Nagamune Kisaburo, Moreno Silvia N J, Sibley L David
Department of Molecular Microbiology, Washington University School of Medicine, 660 S. Euclid Ave., St. Louis, MO 63110, USA.
Antimicrob Agents Chemother. 2007 Nov;51(11):3816-23. doi: 10.1128/AAC.00582-07. Epub 2007 Aug 13.
Artemisinin is a plant sesquiterpene lactone that has become an important drug for combating malaria, especially in regions where resistance to other drugs is widespread. While the mechanism of action is debated, artemisinin has been reported to inhibit the sarcoplasmic endoplasmic reticulum Ca(2+) ATPase (SERCA) in the malaria parasite. Artemisinin is also effective against Toxoplasma in vitro and in vivo, although it is less potent and, hence, is generally not used therapeutically to treat toxoplasmosis. To explore the mechanism of action, we generated chemically derived mutants of Toxoplasma gondii that were resistant to growth inhibition by this compound in vitro. Three artemisinin-resistant (ART(r)) mutant clones that differed in their sensitivities in vitro by three- to fivefold compared with that of the wild-type parasites were obtained. ART(r) mutants were cross-resistant to other derivatives of artemisinin, the most potent of which was artemisone. Resistance was not due to molecular alterations or differences in the expression of SERCA or other putative targets, such as proteins that code for multidrug resistance or translationally controlled tumor protein. ART(r) mutants were resistant to the induction of protein secretion from micronemes, a calcium-dependent process that is triggered by artemisinin. ART(r) mutants were not cross-resistant to secretion induced by thapsigargin but were more sensitive and were unable to regulate cytoslic calcium following treatment with this compound. These studies implicate calcium homeostasis in the mechanism of action of artemisinins against apicomplexan parasites.
青蒿素是一种植物倍半萜内酯，已成为对抗疟疾的重要药物，尤其是在对其他药物耐药性普遍存在的地区。虽然其作用机制存在争议，但据报道青蒿素可抑制疟原虫中的肌浆内质网Ca(2+)ATP酶（SERCA）。青蒿素在体外和体内对弓形虫也有效，尽管其效力较低，因此一般不用于治疗弓形虫病。为了探究其作用机制，我们构建了化学衍生的弓形虫突变体，这些突变体在体外对该化合物的生长抑制具有抗性。获得了三个青蒿素抗性（ART(r)）突变体克隆，它们在体外的敏感性与野生型寄生虫相比相差三到五倍。ART(r)突变体对青蒿素的其他衍生物具有交叉抗性，其中最有效的是蒿甲醚。抗性并非由于SERCA或其他假定靶点（如编码多药耐药性的蛋白质或翻译控制肿瘤蛋白）的分子改变或表达差异所致。ART(r)突变体对由微线体分泌蛋白质的诱导具有抗性，微线体分泌蛋白质是一个由青蒿素触发的钙依赖性过程。ART(r)突变体对毒胡萝卜素诱导的分泌没有交叉抗性，但更敏感，并且在用该化合物处理后无法调节胞质钙。这些研究表明钙稳态参与了青蒿素对顶复门寄生虫的作用机制。
