Inhibition or Knockdown of ABC Transporters Enhances Susceptibility of Adult and Juvenile Schistosomes to Praziquantel

Parasitic flatworms of the genus Schistosoma cause schistosomiasis, a neglected tropical disease that affects hundreds of millions. Treatment of schistosomiasis depends almost entirely on the drug praziquantel (PZQ). Though essential to treating and controlling schistosomiasis, a major limitation of PZQ is that it is not active against immature mammalian-stage schistosomes. Furthermore, there are reports of field isolates with heritable reductions in PZQ susceptibility, and researchers have selected for PZQ-resistant schistosomes in the laboratory. P-glycoprotein (Pgp; ABCB1) and other ATP binding cassette (ABC) transporters remove a wide variety of toxins and xenobiotics from cells, and have been implicated in multidrug resistance (MDR). Changes in ABC transporter structure or expression levels are also associated with reduced drug susceptibility in parasitic helminths, including schistosomes. Here, we show that the activity of PZQ against schistosome adults and juveniles ex vivo is potentiated by co-administration of either the highly potent Pgp inhibitor tariquidar or combinations of inhibitors targeting multiple ABC multidrug transporters. Adult worms exposed to sublethal PZQ concentrations remain active, but co-administration of ABC transporter inhibitors results in complete loss of motility and disruption of the tegument. Notably, juvenile schistosomes (3–4 weeks post infection), normally refractory to 2 µM PZQ, become paralyzed when transporter inhibitors are added in combination with the PZQ. Experiments using the fluorescent PZQ derivative (R)-PZQ-BODIPY are consistent with the transporter inhibitors increasing effective intraworm concentrations of PZQ. Adult worms in which expression of ABC transporters has been suppressed by RNA interference show increased responsiveness to PZQ and increased retention of (R)-PZQ-BODIPY consistent with an important role for these proteins in setting levels of PZQ susceptibility. These results indicate that parasite ABC multidrug transporters might serve as important targets for enhancing the action of PZQ. They also suggest a potentially novel and readily-available strategy for overcoming reduced PZQ susceptibility of schistosomes.

血吸虫属（Schistosoma）的寄生扁形虫可引发血吸虫病，这是一种影响数亿人群的被忽视热带病。血吸虫病的治疗几乎完全依赖吡喹酮（praziquantel, PZQ）。尽管吡喹酮是治疗和控制血吸虫病的必需药物，但其存在重大局限：对未成熟的哺乳动物宿主期血吸虫无活性。此外，已有报道称野外分离株出现了可遗传的吡喹酮敏感性降低现象，且研究人员已在实验室中筛选出耐吡喹酮的血吸虫。
P-糖蛋白（P-glycoprotein, Pgp; ABCB1）及其他ATP结合盒（ATP binding cassette, ABC）转运蛋白可从细胞中清除多种毒素与外源性物质，并与多药耐药（multidrug resistance, MDR）相关。ABC转运蛋白的结构或表达水平改变，也与包括血吸虫在内的寄生蠕虫的药物敏感性降低有关。
本研究证实，联合使用强效Pgp抑制剂他利喹达（tariquidar），或靶向多种ABC多药转运蛋白的抑制剂组合，可增强吡喹酮对体外血吸虫成虫及童虫的活性。暴露于亚致死浓度吡喹酮的成虫仍可保持活动能力，但联合使用ABC转运蛋白抑制剂后，其运动能力完全丧失，体表皮层也遭到破坏。值得注意的是，通常对2 µM吡喹酮具有耐药性的血吸虫童虫（感染后3~4周），在联合使用转运蛋白抑制剂与吡喹酮后会出现麻痹现象。
使用荧光标记吡喹酮衍生物(R)-PZQ-BODIPY开展的实验结果，与转运蛋白抑制剂可提升吡喹酮虫内有效浓度的结论一致。通过RNA干扰（RNA interference, RNAi）抑制ABC转运蛋白表达的成虫，对吡喹酮的反应性增强，且(R)-PZQ-BODIPY的滞留量也有所提升，这表明这些蛋白在调控吡喹酮敏感性水平中发挥重要作用。
上述结果表明，寄生虫ABC多药转运蛋白可作为增强吡喹酮药效的重要靶点。同时，本研究也提出了一种潜在的新型、易于实施的策略，用以克服血吸虫的吡喹酮敏感性降低问题。