SARS-CoV-2 is the viral pathogen causing the COVID19 global pandemic. No effective treatment for COVID-19 has been established yet. TMPRSS2 is essential for viral spread and pathogenicity by facilitating the entry of SARS-CoV-2 onto host cells. The protease inhibitor camostat, an anticoagulant used in the clinic, has potential anti-inflammatory and anti-viral activities against COVID-19. However, the potential mechanisms of viral resistance and antiviral activity of camostat are unclear. Herein, we demonstrate high inhibitory potencies of camostat for a panel of serine proteases, indicating the camostat is a broad-spectrum inhibitor of serine proteases. In addition, we determined the crystal structure of camostat in complex with a serine protease (uPA), which reveals that camostat insert to the S1 pocket of uPA but was hydrolyzed by uPA, and the cleaved camostat covalently binds to the Ser195. We also generated the homology model of the structure of the TMPRSS2 serine protease domain. The model showed that camostat used the same inhibitory mechanism to inhibit the activity of TMPRSS2, and subsequently preventing SARS-CoV-2 spread. Importance section Serine proteases are a large family of enzymes critical for multiple physiological processes and proven diagnostic and therapeutic targets in several clinical indications. A serine protease transmembrane protease serine 2 (TMPRSS2) was recently found to mediate SARS-coronavirus 2 (SARS-CoV-2) entry into the host. camostat mesylate (FOY 305), a serine protease inhibitor active against TMPRSS2 and used for the treatment of oral squamous cell carcinoma and chronic pancreatitis, inhibits SARS-CoV-2 infection of human lung cells. However, the direct inhibition mechanism of camostat mesylate for TMPRSS2 is unclear. Herein, we demonstrate camostat used the same inhibitory mechanism to inhibit the activity of TMPRSS2 as uPA, and subsequently preventing SARS-CoV-2 spread.

The ongoing pandemic of coronavirus disease 2019 (COVID-19) posed a major challenge to the public health. Currently, no proven antiviral treatment for Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) infection is available. Here we report compounds pentalysine β-carbonylphthalocyanine zinc (ZnPc5K) and chlorin e6 (ce6) potently inhibited the viral infection and replication in vitro with EC 50 values at nanomolar level. These compounds were first identified by screening a panel of photosensitizers for photodynamic viral inactivation. Such viral inactivation strategy is implementable, and has unique advantages, including resistance to virus mutations, affordability compared to the monoclonal antibodies, and lack of long-term toxicity.

Receptor dimerization of urokinase-type plasminogen activator receptor (uPAR) was previously identified at protein level and on cell surface. Recently, a dimeric form of mouse uPAR isoform 2 was proposed to induce kidney disease. Here, we report the crystal structure of human uPAR dimer at 2.96 Å. The structure reveals enormous conformational changes of the dimer compared to the monomeric structure: D1 of uPAR opens up into a large expanded ring that captures a β-hairpin loop of a neighboring uPAR to form an expanded β-sheet, leading to an elongated, highly intertwined dimeric uPAR. Based on the structure, we identify E49P as a mutation promoting dimer formation. The mutation increases receptor binding to the amino terminal fragment of its primary ligand uPA, induces the receptor to distribute to the basal membrane, promotes cell proliferation, and alters cell morphology via β1 integrin signaling. These results reveal the structural basis for uPAR dimerization, its effect on cellular functions, and provide a basis to further study this multifunctional receptor.

The pandemic of coronavirus disease 2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has exploded since December 2019, and causes more than 2 million death with more than 95 million people infected as of Jan. 21th, 2021 globally1,2. Angiotensin-converting enzyme 2 (ACE2), expressed in the lungs, arteries, heart, kidney, intestines, and nasal epithelium3, has been shown to be the primary entry point targeted by the surface spike protein of SARS-CoV-2. Currently, no proven antiviral treatment for SARS-CoV-2 infection is available. In this study, we screened a number of photosensitizers for photodynamic viral inactivation, and found compounds pentalysine β-carbonylphthalocyanine zinc (ZnPc5K) and chlorin e6 (ce6) potently inhibited the viral infection and replication in vitro with half-maximal effective concentrations (EC50) values at nanomolar level. Such viral inactivation strategy is implementable, and has unique advantages, including resistance to virus mutations, affordability compared to the monoclonal antibodies, and lack of long-term toxicity.

Receptor dimerization of urokinase-type plasminogen activator receptor (uPAR) was previously identified at protein level and on the cell surface. Recently, a dimeric form of mouse uPAR isoform 2 was proposed, which induced kidney disease. Here, we report the crystal structure of human uPAR dimer at 2.96 Å. The structure reveals enormous conformational changes of the dimer compared to the monomeric structure: D1 of uPAR opens up into a large expanded loop that captures a β-hairpin loop of a neighboring uPAR to form an expanded β-sheet, leading to an elongated, highly intertwined dimeric uPAR. Based on the structure, we identify the E49P mutation promoting dimer formation. The mutation increases receptor binding to amino terminal fragment (ATF) of its primary ligand uPA, induces the receptor to distribute to the basal membrane, promotes cell proliferation, and alters cell morphology via the ERK activation of β1 integrin signaling. These results reveal the structural basis for uPAR dimerization, its effect on cell function, and provide new insight and tools to study this multifunctional receptor.