Targeting NAPE-PLD: assessing its potential in the fight against malaria

Abstract

Malaria remains a major global challenge with increasing artemisinin-resistant Plasmodium falciparum. This calls for an increased need for discovering novel ring-stage drug targets. In this study, we aimed to investigate the compensatory mechanism of N-acyl phosphatidylethanolamine Phospholipase D (NAPE-PLD) and 14 in P. falciparum and assess if they could be potential ring-stage targets. Attempts to study NAPE-PLD 11 and 14 via gene knockdown and knockout were met with technical limitations such as low transfection efficiency and plasmid design constraints. Thus, overexpression cell lines were generated to assess drug sensitivity against LEI-401. Unfortunately, the NAPE-PLD 11 overexpression line did not show a statistically significant IC50 shift, suggesting limited interaction. CETSA-MS analysis failed to detect LEI-401 binding with NAPE-PLD proteins, revealing instead two alternative targets, choline/ethanolamine phosphotransferase (CEPT) and coproporphyrinogen-III oxidase (CPO). Docking analysis indicated stronger LEI-401 binding affinity for CEPT than P. falciparum NAPE-PLD proteins, supporting CEPT’s involvement in lipid metabolism as a viable target. These findings suggest that while NAPE-PLD may not be a primary target of LEI-401, CEPT offers potential as an alternate target. Future studies will focus on optimising genetic constructs for double overexpression and further validating LEI-401’s interaction with CEPT, contributing to the development of ring-stage antimalarials.