Deconstruction of Aspartic Protease Inhibitors Enables Fragment‐Based Discovery of Plasmepsin V Inhibitors

Malaria, caused by the Plasmodium parasites , remains a major global health burden, and resistance to current antimalarials drives the need for drugs with new mechanisms of action. Plasmepsin V (PMV), an essential aspartic protease required for PEXEL processing and protein export, is structurally divergent from human aspartic proteases, offering a path to selective inhibition. Here, we applied a structure‐informed, deconstruction‐based approach to identify non‐peptidomimetic PMV inhibitors by mining catalytic dyad‐binding motifs from experimentally solved aspartic protease–inhibitor complexes and assembling a focused fragment‐like library. Fragment‐inspired model compounds based on pyrrolidine, piperidine, and piperazine cores showed measurable PMV inhibition in a Forster resonance energy transfer (FRET)‐based assay, and a trans ‐3,4‐disubstituted pyrrolidine hit ( 7a ; IC ₅₀ 70  µM) was selected for optimization. Guided by PMV structural data and a renin cocrystal structure of a related pyrrolidine inhibitor, we explored SAR around substituents intended to engage the S1 and S2 regions. Optimization identified N‐ sulfonamide analogs bearing two aromatic substituents as a preferred chemotype, with meta ‐substitution on the N‐ aryl group improving potency to the low micromolar range. The most potent compounds obtained displayed ~10 µM potency, establishing a promising non‐peptidomimetic scaffold for further development of selective PMV inhibitors.