A G358S mutation in the Plasmodium falciparum Na+ pump PfATP4 confers clinically-relevant resistance to cipargamin

Abstract

Diverse compounds target the Plasmodium falciparum Na+ pump PfATP4, with cipargamin and (+)-SJ733 the most clinically-advanced. In a recent clinical trial for cipargamin, recrudescent parasites emerged, with most having a G358S mutation in PfATP4. Here, we show that PfATP4G358S parasites can withstand micromolar concentrations of cipargamin and (+)-SJ733, while remaining susceptible to antimalarials that do not target PfATP4. The G358S mutation in PfATP4, and the equivalent mutation in Toxoplasma gondii ATP4, decrease the sensitivity of ATP4 to inhibition by cipargamin and (+)-SJ733, thereby protecting parasites from disruption of Na+ regulation. The G358S mutation reduces the affinity of PfATP4 for Na+ and is associated with an increase in the parasite’s resting cytosolic [Na+]. However, no defect in parasite growth or transmissibility is observed. Our findings suggest that PfATP4 inhibitors in clinical development should be tested against PfATP4G358S parasites, and that their combination with unrelated antimalarials may mitigate against resistance development.

Generation of a mutator parasite to drive resistome discovery in Plasmodium falciparum

“…Notably, selections with the Dd2-Polδ line yielded a G358S mutant that was recently observed in the majority of treatment failures during a Phase II trial of KAE609 (Schmitt et al, 2021), indicating that in vitro evolution with this line can yield outcomes with clinical relevance. This high-level resistance mutation was also observed previously in selections with a dihydroisoquinolone compound (+)-SJ733 (Jimenez-Diaz et al, 2014), as well as in parallel KAE609 selections with our Dd2-Polδ line by another group 48 . Qui et al generated a CRISPR-edited line with the G358S SNV, demonstrating that this mutation is sufficient to drive a large shift in IC 50 and showed that it protects the Na + -ATPase activity of PfATP4 from inhibition by KAE609, but at the cost of lowering the affinity of the protein to Na + 48 .…”

Optimization of 2,3-Dihydroquinazolinone-3-carboxamides as Antimalarials Targeting PfATP4

“…The G358S mutation is the same mutation that was detected in recrudescent parasites from patients treated with KAE609 in phase II clinical trials. 27,38 Dihydroquinazolinone analogues generally exhibited a 10-fold decrease in potency against the triple mutant PfATP4 strain, and analogues 20 and 36 exhibited a 20-fold decrease in potency. In contrast, the dihydroquinazolinone analogues were generally equipotent against the PfATP4 G358S -resistant strain compared to the Dd2 parental parasite strain, except for compounds 5 and 20 that were 10-fold more potent against the PfATP4 G358S -resistant strain.…”

“…Accordingly, several PfATP4 inhibitors have been identified as preclinical candidates, cipargamin (KAE609) ( 1 ), SJ733 ( 2 ), PA21A092 ( 3 ), and GNF-Pf4492 ( 4 ) (Figure ), and both 1 and 2 are in human clinical trials assessing their curative and transmission-blocking efficacy. Concerningly, a G358S mutation in PfATP4 has been shown to confer a clinically relevant level of resistance to KAE609 and SJ733, which necessitates the need to uncover novel PfATP4 inhibitor chemotypes that have a high barrier to resistance and are active against drug-resistant clinical isolates.…”

A Pyridyl-Furan Series Developed from the Open Global Health Library Block Red Blood Cell Invasion and Protein Trafficking in Plasmodium falciparum through Potential Inhibition of the Parasite’s PI4KIIIB Enzyme

“…To identify the protein target and mechanism of action of the pyridyl-furan compounds, we attempted to select for genetic resistance to OGHL250 through repeated rounds of treatment and recovery against engineered “hypermutator” Dd2 parasites that have a higher background error prone DNA replication and are hereafter called Parental parasites. , After three rounds of selection, the DMSO-treated Parental parasites and OGHL250-challenged parasites were subjected to a 72 h growth assay against a dilution series of OGHL250. However, the EC 50 was little changed, indicating no resistance could be selected.…”