Proteolysis-targeting chimeras as a novel strategy for targeting epigenetic mechanisms in Plasmodium falciparum parasites

Abstract

With the continuous emergence and spread of resistance to the only remaining effective antimalarial, artemisinin, novel drugs with novel modes-of-action (MoA) are needed. Current antimalarial efforts aim to identify small-molecule inhibitors that bind to the active site of essential target proteins, thereby killing the causative organism, Plasmodium falciparum. However, such active site-dependent inhibitors face the major challenge of resistance development. We therefore proposed the use of Proteolysis-targeting chimeras (PROTACs) that can induce the degradation of a target protein via the cells ubiquitin-proteasome system rather than inhibiting it. Ideally, for PROTACs to be effective in malaria research, they should bind to essential proteins, such as epigenetic modulators. Here we aimed to evaluate if PROTACs (MZ1 and VZ185) targeting epigenetic mechanisms have activity against P. falciparum parasites and if they are active due to a unique MoA. Both PROTACs were found to be active against the asexual developmental stages of P. falciparum parasites, while VZ185 was found to also be active against the gametocyte stages, implicating multistage activity. The PROTACs additionally showed nanomolar activity against different drug-resistant parasite strains with potentially improved selectivity towards the parasite compared to mammalian cells. We also confirmed that the activity of the PROTACs in the parasite is reliant on the entire molecule and not only its individual ligand components. We also found that the cis-isomers of the PROTACs have improved or similar activity against the parasite, indicating that the interaction with the E3 ligase in the parasite is not stereochemically driven. Co-treatment of the parasites with the PROTACs and a proteasome inhibitor, epoxomicin, did not affect the activity of MZ1 while VZ185 was adversely affected, indicating that the effect of the latter relies on active proteolysis. We determined if the biological action of VZ185 in the parasite was related to epigenetic mechanisms by studying the changes in the morphology of parasites treated with the PROTAC. We found that asexual blood stage parasites treated with VZ185, and cis-VZ185, altered the parasite's ability to produce daughter cells and inhibit haemoglobin metabolism. This study demonstrated that PROTACs could be used as potential antimalarial compounds and a tool to probe parasite biology.