Pseudo-Dipeptide Bearing α,α-Difluoromethyl Ketone Moiety as Electrophilic Warhead with Activity against Coronaviruses

The synthesis of α-fluorinated methyl ketones has always been challenging. New methods based on the homologation chemistry via nucleophilic halocarbenoid transfer, carried out recently in our labs, allowed us to design and synthesize a target-directed dipeptidyl α,α-difluoromethyl ketone (DFMK) 8 as a potential antiviral agent with activity against human coronaviruses. The ability of the newly synthesized compound to inhibit viral replication was evaluated by a viral cytopathic effect (CPE)-based assay performed on MCR5 cells infected with one of the four human coronaviruses associated with respiratory distress, i.e., hCoV-229E, showing antiproliferative activity in the micromolar range (EC(50) = 12.9 ± 1.22 µM), with a very low cytotoxicity profile (CC(50) = 170 ± 3.79 µM, 307 ± 11.63 µM, and 174 ± 7.6 µM for A549, human embryonic lung fibroblasts (HELFs), and MRC5 cells, respectively). Docking and molecular dynamics simulations studies indicated that 8 efficaciously binds to the intended target hCoV-229E main protease (M(pro)). Moreover, due to the high similarity between hCoV-229E M(pro) and SARS-CoV-2 M(pro), we also performed the in silico analysis towards the second target, which showed results comparable to those obtained for hCoV-229E M(pro) and promising in terms of energy of binding and docking pose.