Artemisinins are extracted from sweet wormwood (Artemisia annua) and are the most potent antimalarials available1, rapidly killing all asexual stages of Plasmodium falciparum2. Artemisinins are sesquiterpene lactones widely used to treat multidrug-resistant malaria1, a disease that annually claims 1 million lives. Despite extensive clinical and laboratory experience3, 4, 5 their molecular target is not yet identified. Activated artemisinins form adducts with a variety of biological macromolecules, including haem, translationally controlled tumour protein (TCTP) and other higher-molecular-weight proteins6. Here we show that artemisinins, but not quinine or chloroquine, inhibit the SERCA orthologue (PfATP6) of Plasmodium falciparum in Xenopus oocytes with similar potency to thapsigargin (another sesquiterpene lactone and highly specific SERCA inhibitor). As predicted, thapsigargin also antagonizes the parasiticidal activity of artemisinin. Desoxyartemisinin lacks an endoperoxide bridge and is ineffective both as an inhibitor of PfATP6 and as an antimalarial. Chelation of iron by desferrioxamine abrogates the antiparasitic activity of artemisinins and correspondingly attenuates inhibition of PfATP6. Imaging of parasites with BODIPY-thapsigargin labels the cytosolic compartment and is competed by artemisinin. Fluorescent artemisinin labels parasites similarly and irreversibly in an Fe2+-dependent manner. These data provide compelling evidence that artemisinins act by inhibiting PfATP6 outside the food vacuole after activation by iron.

Artemisinins target the SERCA of Plasmodium falciparum

Since the first reports of chloroquine-resistant falciparum malaria in southeast Asia and South America almost half a century ago, drug-resistant malaria has posed a major problem in malaria control. By the late 1980s, resistance to sulfadoxine-pyrimethamine and to mefloquine was also prevalent on the Thai-Cambodian and Thai-Myanmar (Thai-Burmese) borders, rendering them established multidrug-resistant (MDR) areas. Chloroquine resistance spread across Africa during the 1980s, and severe resistance is especially found in east Africa. As a result, more than ten African countries have switched their first-line drug to sulfadoxine-pyrimethamine. Of great concern is the fact that the efficacy of this drug in Africa is progressively deteriorating, especially in foci in east Africa, which are classified as emerging MDR areas. Urgent efforts are needed to lengthen the lifespan of sulfadoxine-pyrimethamine and to identify effective, affordable, alternative antimalarial regimens. Molecular markers for antimalarial resistance have been identified, including pfcrt polymorphisms associated with chloroquine resistance and dhfr and dhps polymorphisms associated with sulfadoxine-pyrimethamine resistance. Polymorphisms in pfmdr1 may also be associated with resistance to chloroquine, mefloquine, quinine, and artemisinin. Use of such genetic information for the early detection of resistance foci and future monitoring of drug-resistant malaria is a potentially useful epidemiological tool, in conjunction with the conventional in-vivo and in-vitro drug-sensitivity assessments. This review describes the various features of drug resistance in Plasmodium falciparum, including its determinants, current status in diverse geographical areas, molecular markers, and their implications.

https://www.thelancet.com/pdfs/journals/laninf/PIIS1473-3099(02)00239-6.pdf

Epidemiology of drug-resistant malaria.

https://www.thelancet.com/pdfs/journals/lancet/PIIS0140-6736(04)16767-6.pdf

Mefloquine resistance in Plasmodium falciparum and increased pfmdr1 gene copy number.

Plasmodium falciparum chloroquine resistance is a major cause of worldwide increases in malaria mortality and morbidity. Recent laboratory and clinical studies have associated chloroquine resistance with point mutations in the gene pfcrt. However, direct proof of a causal relationship has remained elusive and most models have posited a multigenic basis of resistance. Here, we provide conclusive evidence that mutant haplotypes of the pfcrt gene product of Asian, African, or South American origin confer chloroquine resistance with characteristic verapamil reversibility and reduced chloroquine accumulation. pfcrt mutations increased susceptibility to artemisinin and quinine and minimally affected amodiaquine activity; hence, these antimalarials warrant further investigation as agents to control chloroquine-resistant falciparum malaria.

http://science.sciencemag.org/content/sci/298/5591/210.full.pdf

Chloroquine resistance in Plasmodium falciparum malaria parasites conferred by pfcrt mutations.

Artemisinin: mechanisms of action, resistance and toxicity.

On the western border of Thailand, Plasmodium falciparum has become resistant to almost all antimalarial agents. The molecular basis of resistance in these parasite populations has not been well characterized. This study assessed genetic polymorphisms in the pfmdr1 gene in 54 parasites collected from the western border of Thailand to determine the relationship of pfmdr1 copy number and codon mutations with parasite sensitivities to mefloquine, chloroquine, halofantrine, quinine, and artesunate assessed in vitro. A point mutation at codon 86 (resulting in a change of Asn to Tyr) was associated with a significantly lower 50% inhibitory concentration (IC(50)) of mefloquine (median, 9 ng/ml versus 52.4 ng/ml; P = 0.003). Overall 35% of the isolates (19 of 54) had an increase in pfmdr1 copy number, and all 19 carried the wild-type allele at codon 86. Increased pfmdr1 copy number was associated with higher IC(50)s of mefloquine (P = 0.04) and artesunate (P = 0.005), independent of polymorphism at codon 86. The relationship between pfmdr1 and resistance to structurally distinct antimalarial agents confirms the presence of a true multidrug-resistant phenotype.

C. Cassar

Papers

The pfmdr1 gene is associated with a multidrug-resistant phenotype in Plasmodium falciparum from the western border of Thailand.