Abstract
TMPRSS2 is a membrane associated serine protease which is important in the viral pathogenesis of coronaviruses and influenza viruses. We developed mechanism-based covalent α-ketobenzothiazole (kbt) inhibitors using substrate specificity PS-SCL screening of TMPRSS2 as a rational guide for inhibitor design. Three distinct focused libraries of tetrapeptide kbts were synthesized and evaluated for their inhibition of TMPRSS2, matriptase and other serine proteases. We also investigated different capping groups for the previously reported tripeptide inhibitor Ac-QFR-kbt (18) to increase its selectivity over the blood coagulation protease factor Xa. The most potent compounds were tested for their ability to inhibit viral replication of SARS-CoV-2 coronavirus and the H1N1 influenza A virus. The most active compounds were profiled for their pharmacokinetics (PK) in mice. Several promising new compounds were identified with improved potency, selectivity, and drug-like properties including Bz-QFR-kbt (23) and Cbz-QFR-kbt (25) with an IC(50) of 150 nM and 60 nM for H1N1, respectively.