In silico studies for improving target selectivity of anti-malarial dual falcipain inhibitors vis-à-vis human cathepsins

Dual falcipain-2 (FP-2) and falcipain-3 (FP-3) inhibitors, NM12 and NM15, displayed micromolar inhibitions but they exhibit similar binding affinities for the human cathepsins, thus indicating potential toxicity. The current study aims to develop a model to enhance the selectivity of the falcipain inhibitors vis-à-vis human cathepsins using previously identified dual falcipain 2 and 3 inhibitors, NM12 and NM15. To improve the selectivity of NM12 and NM15, analogs with weaker interactions with the conserved residues in the FPs and hCatK were designed while enhancing the unique interactions for the FPs. In silico analysis was carried out in the S2 subsite of both plasmodium and human proteases which is considered the preferred selective site due to the presence of less conserved residues. The Fasta sequence alignment and active/conserved binding site superimposition show that FPs contain acidic polar residues (Asp234 for FP2 and Glu243 for FP3) while hCatK has a neutral hydrophobic residue (Leu209) at the S2 subsite. Therefore, analogs of NM12 and NM15 were designed to enhance affinity and selectivity by improving interactions with these acidic residues while avoiding interactions with hydrophobic residues in hCatK. Newly designed analogs (NM12H and NM15G) show better selectivity as well as binding affinity towards FPs (ΔG of NM12H: −74.49 kcal/mol for FP2, −70.97 kcal/mol for FP3; ΔG of NM15G: −70.09 kcal/mol for FP2, −74.52 kcal/mol for FP3) as compared to NM12 and NM15. Thus, the selectivity and binding affinity against dual falcipains vis-à-vis human cathepsin were improved using molecular dynamic simulations.