Link to bioRxiv paper: http://biorxiv.org/cgi/content/short/2020.10.23.347534v1?rss=1 Authors: Mellott, D., Tseng, C.-T., Drelich, A., Fajtova, P., Chenna, B. C., Kostomiris, D., Hsu, J. C., Zhu, J., Taylor, Z., Tat, V., Katzfuss, A., Li, L., Giardini, M. A., Skinner, D., Hirata, K., Beck, S., Carlin, A. F., Clark, A. E., Berreta, L., Maneval, D., Frueh, F., Hurst, B. L., Wang, H., Kocurek, K. I., Raushel, F. M., O'Donoghue, A., Siqueira-Neto, J. L., Meek, T. D., McKerrow, J. H. Abstract: K777 is a di-peptide analog that contains an electrophilic vinyl-sulfone moiety and is a potent, covalent inactivator of cathepsins. Vero E6, HeLa/ACE2, Caco-2, A549/ACE2, and Calu-3, cells were exposed to SARS-CoV-2, and then treated with K777. K777 reduced viral infectivity with EC50 values of inhibition of viral infection of: 74 nM for Vero E6, <80 nM for A549/ACE2, and 4 nM for HeLa/ACE2 cells. In contrast, Calu-3 and Caco-2 cells had EC50 values in the low micromolar range. No toxicity of K777 was observed for any of the host cells at 10-100 M inhibitor. K777 did not inhibit activity of the papain-like cysteine protease and 3CL cysteine protease, encoded by SARS-CoV-2 at concentrations of [≤] 100 M. These results suggested that K777 exerts its potent anti-viral activity by inactivation of mammalian cysteine proteases which are essential to viral infectivity. Using a propargyl derivative of K777 as an activity-based probe, K777 selectively targeted cathepsin B and cathepsin L in Vero E6 cells. However only cathepsin L cleaved the SARS-CoV-2 spike protein and K777 blocked this proteolysis. The site of spike protein cleavage by cathepsin L was in the S1 domain of SARS-CoV-2 , differing from the cleavage site observed in the SARS CoV-1 spike protein. These data support the hypothesis that the antiviral activity of K777 is mediated through inhibition of the activity of host cathepsin L and subsequent loss of viral spike protein processing. Copy rights belong to original authors. Visit the link for more info