Discovery and Crystallographic Studies of Trisubstituted Piperazine Derivatives as Non-Covalent SARS-CoV-2 Main Protease Inhibitors with High Target Specificity and Low Toxicity.
Gao S, Sylvester K, Song L, Claff T, Jing L, Woodson M, Weiße RH, Cheng Y, Schäkel L, Petry M, Gütschow M, Schiedel AC, Sträter N, Kang D, Xu S, Toth K, Tavis J, Tollefson AE, Müller CE, Liu X, Zhan P.
The continuous spread of SARS-CoV-2 calls for more direct-acting antiviral agents to combat the highly infectious variants. The main protease (M<sup>pro</sup>) is an promising target for anti-SARS-CoV-2 drug design. Here, we report the discovery of potent non-covalent non-peptide M<sup>pro</sup> inhibitors featuring a 1,2,4-trisubstituted piperazine scaffold. We systematically modified the non-covalent hit MCULE-5948770040 by structure-based rational design combined with multi-site binding and privileged structure assembly strategies. The optimized compound <b>GC-14</b> inhibits M<sup>pro</sup> with high potency (IC<sub>50</sub> = 0.40 μM) and displays excellent antiviral activity (EC<sub>50</sub> = 1.1 μM), being more potent than Remdesivir. Notably, <b>GC-14</b> exhibits low cytotoxicity (CC<sub>50</sub> > 100 μM) and excellent target selectivity for SARS-CoV-2 M<sup>pro</sup> (IC<sub>50</sub> > 50 μM for cathepsins B, F, K, L, and caspase 3). X-ray co-crystal structures prove that the inhibitors occupy multiple subpockets by critical non-covalent interactions. These studies may provide a basis for developing a more efficient and safer therapy for COVID-19.