Table 3.

Investigating selected reported small‐molecule cysteine protease inhibitors for SARS‐CoV‐2 PL^pro inhibition using SPE‐MS inhibition assays.

	Cysteine protease inhibitor	IC50 [μM][a]		Cysteine protease inhibitor	IC50 [μM][a]
1		0.3±0.1	3		>50
2		>20[b]	4		>50

[a] PL^pro SPE‐MS inhibition assays were performed using SPE‐MS as described in the Supporting Information (Section 5) employing 0.2 μM SARS‐CoV‐2 PL^pro and 2.0 μM of the nsp2/3 peptide 2 (VTNNTFTLKGG/APTKVTFGD) in the presence of N‐terminally acetylated product peptides (0.2 μM Ac‐VTNNTFTLKGG and 0.2 μM Ac‐APTKVTFGD). Inhibitors were commercially‐sourced and used as received, PF‐07321332 ^[2f] and SDZ‐224015 ^[8b] were synthesized as described. Results are means of two independent runs (n=2; mean±SD); [b] no PL^pro inhibition was observed at 20 μM PF‐07321332.