required Activity not localized in the food vacuole

References

1. Robert, Anne, et al. “From Mechanistic Studies on Artemisinin Derivatives to New Modular Antimalarial Drugs.” Accounts of Chemical Research, 2002, Vol. 35, pp. 167-174.

2. Cazelles, Jerome, et al. “Alkylating Capacity and Reaction Products of Antimalarial Trioxanes after Activation by a Heme Model.” The Journal of Organic Chemistry, 2002, Vol. 67, Number 3, pp. 609-619.

3. Wu, Wen-Min, et al. “Unified Mechanistic Framework for the Fe(II)-Induced Cleavage of Qinghaosu and Derivatives/Analogues. The First Spin-Trapping Evidence for the Previously Postulated Secondary C-4 Radical.” J. Am. Chem. Soc., 1998, Vol. 120, pp. 3316-3325.

4. Biot, Christophe, et al. “Synthesis and Antimalarial Activity in Vitro and in Vivo of a New Ferrocene-Chloroquine Analogue.” J. of Medicinal Chemistry, 1997, Vol. 40, pp. 3715-3718.

5. Yarnell, Amanda; “Rethinking How Artemisinin Kills,” Chemical and Engineering News, Aug. 25, 2003, Vol. 81 (24), pp. 6.

6. Eckstein-Ludwig, Ursula, et al. “Artemisinins Target the SERCA of Plasmodium falciparum,” Nature, 2003, Vol. 424, pp.957.

Questions