Improving the selectivity of 3 amidinophenylalanine derived matriptase inhibitors

A rational structure based approach was employed to develop novel 3 amidinophenylalanine derived matriptase inhibitors with improved selectivity against thrombin and factor Xa. Of all 23 new derivatives, several monobasic inhibitors exhibit high matriptase affinities and strong selectivity against thrombin. Some inhibitors also possess selectivity against factor Xa, although less pronounced as found for thrombin. A crystal structure of a selective monobasic matriptase inhibitor in complex with matriptase and three crystal structures of related compounds in trypsin and thrombin have been determined. The structures offer an explanation for the different selectivity profiles of these inhibitors and contribute to a more detailed understanding of the observed structure activity relationship. Selected compounds were tested in vitro against a matriptase dependent H9N2 influenza virus strain and demonstrated a concentration dependent inhibition of virus replication in MDCK II cell

Cleavage and Activation of the Severe Acute Respiratory Syndrome Coronavirus Spike Protein by Human Airway Trypsin-Like Protease ▿

The highly pathogenic severe acute respiratory syndrome coronavirus (SARS-CoV) poses a constant threat to human health. The viral spike protein (SARS-S) mediates host cell entry and is a potential target for antiviral intervention. Activation of SARS-S by host cell proteases is essential for SARS-CoV infectivity but remains incompletely understood. Here, we analyzed the role of the type II transmembrane serine proteases (TTSPs) human airway trypsin-like protease (HAT) and transmembrane protease, serine 2 (TMPRSS2), in SARS-S activation. We found that HAT activates SARS-S in the context of surrogate systems and authentic SARS-CoV infection and is coexpressed with the viral receptor angiotensin-converting enzyme 2 (ACE2) in bronchial epithelial cells and pneumocytes. HAT cleaved SARS-S at R667, as determined by mutagenesis and mass spectrometry, and activated SARS-S for cell-cell fusion in cis and trans, while the related pulmonary protease TMPRSS2 cleaved SARS-S at multiple sites and activated SARS-S only in trans. However, TMPRSS2 but not HAT expression rendered SARS-S-driven virus-cell fusion independent of cathepsin activity, indicating that HAT and TMPRSS2 activate SARS-S differentially. Collectively, our results show that HAT cleaves and activates SARS-S and might support viral spread in patients