Malaria remains a global health concern as drug resistance threatens treatment programs. We identified a piperidine carboxamide (SW042) with anti-malarial activity by phenotypic screening. Selection of SW042-resistant Plasmodium falciparum (Pf) parasites revealed point mutations in the Pf_proteasome β5 active-site (Pfβ5). A potent analog (SW584) showed efficacy in a mouse model of human malaria after oral dosing. SW584 had a low propensity to generate resistance (minimum inoculum for resistance [MIR] >10⁹) and was synergistic with dihydroartemisinin. Pf_proteasome purification was facilitated by His₈-tag introduction onto β7. Inhibition of Pfβ5 correlated with parasite killing, without inhibiting human proteasome isoforms or showing cytotoxicity. The Pf_proteasome_SW584 cryoelectron microscopy (cryo-EM) structure showed that SW584 bound non-covalently distal from the catalytic threonine, in an unexplored pocket at the β5/β6/β3 subunit interface that has species differences between Pf and human proteasomes. Identification of a reversible, species selective, orally active series with low resistance propensity provides a path for drugging this essential target.

疟疾仍然是一个全球性的健康问题，因为耐药性威胁着治疗计划。我们通过表型筛选确定了一种具有抗疟疾活性的哌啶羧酰胺 （SW042）。SW042 耐药性恶性疟原虫 （Pf） 寄生虫的选择揭示了 Pf_蛋白酶体 β5 活性位点 （Pfβ5） 的点突变。口服给药后，强效类似物 （SW584） 在人类疟疾小鼠模型中显示出疗效。SW584 产生耐药性倾向较低 （最低耐药接种量 [MIR] >10⁹），并且与双氢青蒿素具有协同作用。Pf_蛋白酶体的纯化是通过 His₈ 标签引入 β7 来促进的。抑制 Pfβ5 与寄生虫杀伤相关，不抑制人蛋白酶体亚型或显示细胞毒性。Pf_proteasome_SW584冷冻电子显微镜 （cryo-EM） 结构显示，SW584 在催化苏氨酸远端非共价结合，位于 β5/β6/β3 亚基界面处一个未探索的口袋中，该口袋在 Pf 和人蛋白酶体之间存在物种差异。鉴定具有低耐药倾向的可逆、物种选择性、口服活性系列为这一重要靶标的药物提供了一条途径。