Integrative computational and experimental approaches for identifying potent antimalarials by targeting falcipain-2...

Malaria remains a critical global health issue, particularly in tropical and subtropical regions. Understanding its biology and epidemiology is vital for developing effective prevention and control strategies. Falcipain-2 (FP-2), a cysteine protease in Plasmodium falciparum, the deadliest malaria parasite, is essential for parasite survival in red blood cells, making it an attractive drug target. Inhibiting FP-2 disrupts key metabolic processes, leading to parasite death, offering a promising antimalarial drug development avenue. This study utilized computational approaches to design novel antimalarials. We prepared large fragment libraries, Enamine (∼220,174 fragments) and ChemDiv (∼18,713 fragments), for virtual screening against FP-2 (PDB ID: 6JW9). Fragments with binding free energies ≤ -4.0 kcal/mol were selected and evolved into drug-like ligands. SP and XP docking-based screenings prioritized these ligands, refined further using MM-GBSA calculations. Promising ligands underwent 100 ns molecular dynamics simulations to assess conformational changes and complex stability. This study identified two viable inhibitors, T4 and A3, with high affinity, stability, and selectivity towards FP-2 compared to E64.Following computational studies, compound A3 and its ten analogues (KA-series) were synthesized and characterized by using multi-spectroscopic techniques with high purity confirmed by LC-MS. Biological testing against P. falciparum 3D7 strain revealed KA-5 as the most potent compound with an IC₅₀ value of 3.0 μM. Future work will involve synthesizing additional analogue series for an extensive structure-activity relationship (SAR) study. Further experimental validation is essential to develop these compounds as effective therapeutic agents for malaria treatment.