Design, synthesis and biological evaluation of Rac1-targeting carbazole-pyrimidines as host-directed antimalarials

In this research, we validate that inhibitors of host-cell-Rac1 can provide a novel strategy for antimalarial therapy. Host-directed therapies represent an emerging strategy to overcome antimalarial drug resistance by targeting host factors essential for Plasmodium survival. The small GTPase Rac1, the only Rac present in mature human erythrocytes, has been previously identified as a key host factor supporting Plasmodium falciparum intraerythrocytic development. Building on the known Rac1 inhibitor EHop-016, which had shown good antiplasmodial activity in vitro in a previous study, we designed and synthesized a series of carbazole-pyrimidine analogs to identify derivatives with improved potency and selectivity. Replacement of the pyrimidine core resulted in inactive compounds, demonstrating its essential role. Systematic chemical modifications at positions 2-, 5-, and 6- of the pyrimidine ring revealed that both electronic and steric factors critically influence antiplasmodial activity. Several derivatives displayed submicromolar inhibitory concentrations against both chloroquine-sensitive D10 and chloroquine-resistant W2 P. falciparum strains. Their cytotoxicity was evaluated on human cells to determine the selectivity index. Based on these results, five compounds were selected to confirm Rac1inhibition in epithelial cells. Among them, compounds 11 and 5e emerged as lead candidates, exhibiting IC₅₀ values of 46.6 nM and 76.0 nM against D10 parasites, with selectivity indices >45. Both compounds showed increased potency (IC₅₀ < 250 nM) and selectivity towards the resistant W2 strain compared with EHop-016. Stage-specific analyses revealed that both compounds exert antimalarial activity throughout the entire intraerythrocytic developmental cycle. These findings further validate Rac1 as a host target for antimalarial therapy and identify novel EHop-016 derivatives as a promising framework for the development of host-directed antimalarial agents.