Defining molecular pathways in antimalarial drug resistance

Malaria is one of the deadliest diseases in the world and a serious threat to the progress of affected areas. Plasmodium falciparum is one of the parasites that causes the disease and has a high capacity to develop resistance to drugs, making most therapies ineffective. Only artemisinin-based combination therapies remain effective treatments, however recent studies point to a reduction in their effectiveness. Consequently, it is necessary to identify the main markers of resistance and understand its evolution. It is also important to understand and identify the biological mechanisms of the parasite to circumvent resistance mechanisms or identify new effective drug targets. In this thesis, Clinical trial samples from Africa were analyzed for piperaquine resistance markers and the transcriptional patterns of pfk13, the gene most predictive of artemisinin tolerance, were explored. Duplications of the plasmepsin 2 gene, a marker associated with piperaquine resistance in Southeast Asia, were found in Africa, indicating the possible presence of parasites with mutations favorable to the emergence of piperaquine resistance. Analysis of the pfk13 transcriptional response after treatment with artemether-lumefantrine revealed a correlation between pfk13 downregulation and a longer parasite elimination time. Therefore, the transcriptional response is a possible mechanism of artemisinin tolerance. To explore the mechanism of piperaquine resistance, this work generated strains of P. falciparum genetically modified with duplication of plasmepsin 2 and plasmepsin 3-1. These parasites show greater resistance to high concentrations of piperaquine, a phenotype that was increased through the use of chemical blockers of the PfMDR1 and PfCRT transporters. These data suggest a multigenic phenotype and an interaction of multiple systems present in the parasite’s digestive vacuole. This work also explored the role of the cytoplasmic transporter protein PfMRP1, which has no particular impact on commonly used antimalarial drugs. However, antifolate resistance was observed in a phenotype incompatible with the previously proposed exporter functionality. The accumulation of fluorescent antifolate molecules was reduced in parasites with the pfmrp1 gene disrupted. Through a phylogenetic analysis, we demonstrated that pfmrp1 is distant from other transporters of the same class in several organisms. Therefore, this work proposes PfMRP1 as an import protein.

Project details

Drug Resistance

Dec 2021 — Jun 2022

$3,131

Portugal