Targeting Artemisinin-Resistant Malaria by Repurposing the Anti-Hepatitis C Virus Drug Alisporivir.

The emergence of Plasmodium falciparum resistance raises an urgent need to find new antimalarial drugs. Here, we report the rational repurposing of the anti-hepatitis C virus drug, alisporivir, a nonimmunosuppressive analog of cyclosporin A, against artemisinin-resistant strains of P. falciparum. In silico docking studies and molecular dynamic simulation predicted strong interaction of alisporivir with Pf Cyclophilin 19B, confirmed through biophysical assays with a K _d value of 354.3 nM. Alisporivir showed potent antimalarial activity against chloroquine-resistant ( Pf RKL-9 with resistance index [Ri] 2.14 ± 0.23) and artemisinin-resistant ( Pf Kelch13 ^R539T with Ri 1.15 ± 0.04) parasites. The Ri is defined as the ratio between the IC ₅₀ values of the resistant line to that of the sensitive line. To further investigate the mechanism involved, we analyzed the expression level of Pf Cyclophilin 19B in artemisinin-resistant P. falciparum ( Pf Kelch13 ^R539T ). Semiquantitative real-time transcript, Western blot, and immunofluorescence analyses confirmed the overexpression of Pf Cyclophilin 19B in Pf Kelch13 ^R539T . A 50% inhibitory concentration in the nanomolar range, together with the targeting of Pf Cyclophilin 19B, suggests that alisporivir can be used in combination with artemisinin. Since artemisinin resistance slows the clearance of ring-stage parasites, we performed a ring survival assay on artemisinin-resistant strain Pf Kelch13 ^R539T and found significant decrease in parasite survival with alisporivir. Alisporivir was found to act synergistically with dihydroartemisinin and increase its efficacy. Furthermore, alisporivir exhibited antimalarial activity in vivo . Altogether, with the rational target-based Repurposing of alisporivir against malaria, our results support the hypothesis that targeting resistance mechanisms is a viable approach toward dealing with drug-resistant parasite.