Alpha-ketoamides as broad-spectrum inhibitors of coronavirus and enterovirus replication

Linlin Zhang; Daizong Lin; Yuri Kusov; Yong Nian; Qingjun Ma; Jiang Wang; Albrecht von Brunn; Pieter Leyssen; Kristina Lanko; Johan Neyts; Adriaan de Wilde; Eric J. Snijder; Hong Liu; Rolf Hilgenfeld

The copyright holder for this preprint (which was not peer-reviewed) is the . https://doi.org/10.1101/2020.02.10.936898 doi: bioRxiv preprint ment of the phenyl group by the cyclohexyl group led to a significant improvement of the inhibitory activity against the recombinant SARS-CoV M pro and to a dramatic improvement in case of CVB3 3C pro . Even for the HCoV-NL63 M pro , against which 11a was completely inactive, greatly improved albeit still weak activity was observed (Table 1 ). In the viral replicons, 11r performed very well, with EC50 = 0.8 -0.9 µM for the EV-A71 replicon, 0.45 µM for CVB3, and 1.4 µM for SARS-CoV (Table 2 ). In the virus-infected cell culture assays (Table 3) , 11r exhibited EC50 = 3.7 µM against EV-A71 in RD cells and EC50 = 0.48 -0.7 µM against EV-D68, HRV2, and HRV14 in HeLa cells. Against HCoV 229E in Huh7 cells, the EC50 was surprisingly low (1.8 µM). Interestingly, the compound proved extremely potent against MERS-CoV in Huh7 cells, with EC50 = 0.0004 µM (400 picomolar!). Even in Vero cells, EC50 against MERS-CoV was 5 µM, and the EC50 against SARS-CoV in Vero E6 cells was 1.8 -2.1 µM, i.e. the best activity we have seen for an M pro inhibitor against SARS-CoV in this type of cells. The therapeutic index (CC50/EC50) of 11r against EV-D68, HRV2, and HRV14 was >15 in HeLa Rh cells as well as against CVB3 in Huh-T7 cells, but only ~5 for EV-A71 in RD cells.