Here, we present remarkable epoxyketone-based proteasome inhibitors with low nanomolar in vitro potency for blood-stage Plasmodium falciparum and low cytotoxicity for human cells. Our best compound has more than 2,000-fold greater selectivity for erythrocytic-stage P. falciparum over HepG2 and H460 cells, which is largely driven by the accommodation of the parasite proteasome for a D-amino acid in the P3 position and the preference for a difluorobenzyl group in the P1 position. We isolated the proteasome from P. falciparum cell extracts and determined that the best compound is 171-fold more potent at inhibiting the β5 subunit of P. falciparum proteasome when compared to the same subunit of the human constitutive proteasome. These compounds also significantly reduce parasitemia in a P. berghei mouse infection model and prolong survival of animals by an average of 6 days. The current epoxyketone inhibitors are ideal starting compounds for orally bioavailable anti-malarial drugs.

Calibr a division of The Scripps Research Institute 11119 N Torrey Pines Rd La Jolla California 92093 USA

Center for Marine Biotechnology and Biomedicine Scripps Institution of Oceanography University of California San Diego 9500 Gilman Drive La Jolla California 92093 USA

Department of Pediatrics School of Medicine University of California San Diego 9500 Gilman Drive La Jolla California 92093 USA

Department Pharmaceutical Sciences College of Pharmacy University of Jordan Amman 11942 Jordan

Institute of Organic Chemistry and Biochemistry of the Czech Academy of Sciences 16610 Prague Czech Republic

Skaggs School of Pharmacy and Pharmaceutical Sciences University of California San Diego 9500 Gilman Drive La Jolla California 92093 USA

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Structural elucidation of recombinant Trichomonas vaginalis 20S proteasome bound to covalent inhibitors